2024-03-28T17:00:36Z
http://surface.syr.edu/do/oai/
oai:surface.syr.edu:bio_etd-1002
2010-09-01T15:27:46Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The role of steroids in the regulation of oocyte cyst breakdown and primordial follicle assembly in the neonatal mouse ovary
Chen, Ying
2008-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Melissa E. Pepling
Steroids
Oocyte cyst breakdown
Primordial follicle assembly
Ovary
Biology
Mouse germ cells develop in special structures called germline cysts formed by several rounds of mitosis and incomplete cytokinesis. Right after birth, cysts break down and each individual germ cell becomes enclosed in a primordial follicle, which represents the total germ cell population a female has throughout her reproductive life. At the same time, about two thirds of the germ cells die with only one third surviving. The mechanisms regulating cyst breakdown and selective oocyte loss are not completely understood. A more detailed understanding of these processes will give us more insights into normal early oocyte development and female fertility problems.
In this study, utilizing the ovary organ culture system, the role of hormone signaling on cyst breakdown is investigated. Estrogen, progesterone, as well as phytoestrogen genistein all displayed an inhibitory effect on cyst breakdown, causing large cysts to persist during culture. Primordial follicle assembly is also inhibited. Ovaries cultured in vitro cut off from their maternal estrogen supply from the mother, undergo premature cyst breakdown, oocyte death and follicle development. Adding estradiol or progesterone back to the culture media rescues this premature development. Using estrogen receptor specific agonists and antagonists, we established that both estrogen receptor a and estrogen receptor β, as well as a membrane estrogen receptor, are involved in estrogen signaling in the neonatal ovary. ICI182,780, a pure antiestrogen, can reverse the effect of estradiol on cyst breakdown. Our current working model is that before birth, the fetus is exposed to estrogen from the mother, and the relatively high estrogen level inhibits the cysts from breaking down; Right after birth, estrogen levels drop dramatically, and this triggers some oocytes to die and the cysts to break down into individual oocytes.
https://surface.syr.edu/bio_etd/4
oai:surface.syr.edu:bio_etd-1004
2015-01-12T15:51:20Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The voltage dependent anion channel affects mitochondrial cholesterol distribution and function
Campbell, Andrew M.
2007-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Samuel H. P. Chan
Mitochondria
Warburg effect
Voltage dependent anion channel
Cholesterol
Biology
<p>Normal cells of aerobic organisms synthesize the energy they require in the form of ATP via the process of oxidative phosphorylation. This complex system resides in the mitochondria of cells and utilizes oxidizable substrates to fuel the conversion of ADP to ATP and is dependent on molecular oxygen as the final electron acceptor. In humans, 90% of cellular energy is produced by this process. However, in most tumors, especially those with elevated cholesterogenesis, there is an increased reliance on glycolysis for energy, even in conditions where oxygen is available. This aerobic glycolysis (the Warburg effect) has far reaching ramifications on the tumor itself and the cells that surround it. Here, we have observed abnormally high membrane cholesterol levels and a subsequent deficiency of oxidative energy production in mitochondria from cultured Morris hepatoma cells (MH-7777). Using cholesterol affinity chromatography and MALDI-TOF MS, we have identified the voltage dependent anion channel (VDAC) as a necessary component of a protein complex involved in mitochondrial membrane cholesterol distribution and transport. VDAC is known to associate strongly with hexokinase, particularly in glycolytic cancers. By constructing an E72Q mutant form of VDAC that inhibits its binding of hexokinase, we report an increase in OXPHOS activity of MH7777 cells, as well as reduced membrane cholesterol ratios to levels near that of normal liver mitochondria. Work in our laboratory demonstrates that the ability of VDAC to influence mitochondrial membrane cholesterol distribution may have implications on mitochondrial characteristics such as oxidative phosphorylation and induction of apoptosis, as well as the propensity of cancer cells to exhibit a glycolytic phenotype.</p>
https://surface.syr.edu/bio_etd/2
oai:surface.syr.edu:bio_etd-1003
2012-12-12T14:38:13Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Study of hydrogen bonding properties with ab initio calculations, neutron scattering spectroscopy and 1H NMR
Lan, Yanmei
2007-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Bruce S. Hudson
Hydrogen bonding
Ab initio calculation
1H NMR
Neutron scattering spectroscopy
Aggregation
Cooperative hydrogen bonding
Biology
<p>The H-bond is known to be very importance in solid aggregation, molecular biology, and medicine. It is also very important in proteins and nucleic acids and therefore in life-sustaining processes. The understanding and control of non-covalent interactions, such as H-bonding, will be very helpful to the assembly of functional nano-systems with a precision analogous to that found in the natural world. This thesis is to investigate H-bond properties of small organic molecules with neutron scattering spectroscopy (INS), ab initio calculations, and 1 H NMR. I first show the results of the theoretical and experimental study of INS vibrational spectra of a moderate H-bonding system. The calculation methods are correctly used to simulate the vibrational spectra of the system. I also show a detailed study on 1,3-cyclohexanedione (CHD) and its methylated analogues that exhibit cooperative, resonance assisted H-bonding (RAHB) in solid state. INS simulations with hartree fork (HF) and density functional theory (DFT) are carried out for non-deuterated and deuterated species, with results in good agreement with experiment. 1 H NMR experimental methods are used in connection with HF and DFT calculations. The results of gas phase calculations on theoretical energy of a monomer, dimer, and trimer of CHD and its analogues indicate that cooperative behavior is expected with aggregation. Evidence from the comparison of crystal structure with the structure calculated for several aggregates also suggests a basis for such cooperative behavior. Measures of the association equilibria of CHD, however, provide no evidence for cooperativity due to a large effect of the CDCl 3 solvent. Calculations of CHD in CHCl 3 medium using Onsager Spherical Model indicate that the polar solvent has a big effect on the H-bonding equilibrium of CHD. The cooperative energy on going from dimer to trimer is compensated in the free energy by a change in the enthalpy and entropy so the cooperative effect is not reflected in the equilibrium constants. This thesis first solved the problem of the keto-enol equilibrium of CHD and its aggregations in solution. This is a new method that combines both experiment and theory to examine the hydrogen bonding properties of CHD in solution.</p>
https://surface.syr.edu/bio_etd/3
oai:surface.syr.edu:bio_etd-1006
2010-09-14T17:30:39Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Ecology of the Trans-Himalayan grazing ecosystem
Bagchi, Sumanta
2009-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Mark E. Ritchie
Grazing ecosystem
Herbivory
Climate change
Biodiversity
Soil quality
Ecology and Evolutionary Biology
Life Sciences
This study investigates ecosystem function in the Trans-Himalayan grazing ecosystem in northern India. First, herbivory-tolerance in plants was studied under a full-factorial arrangement of experimental clipping, irrigation and fertilization. In absence of fertilization, plants compensated for defoliation if not irrigated, but failed to compensate under irrigation. When fertilized, plants compensated for defoliation only under irrigation. These results indicate that grazers can alter the strength of resource co-limitation faced by plants. Second, using herbivore-exclusion experiments, direct and indirect impacts of grazers on ecosystem function were measured. Results show that grazing promotes aboveground production but reduces it belowground. Grazing also reduces C:N ratio in plant tissue and litter, which coincides with increased plant-available inorganic N in soil. These results indicate that herbivores can have positive effects on ANPP if they accelerate nutrient cycling. Third, simultaneous grazer control on plant production and community composition were estimated. Results indicated that after accounting for influence of co-varying variables, these two aspects are not related. Fourth, differences between effects of native and introduced grazers on ecosystem function was estimated. Results indicate that introduced livestock grazers reduce wholeecosystem productivity and lower the potential for C-input in soil. This reduces potential soil-C sequestration.
https://surface.syr.edu/bio_etd/8
oai:surface.syr.edu:bio_etd-1007
2010-09-14T18:37:01Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Identification and characterization of proteins involved in Myxococcus xanthus sporulation
Tengra, Farah Khurshed
2009-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Anthony Garza
Sporulation
Myxococcus xanthus
Stress resistance
Life Sciences
Microbiology
Myxococcus xanthus is a Gram-negative, rod-shaped soil dweller that undergoes a complex developmental cycle in response to nutrient starvation. A complex multi-cellular structure, referred to as a fruiting body, is formed following the aggregation of approximately 10 5 cells under starvation conditions. The rod-shaped cells within this structure differentiate into stress-resistant, dormant spores that are relatively resistant to dessication, UV irradiation, toxic molecules, and extreme temperatures. Myxococcus xanthus ( M. xanthus ) spores are composed of a core, cortex, and coat, but the proteins associated with the layers of the spore are not well studied. Two approaches were taken to identify proteins involved in sporulation of M. xanthus. We looked for genes with sequence similarity to the well studied sporulation model, Bacillus subtilis. We also took a proteomic approach to identify spore related proteins in M. xanthus.
CbgA was first identified as a potential M. xanthus sporulation protein based upon its sequence similarity to the Bacillus subtilis sporulation protein SpoVR. SpoVR is known to play an important role in the formation the endospore cortex, a protective layer of peptidoglycan. Here we show that the cbgA mutant aggregates and forms fruiting bodies, although cbgA fruiting bodies have abnormal shapes. Transmission electron microscopy revealed that cbgA mutant spores lack cortexes or have relatively thin cortex layers. Given that cbgA spores are spherical like their wild-type counterparts, this finding suggests that the cortex is not essential for M. xanthus spores to maintain their characteristic shape. Heat sensitivity is a property associated with spore cortex defects, and we found that spores produced by the cbgA mutant are more sensitive to heat than wild-type spores. We also found that cbgA spores are more sensitive to sodium dodecyl sulfate (SDS) than wild-type spores. However, cbgA mutant spores show no loss in sonication, UV irradiation or lysozyme resistance. These results indicate that CbgA plays an important role in spore cortex formation and the acquisition of a subset of spore stress-resistance properties.
The proteome of liquid-grown vegetative cells was compared with the proteome of mature fruiting body spores to look for proteins involved in forming resistant fruiting body spores. Two proteins, protein S and protein S1, were differentially expressed in spores as has been previously reported. In addition, we identified three previously uncharacterized proteins differentially expressed in spores. The genes for the three novel m ajor s pore p roteins ( mspA, mspB, and mspC ), were inactivated by insertion mutagenesis, and the resulting mutant strains were characterized for development. All three mutants formed aggregates, but the aggregates of two of the strains the fruiting bodies remained as flattened mounds of cells instead of taking on the characteristic dome shape of wild-type fruiting bodies. The most pronounced structural defect of spores produced by all three of these mutants was an altered cortex layer. mspA and mspB mutant spores are more sensitive specifically to heat and sodium dodecyl sulfate than wild-type spores, while mspC mutant spores are more sensitive to all stress treatments tested. Hence, the products of mspA, mspB, and mspC play significant roles in morphogenesis of M. xanthus spores and in the ability of spores to survive stress.
https://surface.syr.edu/bio_etd/7
oai:surface.syr.edu:bio_etd-1008
2010-09-16T17:56:53Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Plumage evolution in bearded manakins (Manacus spps.)
Stein, Adam Carsten
2009-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
J. Albert C. Uy
Plumage
Bearded manakins
Manacus
Animal Sciences
Ecology and Evolutionary Biology
Evolution
Life Sciences
Zoology
The evolution of bright, colorful and elaborate male secondary sexual characteristics is now widely accepted to be the product of sexual selection by female-choice and/or male-male competition. More recently, sexual selection by female choice has also been argued to play an important role in the speciation process. In particular, it has been argued that when a female preference for a particular sexual trait diverges and there is a corresponding divergence in that male trait, it could inhibit different, but essentially genetically similar, populations from interbreeding. Many studies have now begun to elucidate the role of divergent sexual selection in reproductive isolation, but few have focused on the underlying mechanisms that drive the change in female preference. We studied a unique avian hybrid zone between Manacus candei and M. vitellinus, where the bright white secondary male plumage of Manacus candei is evolving into bright yellow plumage through the introgression of plumage traits from Manacus vitellinus. We attempt to discover what mechanisms may be responsible for causing sexual signals to diverge. To test whether male secondary sexual color was an important corollary to mating success, we monitored a group of male Manacus vitellinus throughout the breeding season and related the number of females they copulated with (mating success) to their respective plumage traits. We then focused on a population of males where the two color forms came together at the hybrid zone. This population contained both yellow and white males that competed for the same females. We monitored selected pairs of males that consisted of one naturally white male and one naturally yellow male to see if there was any difference between color types in mating success that may explain the observed introgression of yellow plumage traits. In attempt to distinguish between female choice and male male-competition for any observed difference in mating success between the two color forms, we measured other physical and behavioral attributes considered important to male-male competition between naturally yellow and naturally white males. Further, we monitored both a population of male Manacus candei and a population of male Manacus vitellinus during the height of their breeding season to determine the mating success of each individual. We then modeled how females perceive their specific plumage traits in their particular environment to determine if females were simply choosing the males with the most conspicuous plumage traits. Lastly, we examined if this sexually-selected color trait could serve another additional function outside of attracting females by performing a plumage manipulation experiment in the population containing both yellow and white individuals.
We found that aspects of color were indeed strong predictors of mating success in this species complex and that the introgression of novel yellow plumage traits into Manacus candei as a result of this hybridization was most likely the result of sexual selection via female choice. Females, however, did not choose the most conspicuous males in leks outside of the hybrid zone, leading us to conclude that female preference for particular colors may be more complex than just favoring the most effective signal. Results of the plumage manipulation experiment demonstrated that males were also using color, most likely to discriminate between familiar and unfamiliar males.
https://surface.syr.edu/bio_etd/9
oai:surface.syr.edu:bio_etd-1009
2010-09-20T14:29:18Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Agglutination-dependent and -independent adhesin interactions during mating in Saccharomyces cerevisiae
Huang, Guohong
2006-06-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Scott E. Erdman
Fungal adhesins
Mating
Saccharomyces cerevisiae
Posttranslational modification
WCPL/CX4C domains
GPI
Glycophosphatidylinositol
Agglutination
Microbiology
Molecular Biology
Fungal adhesins are cell surface proteins that mediate adherence of fungal cells to host substrates and each other, which affects their access to nutrients, sexual conjugation, and survival in hosts. Most fungal adhesins receive two posttranslational modifications, glycophosphatidylinositol (GPI) anchor addition and O-glycosylation, and a subset of fungal adhesins contains highly conserved WCPL and CX 4 C domains of unknown function. This study was directed toward understanding the in vivo functional requirements of the posttranslational modifications and the WCPL/CX 4 C domains for fungal adhesins, using the mating system in Saccharomyces cerevisiae as a model.
Deletion of the signal sequence for GPI addition of the yeast a -agglutinin subunit, Aga1p, eliminated its activity, while deletions of different internal domains had varying effects on function. Substitution of the Aga1p GPI signal domain with those of other GPI-anchored proteins, a single transmembrane domain, or a cysteine capable of forming a disulfide all produced functional adhesins. Aga1p and the α-agglutinin Agα1p were shown to be under glycosylated in cells lacking the protein mannosyltransferase genes PMT1 and PMT2 , with phenotypes manifested only in MATα cells for single mutants but in both cell types when both genes are absent.
Besides the well-known agglutinin interaction between the a -agglutinin complex (Aga1p-Aga2p) and Agα1p, three more pairs of interactions were revealed by this study, including bilateral heterotypic interactions between Aga1p and Fig2p, and a weaker homotypic interaction between Fig2p and Fig2p, in cells of the opposite mating type. These four pairs of adhesin interactions are collectively required for maximum mating efficiency and normal zygote morphogenesis. GPI-less, epitope-tagged forms of Aga1p and Fig2p can be co-immunoprecipitated from the culture medium of mating cells in a manner dependent on the WCPL/CX 4 C domains in Aga1p. Using site-directed mutagenesis, the conserved residues in Aga1p that interact with Fig2p were identified.
To conclude, posttranslational modifications and the conserved WCPL/CX 4 C domains are necessary for the biogenesis and activity of Aga1p. Aga1p is involved in two distinct adhesive functions that are independent of each other, which raises the possibility for combinatorial interactions of this protein with its different adhesion receptors, Aga2p and Fig2p, a property previously unknown for fungal adhesins.
https://surface.syr.edu/bio_etd/10
oai:surface.syr.edu:bio_etd-1010
2010-09-20T16:39:51Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Exploring transcriptional regulators that control emergent behaviors in Myxococcus xanthus
Taylor, Rion G.
2008-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Myxococcus xanthus
Chemotaxis
Development
Self-organization
Emergence
Life Sciences
Microbiology
A swarm of the δ-proteobacterium Myxococcus xanthus is a distributed system; a population of superposable automata whose distribution is transparent so that it appears as one machine. These swarms contain millions of cells that act as a collective, exhibiting coordinated movement through a series of signals to create complex, dynamic patterns as a response to environmental cues. These self-organizing patterns are considered emergent, as they cannot be predicted by observing the behavior of the individual cells.
Self-organization in M. xanthus is ultimately controlled through gene expression via transcriptional regulators (TRs). We selected to examine the effect of TRs on development, an example of a distinct emergent pattern, and designed new assays to quantify this behavior. We measured the swarm's ability to develop using TR mutant strains and found that mutants previously characterized as producing no disparate phenotype actually did.
We identified, characterized, and quantified a second distinct emergent pattern in M. xanthus called chemotaxis. We define chemotaxis as the directed movement of a swarm up a nutrient gradient toward its source. This behavior can only be accomplished when the cells are in a cooperative, multicellular swarm. We have also demonstrated that chemotaxis is under transcriptional control by several members of the highly duplicated NtrC-like family of TRs. The combination of experimental and genetic evidence suggests that chemotaxis evolved after the establishment of the swarm by means of duplication and divergence of multiple signaling pathways.
https://surface.syr.edu/bio_etd/12
oai:surface.syr.edu:bio_etd-1011
2010-09-20T18:55:31Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Resource allocation to growth and thermoregulation during early development in altricial nestlings
Kovatch, Jeffrey J.
2008-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Altricial birds
Brooding
Resource allocation
Thermoregulation
Nestlings
Ecology and Evolutionary Biology
Life Sciences
Physiology
Patterns of limited resource allocation to the mutually exclusive processes of growth and maintenance are products of natural selection and presumably confer fitness benefits. Hypothesized universal scaling of metabolic rate, growth, and resource allocation may not apply for altricial birds. Altricial birds grow rapidly by delaying thermal maintenance costs and preferentially allocating energy to growth. Endothermic nestling thermoregulation is aided by intermittent parental brooding. A general growth model was used as a null-model to test if deviations of altricial growth from universal patterns suggested energetic and fitness advantages. Differences between altricial and universal growth were not explained by changes in proportional water content of tissue or observed systematic increases in body temperature during ontogeny. Nestling body temperatures are maintained below 37ºC for the first few days. Mass- and temperature-normalization of nestling metabolic rates suggest the rapid development of metabolic capacity may explain observed growth patterns, and thus deviations from universal patterns. The hypothesis that the altricial thermoregulatory strategy has a cost savings relative to the endothermic-at-hatch (precocial) alternative was tested for young house wren ( Troglodytes aedon ) nestlings. Nestling and parental thermoregulatory contributions were estimated from rates of wren broods cooling and heating under natural conditions, and costs for brood-level endothermy were modeled. Intermittent brooding was energetically cheaper than endothermy, ∼250 kJ savings or ∼168% over the first 5 days of development at moderate environmental temperatures, ∼21ºC. Relative cost savings for brooding increased with lower environmental temperatures. Projected metabolic capacities necessary for endothermy are unrealistically high, up to 14-fold, for young nestlings in broods. The shift in the brooding strategy cost effectiveness coincides with the age when relative tissue water decreases and growth rate is maximum, suggesting a shift in preferential resource allocation from growth to maintenance. The hypothesis of temperature dependent growth was tested in wrens by experimentally increasing body temperatures ∼2ºC for the first 7 days. No change in growth rate was observed suggesting that nestling metabolic and growth rates may be maximized. Because there is no growth rate advantage with higher temperatures, maintaining lower body temperatures for young nestlings is another cost savings of the altricial strategy.
https://surface.syr.edu/bio_etd/11
oai:surface.syr.edu:bio_etd-1013
2010-09-21T17:59:38Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Understanding prokaryotic diversity in the post-genomics era
Suen, Garret
2008-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Functional genomics
Protein domains
Gammaproteobacteria
Bioinformatics
Life Sciences
Microbiology
Prokaryotes, one of nature's most pervasive organisms, are ubiquitous in the environment and impact human society, both as harmful and beneficial agents. A challenge of microbial research is to understand prokaryotes within the context of their environment. Currently, assessing prokaryotic diversity is difficult because we cannot culture the majority of microbes from the environment. Therefore, new techniques must be developed to not only assess this diversity, but to understand how such diversity is linked to genes in a prokaryote's genome. Access to the genome sequences of a large number of prokaryotes has radically changed microbial research, and the expected availability of thousands of genomes in the future has transitioned the field into the post-genomics era. This thesis presents the development and analysis of two post-genomics tools. The first is in functional genomics, which seeks to determine the genetic interactions that underlie prokaryotic traits. Phylogenomics predicts genetic interactions based on the incidence of coinherited proteins across evolutionary phyla, and assigns function to a gene. This was experimentally tested for the model organism Myxococcus xanthus and further applied to the genome annotation of Sorangium cellulosum . Additional analysis was performed in the area of integrative functional genomics, where it is thought that the combination of multiple functional genomics approaches improves the accuracy of individual predictions. This assumption was tested by combining the predictions from phylogenomics and gene expression mapping for four prokaryotes. It was found that, while this assumption holds, the rate that predictions converge upon a stable set of interactions is significantly slower than previously thought. The second tool relates a prokaryote's genome to its niche. Prokaryotes with similar genetic content, as measured by protein domains, were clustered into mountains on a niche map. When compared to a second map based on 16S rRNA, the niche map better clustered prokaryotes according to an environment/function metric. Analysis of mountains on the niche map showed correlation to niche, including the clustering of marine Gammaproteobacteria, obligate pathogens and symbionts, and prokaryotes that exist at the soil, plant, and human interface. Taken together, both tools can help us better understand prokaryotic diversity in the post-genomics era.
https://surface.syr.edu/bio_etd/13
oai:surface.syr.edu:bio_etd-1014
2010-09-22T12:59:13Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Direct and indirect ecological effects of Dreissenid mussels (the zebra mussel Dreissena polymorpha and the quagga mussel D. bugensis) on submerged macrophytes in North American lakes
Zhu, Bin
2006-06-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
C. M. Mayer
Mark E. Ritchie
Dreissenid mussels
Submerged macrophytes
Great Lakes
Hydroacoustics
Terrestrial and Aquatic Ecology
Dreissenid mussels (the zebra mussel, Dreissena polymorpha and the quagga mussel, D. bugensis ) are ecosystem engineers that increase water clarity and contribute to oligotrophication and benthification in North American lakes. Several researchers have observed changes in submerged macrophyte communities associated with the dreissenid mussel invasion, and attributed these changes to increased water clarity. In contrast, phosphorus reduction following the implementation of the Great Lakes Water Quality Agreement in 1972 appears not to have had significant impacts on macrophytes. I conducted both observational and experimental studies to investigate both direct and indirect ecological effects of dreissenid mussels on submerged macrophytes in North American lakes. The thesis consists of four chapters: (1) an observational study of the indirect impact of dreissenid mussels on submerged macrophytes through mediating water clarity in Oneida Lake, NY, (2) an observational study of the indirect impact in two bays of Lake Ontario, (3) an experimental study of the indirect impact in a pond, and (4) an observational and experimental study of direct light and nutrient effects of dreissenid mussels on submerged macrophyte growth.
Results from the first three chapters showed water clarity increased in lakes after the dreissenid invasion and macrophyte species diversity, their frequency of occurrence, the maximum depth of colonization, and their coverage increased too. In addition, macrophyte species composition changed from low-light-tolerant species to those tolerating a wide range of light conditions. In the fourth chapter, the study of direct impacts of dreissenid mussels on submerged macrophytes revealed species-specific impacts of the shell attachment and no effect of nutrient relocation. However, the nutrient impacts in lakes need further investigations.
Together, these results showed light is the overriding factor that determines the distribution and diversity of submerged macrophytes and the indirect effect of dreissenid mussels by mediating water clarity and light penetration is the most important. Other direct and indirect effects of dreissenid mussels are probably not important although more investigations may be needed. Therefore the consequence of dreissenid mussels for ecosystems is indeed indirect and appropriately called ecosystem engineering, which contributes to benthification in North American lakes.
https://surface.syr.edu/bio_etd/16
oai:surface.syr.edu:bio_etd-1015
2010-09-22T13:45:00Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
DNA helix imperfections: Structure and flexibility
McGee, Christopher J.
2006-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Philip N. Borer
DNA helix
Flexibility
Hairpin loop
Biochemistry, Biophysics, and Structural Biology
This dissertation reports on the NMR solution structure of a unique DNA 22-mer hairpin. The secondary structure of this hairpin has a four-base thymine loop, a 3'-dangling adenosine, and a one-base thymine bulge in the center of the stem flanked by four base pairs. Unique structures and mutations in DNA continue to be a research topic of great interest. Models such as this one may be of use in the study of DNA repair where it has been suggested that the structure, flexibility and dynamics of DNA might be responsible for activating the appropriate repair system. This oligomer has been synthesized by automated solid phase synthesis and NMR samples have been prepared at different concentrations. These samples have been used to acquire various NMR spectra with a 500 MHz spectrometer in both water and deuterium oxide solutions. Resonance assignments of NOESY, 13 C- 1 H HSQC, 1 H- 1 H COSY, 1 H- 31 P COSY and TOCSY spectra have been made to facilitate the computation of a DNA solution structure. Distances, derived from the cross peak intensities in various NOESY experiments, as well as torsion angle ranges were combined with the calculations from both the DYANA and AMBER protocols, to yield the three dimensional structure. Furthermore, in an effort to expand our understanding of the dynamical properties of DNA, additional 13 C- 1 H NMR measurements have been made. These include the steady state NOE, the spin-lattice relaxation and the spin-spin relaxation rates. Correlation times and order parameters were also calculated and presented based upon a "model-free" analysis. Our results indicate that the structure of this nucleic acid is consistent with a B-form DNA double helical stem which incorporates the one-base bulge in a well restrained structure. The tetraloop is less refined than the stem, indicating flexibility at the 3'-end of the loop. The relaxation data gathered further supports these conclusions.
https://surface.syr.edu/bio_etd/15
oai:surface.syr.edu:bio_etd-1016
2010-09-23T16:46:10Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Effects of light, nutrients and Dreissena (Dreissena polymorpha and Dreissena bugensis) on benthic ecosystems in lakes
Qin, Peibing
2007-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Nutrients
Lakes
Benthic ecosystems
Dreissena
Light penetration
Periphytons
Ecology and Evolutionary Biology
Life Sciences
Terrestrial and Aquatic Ecology
Filtration by invasive mollusks ( Dreissena spp. ) and nutrient reductions (e.g., phosphorus abatement) have dramatically increased water clarity and successfully lowered phosphorus inputs in the Great Lakes in recent decades. Field studies have shown that high Dreissena filtration greatly decreased phytoplankton biomass. However, we conducted field, mesocosm and lab experiments to examine light, Dreissena and nutrient effects on benthic algal biomass, production, and stoichiometry and benthic invertebrate biomass. This thesis consists of four chapters: (1) a series of field experiments to test effects of light and nutrients on periphyton food quantity and quality in lakes, (2) a mesocosm experiment to separate synergistic effects of Dreissena filtration and phosphorus abatement on benthic primary producers, (3) a mesocosm experiment to compare effects on benthic ecosystem of Dreissena to those of native grazers, and (4) three lab and field light experiments to test if single and natural periphyton stoichiometry response to light same as its production response to light.
Our field experiments showed that both light and nutrients affected periphyton biomass and stoichiometry. Periphyton biomass increased and periphyton C:P decreased as total phosphorus increased under high light. Periphyton biomass and C:P were higher in light than shade. Benthic invertebrate biomass was higher with high periphyton biomass and low periphyton C:P. Three lab and field light experiments also indicated that periphyton C: nutrients increased under high light due to more carbon fixation.
The two mesocosm experiments suggest that Dreissena are ecosystem engineers that modify habitat in a unique way by increasing light, habitat complexity and available P. Dreissena had a strong positive effect on periphyton PP, and biomass of macrobenthos on hard substrate. Light and Dreissena increased benthic invertebrate biomass. P1 alone did not affect biomass of periphyton and macrobenthos in ourutrophic mesocosms. High light and low P increased periphyton C:P on rock, but Dreissena did not affect periphyton stoichiometry, because it increased both C and P content of periphyton.
Together, these results showed Dreissena locally increased periphyton and benthic invertebrate biomass on hard substrate and increased benthic production lake-wide by mediating water clarity and light penetration. Our experiments allow us to tease apart concurring environmental stressors that Dreissena but not P abatement increased periphyton and benthic invertebrate biomass. Additionally, light should be considered as an important factor affecting periphyton stoichiometry on hard substrate and macrobenthos biomass lake-wide.
https://surface.syr.edu/bio_etd/19
oai:surface.syr.edu:bio_etd-1018
2010-09-23T17:41:50Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Genetic and molecular analysis of ego-2, a positive regulator of Notch-type signaling in Caenorhabditis elegans
Liu, Ying
2007-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Notch signaling
GLP-1
ego-2
LIN-12
Genetics and Genomics
Life Sciences
Notch-type signaling plays a fundamental role in regulating the development of metazoans. In the gonad of nematode Caenorhabditis elegans, the germline mitosis/meiosis decision is regulated by somatic gonadal cells, including the distal tip cell (DTC) and the proximal anchor cell (AC), via GLP-1/Notch signaling. Although this signaling has four main components, more have been isolated that regulate GLP-1 signaling activity, one of which is the ego-2 ( e nhancer of g lp-1 ) gene, a genetic enhancer of a glp-1 loss-of-function (lf) mutation. GLP-1 signaling also regulates embryonic development. Another Notch receptor in C. elegans, LIN-12, functions in the soma.
My dissertation research has focused on the molecular identification of the ego-2 gene, characterization of its molecular structure and functions, and description of its relationship with its paralog, alx-1, and other components of the GLP-1/LIN-12/Notch pathway. EGO-2 not only promotes GLP-1 signaling in both germline and embryonic development, but also positively regulates the Z1.ppp-Z4.aaa interaction via LIN-12 during somatic gonad development, suggesting that EGO-2 is a general regulator of Notch signaling. ego-2 activity is mainly required in the soma to promote germline proliferation, consistent with the genetic data showing that it acts upstream or in parallel with glp-1. The ego-2(lf) also causes Spe ( Spe rmatogenesis) defect in the germ line and many other somatic phenotypes. Therefore, EGO-2 activity is involved in many aspects of development, which may or may not involve GLP-1/Notch signaling. EGO-2 and ALX-1 have opposite functions with respect to GLP-1 signaling in germline proliferation, but act in concert during spermatogenesis, suggesting their relationship is complex.
The EGO-2 protein is predicted to contain a Bro1 domain, which has been shown to be involved in endocytosis, a process which can positively regulate Notch signaling in many species. Based on our data, we hypothesize that in the DTC, as well as some other somatic gonadal and germ cells, EGO-2 functions in the endosomal process(es) to promote Notch signaling. To further analyze EGO-2 function, I first assembled a wildtype ego-2 hybrid genomic/cDNA construct, which can rescue some ego-2(lf) phenotypes. I tagged this construct with green fluorescent protein to examine the EGO-2 expression pattern in the future.
https://surface.syr.edu/bio_etd/17
oai:surface.syr.edu:bio_etd-1017
2010-09-23T17:28:58Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Expressional, functional and genomic studies of proteasome subunits during spermatogenesis of Drosophila melanogaster
Zhong, Lei
2007-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
John M. Belote
Proteasome subunits
Spermatogenesis
Genetics and Genomics
Life Sciences
Most regulated proteolysis in eukaryotic cells is carried out by a large, multisubunit protease called the 26S proteasome, which is composed of a 20S core particle and two 19S regulatory caps at each end. In Drosophila melanogaster , 6 subunits of the 20S core and 5 subunits of the 19S cap have multiple isoforms encoded by separate genes. The additional isoforms are invariably testis-specific. In the work reported here, I characterized the expression patterns of previously studied subunits Prosα3, Prosα3T and Prosα6T in more detail. In addition, I examined the expression patterns of several other proteasome subunits that had not been studied before, using eGFP-tagged reporter proteins. The housekeeping subunits, Prosα3, Prosα6, and Prosβ5 are expressed in early stages of spermatogenesis in both nuclei and cytoplasm, but gradually fade away in elongating spermatids, and are not detectable in mature sperm. In contrast, testis-specific Prosα3T, Prosα6T, Prosβ5R1 and Prosβ5R2 are expressed after meiosis, and persist in the mature sperm. Another interesting expression pattern is that some of these subunits localize to numerous punctate "speckles" trailing the actin cone complex in spermatid bundles undergoing individualization.
The functions of the Prosα6T subunit was studied by generating a null mutation Prosα6T 1 . Most of the spermatogenesis stages of the male-sterile Prosα6T 1 look normal under phase-contrast microscopy, but no mature sperm are produced. The Individualization Complexes (IC) becomes disorganized as they move along the tail bundles. Spermatid nuclei are either scattered or curled. Histone removal is delayed in mutant spermatid nuclei, but protamine incorporation is not affected. The expression analysis and the mutational study indicate that Prosα6T plays an essential role in Drosophila individualization.
In the second part of my study, I identified the proteasome subunit genes in 11 Drosophila species and three non- Drosophila insect species by database searches. The results showed that proteasome subunit gene duplication is a common phenomenon and that many additional genes are the results of retro transposition.
https://surface.syr.edu/bio_etd/18
oai:surface.syr.edu:bio_etd-1020
2010-09-27T15:05:53Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Nla4 and Nla18, key regulatory proteins required for normal growth and development of Myxococcus xanthus
Ossa-Ramirez, Faisury
2006-12-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Anthony G. Garza
Myxococcus xanthus
Nla4
Nla18
Regulatory proteins
Microbiology
Changes in gene expression are important for the landmark morphological events that occur during M. xanthus fruiting body development. NtrC-like activators play a prominent role in the coordinated expression of developmental genes. Mutations in the ntrC -like activator genes nla4 and nla18 are known to affect the timing of fruiting body formation, the morphology of mature fruiting bodies, and the sporulation efficiency. In this study we show that early developmental gene expression is altered in the early stages of fruiting body development in nla4 and nla18 cells. Production of A-signal, a quorum-sensing signal required early in development, is severely affected. The earliest event in development, accumulation of the intracellular starvation signal ppGpp is reduced. Thus, there is an early and strong modification of the developmental program in nla4 and nla18 cells.
Further studies revealed that nla4 cells fail to express normal levels of genes whose products are implicated in the regulation of ppGpp accumulation. Inactivation of nla4 reduces GTP accumulation, a precursor of (p)ppGpp, which also stimulates growth related processes in other bacteria. Furthermore, nla4 inactivation causes a dramatic decrease in the M. xanthus vegetative growth rate. It is likely that Nla4 is an important key in modulating (p)ppGpp accumulation by activating the expression of the stringent response related genes during the M. xanthus life cycle.
This study shows that inactivation of nla18 produces a strong defect in M. xanthus vegetative growth. DNA microarray analysis revealed that the vegetative expression patterns of more than 700 genes are altered in nla18 cells. Among the largest classes of altered genes are those which code for putative membrane and membrane-associated proteins. Our findings show that the membrane protein profiles isolated from vegetative nla18 and wild-type cells are noticeably different. Inactivation of nla18 causes misexpression of many genes that are likely to be important for protein synthesis. It therefore appears that nla18 cells are unable to regulate genes that encode important components of the translational machinery, thereby affecting stringent response. Our data are consistent with a model in which Nla18 controls vegetative growth and development by regulating the expression of genes involved in protein synthesis and membrane structure.
https://surface.syr.edu/bio_etd/21
oai:surface.syr.edu:bio_etd-1021
2010-09-28T14:14:26Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Postcopulatory sexual selection in Drosophila
Bjork, Adam Clarence
2006-12-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Scott Pitnick
Postcopulatory
Sexual selection
Entomology
The arena of sexual selection expands after copulation to include the female reproductive tract when promiscuous females house ejaculates from more than one male. Male-male competition continues in the form of sperm competition, and female choice continues in the form of cryptic female choice. This study aims to contribute (1) to the understanding of the mechanisms underlying postcopulatory sexual selection and (2) to the unification of postcopulatory sexual selection theory and traditional sexual selection theory.
Strong offensive and defensive sperm competitors are favored by postcopulatory sexual selection. Offensive and defensive sperm competitive ability were only found to be significantly repeatable in D. melanogaster across multiple sperm competition bouts between the same two males within the same female. Additionally, experimental evolution techniques revealed that the heritabilities of sperm offense and defense are low in genetically variable populations. These experiments highlight the complex nature of sperm precedence and the maintenance of genetic variation in ejaculate characteristics.
Postcopulatory sexual selection on males can lead to decreased sperm numbers by favoring larger sperm. However, a decline in sperm numbers is predicted to weaken selection on males and increase selection on females. Sexual selection for longer sperm, therefore, is expected to be self-limiting. Competitive mating experiments confirmed this "big-sperm paradox" in Drosophila . A resolution is provided by incorporating knowledge of postcopulatory processes into the interpretation of measures of sexual selection intensity.
Males with little sperm competition risk or few mating opportunities should divert resources away from gamete production because sperm are no longer regarded as energetically cheap and effectively limitless in supply. This prediction was met in the giant-sperm producing D. bifurca . Solitary males with infrequent access to females were found to produce sperm at a rate much slower than males raised in constant association with females and other males.
Understanding the adaptive significance of polyandry is a challenge. No direct fitness advantages of polyandry were found in D. melanogaster . If polyandry evolved to promote postcopulatory competition between males successful at precopulatory competition, then promiscuous females may benefit if these males disproportionately fertilize their eggs. However, no such indirect genetic benefits were revealed.
https://surface.syr.edu/bio_etd/22
oai:surface.syr.edu:bio-1004
2012-09-21T13:47:51Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
EGO-1, a Putative RNA-Directed RNA Polymerase, Promotes Germline Proliferation in Parallel With GLP-1/Notch Signaling and Regulates the Spatial Organization of Nuclear Pore Complexes and Germline P Granules in Caenorhabditis elegans
Vought, Valarie E.
Ohmachi, Mitsue
Lee, Min-Ho
Maine, Eleanor M
Article
2005-01-01T08:00:00Z
C. elegans EGO-1
Germline proliferation
GLP-1
Biology
<p>Caenorhabditis elegans EGO-1, a putative cellular RNA-directed RNA polymerase, promotes several aspects of germline development, including proliferation, meiosis, and gametogenesis, and ensures a robust response to RNA interference. In C. elegans, GLP-1/Notch signaling from the somatic gonad maintains a population of proliferating germ cells, while entry of germ cells into meiosis is triggered by the GLD-1 and GLD-2 pathways. GLP-1 signaling prevents germ cells from entering meiosis by inhibiting GLD-1 and GLD-2 activity. We originally identified the ego-1 gene on the basis of a genetic interaction with glp-1. Here, we investigate the role of ego-1 in germline proliferation. Our data indicate that EGO-1 does not positively regulate GLP-1 protein levels or GLP-1 signaling activity. Moreover, GLP-1 signaling does not positively regulate EGO-1 activity. EGO-1 does not inhibit expression of GLD-1 protein in the distal germline. Instead, EGO-1 acts in parallel with GLP-1 signaling to influence the proliferation vs. meiosis fate choice. Moreover, EGO-1 and GLD-1 act in parallel to ensure germline health. Finally, the size and distribution of nuclear pore complexes and perinuclear P granules are altered in the absence of EGO-1, effects that disrupt germ cell biology per se and probably limit germline growth</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/3
oai:surface.syr.edu:bio-1003
2012-09-21T13:46:37Z
publication:dbio
publication:coscde
publication:cas
publication:bio
EGO-1, a Putative RNA-Dependent RNA Polymerase, Is Required for Heterochromatin Assembly on Unpaired DNA during C. elegans Meiosis
Maine, Eleanor M
Hauth, Jessica
Ratliff, Thomas
Vought, Valarie E.
She, Xingyu
Kelly, William G.
Article
2005-11-08T08:00:00Z
C. elegans
EGO-1
Heterochromatin assembly
Meiosis
Biology
<p>During meiosis in C. elegans, unpaired chromosomes and chromosomal regions accumulate high levels of histone H3 lysine 9 dimethylation (H3K9me2), a modification associated with facultative heterochromatin assembly and the resulting transcriptional silencing [1, 2]. Meiotic silencing of unpaired DNA may be a widely conserved genome defense mechanism [3–5]. The mechanisms of meiotic silencing remain unclear, although both transcriptional and posttranscriptional processes are implicated [3–5]. Cellular RNA-dependent RNA polymerases (RdRPs) function in development and RNA-mediated silencing in many species [3, 6, 7] and in heterochromatin assembly in S. pombe [3, 8]. There are four C. elegans RdRPs, including two with known germline functions. EGO-1 is required for fertility and robust germline RNAi [9–11]. RRF-3 acts genetically to repress RNAi and is required for normal meiosis and spermatogenesis at elevated temperatures [12] (S. L’Hernault, personal communication). Among C. elegans RdRPs, we find that only EGO-1 is required for H3K9me2 enrichment on unpaired chromosomal regions during meiosis. This H3K9me2 enrichment does not require Dicer or Drosha nuclease or any of several other proteins required for RNAi. ego-1 interacts genetically with him-17, another regulator of chromatin and meiosis [13], to promote germline development. We conclude that EGO-1 is an essential component of meiotic silencing in C. elegans.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/1
oai:surface.syr.edu:bio-1002
2012-09-21T13:44:41Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Eukaryotic Translation Initiation Factor 5B Activity Regulates Larval Growth Rate and Germline Development in Caenorhabditis elegans
Yu, Xiang
Vought, Valarie E.
Conradt, Barbara
Maine, Eleanor M
Article
2006-01-01T08:00:00Z
eIF5B
C. elegans
germ line
GLP-1/Notch
translation initiation factor
Biology
<p>In C. elegans, a population of proliferating germ cells is maintained via GLP-1/Notch signaling; in the absence of GLP-1 signaling, germ cells prematurely enter meiosis and differentiate. We previously identified ego (enhancer of glp-1) genes that promote germline proliferation and interact genetically with the GLP-1 signaling pathway. Here, we report that iffb-1 (initiation factor five B) is an ego gene. iffb-1 encodes the sole C. elegans isoform of eukaryotic translation initiation factor 5B, a protein essential for translation. We have used RNA interference and a deletion mutation to determine the developmental consequences of reduced iffb-1 activity. Our data indicate that maternal iffb-1 gene expression is sufficient for embryogenesis, and zygotic iffb-1 expression is required for development beyond late L1/ early L2 stage. Partial reduction in iffb-1 expression delays larval development and can severely disrupt proliferation and differentiation of germ cells. We hypothesize that germline development is particularly sensitive to iffb-1 expression level.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/2
oai:surface.syr.edu:bio-1000
2012-09-21T13:36:02Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Grassland Root Communities: Species Distributions and How They Are Linked to Aboveground Abundance.
Frank, Douglas
Pontes, Alyssa
Maine, Eleanor M
Caruana, Julie
Raina, Ramesh
Raina, Surahbi
Fridley, Jason
Article
2010-01-01T08:00:00Z
FFLP ; Fluorescent fragment length polymorphism ; Grassland ; Plan competition ; Roots ; Yellowstone National Park
Biology
<p>There is little comprehensive information on the distribution of root systems among coexisting species, despite the expected importance of those distributions in determining the composition and diversity of plant communities. This gap in knowledge is particularly acute for grasslands, which possess large numbers of species with morphologically indistinguishable roots. In this study we adapted a molecular method, fluorescent fragment length polymorphism, to identify root fragments and determine species root distributions in two grasslands in Yellowstone National Park. Aboveground biomass was measured and soil cores (2 cm diam) were collected to 40 cm and 90 cm in an upland, dry grassland and a mesic, slope-bottom grassland, respectively, at peak foliar expansion. Cores were subdivided and species that occurred in each 10 cm interval were identified. The results indicated that the average number of species in 10 cm intervals (31 cm3) throughout the sampled soil profile was 3.9 and 2.8 at a dry and a mesic grassland, respectively. By contrast, average species number per 0.5 m2 determined by the presence of shoot material was 6.7 and 14.1 at dry and mesic sites, respectively. There was no relationship between soil depth and number of species per 10 cm interval in either grassland, despite the exponential decline of root biomass with soil depth at both sites. There also was no relationship between root frequency (i.e., the percentage of samples in which a species occurred) and soil depth for the vast majority of species at both sites. The preponderance of species were distributed throughout the soil profile at both sites. Assembly analyses indicated that species root occurrences were randomly assorted in all soil intervals at both sites, with the exception that F. idahoensis segregated from A. tridentata and P. spicata in 10-20 cm soil at the dry grassland. Root frequency throughout the entire sampled soil profile was positively associated with shoot biomass among species. Together these results indicated the importance of large, well proliferated root systems in establishing aboveground dominance. The findings suggest that spatial belowground segregation of species probably plays a minor role in fostering resource partitioning and species coexistence in these YNP grasslands.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/5
oai:surface.syr.edu:bio-1001
2012-09-21T13:42:23Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
The Bro1-Domain Protein, EGO-2, Promotes Notch Signaling in Caenorhabditis elegans
Liu, Ying
Maine, Eleanor M
Article
2007-01-01T08:00:00Z
C. elegans
Notch signaling
GLP-1 and LIN-12
EGO-2
Brol domain
Biology
<p>In Caenorhabditis elegans, as in other animals, Notch-type signaling mediates numerous inductive events during development. The mechanism of Notch-type signaling involves proteolytic cleavage of the receptor and subsequent transport of the receptor intracellular domain to the nucleus, where it acts as a transcriptional regulator. Notch-type signaling activity is modulated by post-translational modifications and endocytosis of ligand and receptor. We previously identified the ego-2 (enhancer of glp-1) gene as a positive regulator of germline proliferation that interacts genetically with the GLP-1/Notch signaling pathway in the germline. Here, we show that ego-2 positively regulates signaling in various tissues via both GLP-1 and the second C. elegans Notch-type receptor, LIN-12. ego-2 activity also promotes aspects of development not known to require GLP-1 or LIN-12. The EGO-2 protein contains a Bro1 domain, which is known in other systems to localize to certain endosomal compartments. EGO-2 activity in the soma promotes GLP-1 signaling in the germline, consistent with a role for EGO-2 in production of active ligand. Another C. elegans Bro1-domain protein, ALX-1, is known to interact physically with LIN-12/Notch. We document a complex phenotypic interaction between ego-2 and alx-1, consistent with their relationship being antagonistic with respect to some developmental processes and agonistic with respect to others.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/4
oai:surface.syr.edu:bio_etd-1022
2010-09-29T13:13:46Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Signal transduction by short-wavelength opsins
Dukkipati, Abhiram
2006-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Robert R. Birge
Signal transduction
Short-wavelength
Opsins
Color vision
Biophysics
Daytime color vision is mediated by cone opsin proteins, which belong to the family of GPCRs and are found in the cone photoreceptor cells of the retina. The light sensitive part of the opsin is an 11- cis retinal chromophore attached via a protonated Schiff base linkage to the protein. Absorption of a photon causes the 11- cis to all- trans isomerization of the chromophore, which in turn drives the protein through a sequence of thermally trappable conformations before forming the physiologically active state. The chromophore binding pocket plays a major role in tuning the spectral sensitivity of the chromophore in different opsin groups. These spectral tuning mechanisms are not clearly understood, especially for the opsin group that mediates color sensitivity in the violet and ultraviolet region of the energy spectrum. In the study presented in this thesis, several residues from the 2 nd and 3 rd transmembrane domains that are important for regulating spectral tuning of the dark as well as the primary photoproduct and signal transduction properties of a violet sensitive cone opsin have been identified. Based on the spectroscopic results combined with theoretical analysis, a spectral tuning mechanism unique to short-wavelength sensitive opsins is proposed. The ultraviolet sensitive opsins are unique since it is believed that these are the only visual pigments that have an unprotonated chromophore-retinylidene Schiff base linkage in the dark state. The photobleaching pathway of an ultraviolet sensitive opsin has been characterized using cryogenic and semi-low temperature spectroscopy. Evidence is provided to show that a conserved counterion mediated protonation of the chromophore-protein retinylidene Schiff base linkage is necessary for normal signal transduction in ultraviolet sensitive visual pigments.
https://surface.syr.edu/bio_etd/23
oai:surface.syr.edu:bio-1005
2012-09-21T13:49:08Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Caenorhabditis elegans atx-2 Promotes Germline Proliferation and the Oocyte Fate
Maine, Eleanor M
Hansen, Dave
Springer, Deborah
Vought, Valarie E.
Article
2004-01-01T08:00:00Z
C. elegans atx2
Gerline proliferation
Meiosis
GLP-1 signaling
Biology
<p>In the Caenorhabditis elegans germline, proliferation is induced by Notch-type signaling. Entry of germ cells into meiosis is triggered by activity of the GLD-1 and GLD-2 pathways, which function redundantly to promote meiosis and/or inhibit proliferation. Activation of the germline Notch-type receptor, GLP-1, ultimately inhibits the activities of the GLD-1 and GLD-2 pathways. We previously identified several ego (enhancer of glp-1) genes that promote germline proliferation and interact genetically with the GLP-1 signaling pathway. Here, we show that atx-2 is an ego gene. Our data suggest that ATX-2 is not a positive regulator of the GLP-1 signaling pathway and GLP-1 signaling is not the sole positive regulator of ATX-2 activity. Moreover, our data indicate that GLP-1 must have an additional function, which may be to repress activity of a third meiotic entry pathway that would work in parallel with the GLD-1 and GLD-2 pathways. In addition to its role in proliferation, ATX-2 acts downstream of FOG-2 to promote the female germline fate.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/12
oai:surface.syr.edu:bio-1011
2012-09-21T13:55:51Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
A Phylogenetic Analysis of Vertebrate and Invertebrate Notch-Related Genes
Maine, Eleanor M
Lissemore, James L.
Starmer, William T.
Article
1995-01-01T08:00:00Z
Notch
Cell surface receptor
Membrane protein
Protein Notch
Genetics
Invertebrate
Phylogeny
Vertebrate
Biology
<p>Members of the Notch gene family are thought to mediate inductive cell-cell interactions during develop ment of a wide variety of vertebrates and invertebrates. These genes encode transmembrane proteins that appear to act as receptors and contain three repeated sequence motifs. Two of these motifs (an epidermal growth factor like sequence and a cdc10/SW16/ankyrin sequence) have been found in a large number of unrelated proteins, while the third motif (a lin-12/Notch/glp-1 sequence) is unique to proteins of the Notch family. We present a phylogenetic analysis of 17 Notch-related genes from eight species that has implications as to the origins and relative functions of these genes in different species. Several independent gene duplications have occurred and at least one such duplication in the vertebrate lineage preceded the avian/mammalian divergence. Significantly, the overall organization of individual members of each internally repeated motif ap pears to have been conserved among species, suggesting that each repeat plays a unique role in protein function. Yet, where sequence divergence does occur among genes in vertebrate, dipteran, and nematode lineages, it may signify functional differences for specific regions in Notch-related proteins.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/6
oai:surface.syr.edu:bio-1007
2012-09-21T13:50:39Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Oogenesis in Caenorhabditis elegans
Maine, Eleanor M
Article
2001-01-01T08:00:00Z
Oogenesis
Caenorhabditis elegans
germ line
meiosis
Biology
<p>Oogenesis is the process of forming the female gamete, i.e., the ovum or egg. In Caenorhabditis elegans, gametes derive from a tissue called the germ line, which is specified early in embryonic development. Two major events occur during oogenesis: the oocyte precursor germ cell undergoes meiotic division and it accumulates substantial cytoplasm. In meiosis, two sequential rounds of cell division produce a haploid egg, with only one copy of each chromosome, from the diploid oocyte precursor cell. Simultaneously, a large volume of cytoplasm is accumulated; it contains yolk and numerous other components that are essential for early embryonic development. Meiotic progression seems to be an integral part of oogenesis, since a number of proteins are required meiotic progression and for the development of functional oocytes. For example, GLD-1, an RNA-binding protein, is required for maintenance of oocyte precursors in pachytene stage (see below); in its absence, female germ cells will enter meiosis and progress to pachytene stage, but then exit meiosis and return to mitotic proliferation. In contrast, male germ cells do not require GLD-1 for meiosis and gametogenesis.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/10
oai:surface.syr.edu:bio-1010
2012-09-21T13:57:59Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Enhancers of gZP1, a Gene Required for Cell-Signaling in Caenorhabditis elegans, Define a Set of Genes Required for Germline Development
Qiao, Li
Lissemore, James L.
Shu, Pei
Smardon, Anne
Gelber, Melanie B.
Maine, Eleanor M
Article
1995-01-01T08:00:00Z
Cmorhabditis ekgans
Germline
cell signaling
development
cell fate choice
Biology
<p>The distal tip cell (DTC) regulates the proliferation or differentiation choice in the Cmorhabditis ekgans germline by an inductive mechanism. Cell signaling requires a putative receptor in the germline, encoded by the glp-1 gene, and a putative signal from the DTC, encoded by the lag-2 gene. Both glp-1 and lag-2 belong to multigene gene families whose members are essential for cell signaling during development of various tissues in insects and vertebrates as well as C. elegans. Relatively little is known about how these pathways regulate cell fate choice. To identify additional genes involved in the glp-1 signaling pathway, we carried out screens for genetic enhancers of glp-1. We recovered mutations in five new genes, named ego (enhancer of glp-1), and two previously identified genes, lag-1 and glp-4, that strongly enhance a weak glp-1 loss-of-function phenotype in the germline. Ego mutations cause multiple phenotypes consistent with the idea that gene activity is required for more than one aspect of germline and, in some cases, somatic development. Based on genetic experiments, glp-1 appears to act upstream of ego1 and ego3W. e discuss the possible functional relationships among these genes in light of their phenotypes and interactions with glp-1.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/7
oai:surface.syr.edu:bio-1006
2012-09-21T13:52:03Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
RNAi As a Tool for Understanding Germline Development in Caenorhabditis elegans: Uses and Cautions
Maine, Eleanor M
Article
2001-01-01T08:00:00Z
Caenorhabditis elegans
C. elegans
RNA interference
RNAi
germ line
germ cells
development
germline development.
Biology
<p>RNA-mediated genetic interference (RNAi) has become a very useful tool for analyzing gene function in development and other processes. RNAi can be used as a complement to traditional genetic studies or as a primary means of determining biological function. However, the efficacy of RNAi depends on a variety of factors that the researcher must take into consideration. This review focuses on germline development in the nematode, Caenorhabditis elegans, and discusses the uses and limitations of RNAi in providing new information about gene function as well as the possible endogenous role RNAi plays in germline physiology.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/11
oai:surface.syr.edu:bio-1009
2012-09-21T13:53:08Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
A Conserved Mechanism for Post-Transcriptional Gene Silencing?
Maine, Eleanor M
Article
2000-09-13T07:00:00Z
Post-transcriptional gene silencing
Neurospora crassa
RNAi
Caenorhabditis elegans
Arabidopsis thaliana
Biology
<p>Proteins with homology to RNA-directed RNA polymerases function in post-transcriptional gene silencing: in quelling in the fungus Neurospora crassa, RNAi in the nematode Caenorhabditis elegans, and co-suppression in the mustard plant Arabidopsis thaliana. These findings are consistent with a conserved mechanism operating in these diverse species.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/8
oai:surface.syr.edu:bio-1008
2012-09-21T13:54:21Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
EGO-1 is related to RNA-directed RNA polymerase and functions in germ-line development and RNA interference in C. elegans
Smardon, Anne
Spoerke, Jill M.
Stacey, Steven C.
Klein, Marcia E.
Mackin, Nancy
Maine, Eleanor M
Article
2000-01-01T08:00:00Z
C. elegans
Germ-line development
Cell-fate determination
Biology
<p>Background: Cell-fate determination requires that cells choose between alternative developmental pathways. For example, germ cells in the nematode worm Caenorhabditis elegans choose between mitotic and meiotic division, and between oogenesis and spermatogenesis. Germ-line mitosis depends on a somatic signal that is mediated by a Notch-type signaling pathway. The ego-1 gene was originally identified on the basis of genetic interactions with the receptor in this pathway and was also shown to be required for oogenesis. Here, we provide more insight into the role of ego-1 in germ-line development. Results: We have determined the ego-1 gene structure and the molecular basis of ego-1 alleles. Putative ego-1 null mutants had multiple, previously unreported defects in germ-line development. The ego-1 transcript was found predominantly in the germ line. The predicted EGO-1 protein was found to be related to the tomato RNA-directed RNA polymerase (RdRP) and to Neurospora crassa QDE-1, two proteins implicated in post-transcriptional gene silencing (PTGS). For a number of germ-line-expressed genes, ego-1 mutants were resistant to a form of PTGS called RNA interference. Conclusions: The ego-1 gene is the first example of a gene encoding an RdRP-related protein with an essential developmental function. The ego-1 gene is also required for a robust response to RNA interference by certain genes. Hence, a protein required for germ-line development in C. elegans may be a component of the RNA interference/PTGS machinery.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/9
oai:surface.syr.edu:bio-1012
2012-09-21T13:59:43Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Analysis of the Multiple Roles of gld-I in Germline Development: Interactions With the Sex Determination Cascade and the glP-1 Signaling Pathway
Francis, Ross
Maine, Eleanor M
Schedl, Tim
Article
1995-01-01T08:00:00Z
Caenorhabditis elegansgene gld-1
Oocyte development
Genetic epistasis analysis
Germline
Tumor formation
Germ cell proliferation
Biology
<p>The Caenorhabditis elegansgene gld-1 is essential for oocyte development; in gld-1 (null) hermaphrodites, a tumor forms where oogenesis would normally occur. We use genetic epistasis analysis to demonstrate that tumor formation is dependent on the sexual fate of the germline. When the germlines sex determination pathway is set in the female mode (terminal fem/fog genes inactive), gld-1 (null) germ cells exit meiotic prophase and proliferate to form a tumor, but when the pathway is set in the male mode, they develop into sperm. We conclude that the gld-1 (null) phenotype is cell-type specific and that gld-1 ( + ) acts at the end of the cascade to direct oogenesis. We also use cell ablation and epistasis analysis to examine the dependence of tumor formation on the glp-I signaling pathway. Although glp-1 activity promotes tumor growth, it is not essential for tumor formation by gld-1 (null) germ cells. These data also reveal that gld-1 ( + ) plays a nonessential (and sex nonspecific) role in regulating germ cell proliferation before their entry into meiosis. Thus gld-1 ( + ) may negatively regulate proliferation at two distinct points in germ cell development: before entry into meiotic prophase in both sexes (nonessential premeiotic gld-1 function) and during meiotic prophase when the sex determination pathway is set in the female mode (essential meiotic gld-1 function).</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/13
oai:surface.syr.edu:bio-1013
2012-09-21T14:02:01Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Intragenic Dominant Suppressors of glp-1, a Gene Essential for Cell-Signaling in Caenorhabditis elegans, Support a Role for cdcl O/SWZ6/Ankyrin Motifs in GLP-1 Function
Lissemore, James L.
Currie, Peter D.
Turk, Christine M.
Maine, Eleanor M
Article
1993-01-01T08:00:00Z
Caenorhabditis elegans
Development
Temperature sensitive mutation
Suppressor gene
Reversion
Membrane protein
Molecular interaction
Ankyrin
Biology
<p>The glp-1 gene product mediates cell-cell interactions required for cell fate specification during development in Caenorhabditis elegans. To identify genes that interact with glp-1, we screened for dominant suppressors of two temperature-sensitive glp-1 alleles and recovered 18 mutations that suppress both germline and embryonic glp-1 phenotypes. These dominant suppressors are tightly linked to glp-1 and do not bypass the requirement for a distal tip cell, which is thought to be the source of a signal that is received and transduced by the GLP-1 protein. Using single-strand conformation polymorphism (SSCP) analysis and DNA sequencing, we found that at least 17 suppressors are second-site intragenic revertants. The suppressors, like the original glp-1(ts) mutations, are all located in the cdc10/SWI6/ankyrin domain of GLP-1. cdc10/SWI6/ankyrin motifs have been shown to mediate specific protein-protein interactions in other polypeptides. We propose that the glp-1(ts) mutations disrupt contact between GLP-1 and an as yet unidentified target protein(s) and that the dominant suppressor mutations restore appropriate protein-protein interactions.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/17
oai:surface.syr.edu:bio-1014
2012-09-21T14:00:46Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Suppressors of glp-1, a Gene Required for Cell Communication During Development in Caenorhabditis elegans, Define a Set of Interacting Genes
Maine, Eleanor M
Kimble, Judith
Article
1993-01-01T08:00:00Z
Alleles
Caenorhabditis elegans
Cell Communication/genetics
Chromosome Mapping
Suppressor genes
Helminth genes
Genetic Complementation Test
Mutation
Phenotype
Biology
<p>The glp-1 gene is essential for two cell interactions that control cell fate in Caenorhabditis elegans: induction of anterior pharynx in the embryo and induction of mitotic proliferation in the germ line. To identify other genes involved in these cell interactions, we have isolated suppressors of two temperature sensitive alleles of glp-1. Each of 14 recessive suppressors rescues both embryonic and germline glp-1(ts) defects. These suppressors are extragenic and define a set of six genes designated sog, for suppressor of glp-1. Suppression of glp-1 is the only obvious phenotype associated with sog mutations. Mutations in different sog genes show allele-specific intergenic noncomplementation, suggesting that the sog gene products may interact. In addition, we have analyzed a semidominant mutation that suppresses only the glp-1 germline phenotype and has a conditional feminized phenotype of its own. None of the suppressors rescues a glp-1 null mutation and therefore they do not bypass a requirement for glp-1. Distal tip cell function remains necessary for germline proliferation in suppressed animals. These suppressor mutations identify genes that may encode other components of the glp-1 mediated cell-signaling pathway or regulate glp-1 expression.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/16
oai:surface.syr.edu:bio-1015
2012-09-21T14:02:59Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Molecular Basis of Loss-of-Function Mutations in the glp-1 Gene of Caenorhabitis elegans
Kodoyianni, Voula
Maine, Eleanor M
Kimble, Judith
Article
1992-01-01T08:00:00Z
Mutation
Caenorhabditis elegans
Multigene family
Gene organization
Phenotype
Biology
<p>The glp-1 gene encodes a membrane protein required for inductive cell interactions during development of the nematode Caenorhabditis elegans. Here we report the molecular characterization of 15 loss-of-function (lf) mutations of glp-1. Two nonsense mutations appear to eliminate glp-1 activity; both truncate the glp-1 protein in its extracellular domain and have a strong loss-of-function phenotype. Twelve missense mutations and one in-frame deletion map to sites within the repeated motifs of the glp-1 protein (10 epidermal growth factor [EFG]-like and 3 LNG repeats extracellularly and 6 cdc10/SW16, or ankyrin, repeats intracellularly). We find that all three types of repeated motifs are critical to glp-1 function, and two individual EFG-like repeats may have distinct functions. Intriguingly, all four missense mutations in one phenotypic class map to the N-terminal EGF-like repeats and all six missense mutations in a second phenotypic class reside in the intracellular cdc10/SW16 repeats. These two clusters of mutations may identify functional domains within the glp-1 protein.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/15
oai:surface.syr.edu:bio-1016
2012-09-21T14:04:19Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Carboxy-Terminal Truncation Activates glp-1 Protein to Specify Vulval Fates in Caenorhabditis elegans
Mango, Susan E.
Maine, Eleanor M
Kimble, Judith
Article
1991-08-29T07:00:00Z
Amino acid sequence
Caenorhabditis elegans Proteins
Cell division
Caenorhabditis/embryology
Caenorhabditis/genetics
Germ cells
Membrane proteins
Molecular sequence data
Mutation
Phenotype
Vulva/embryology
Vulva growth and development
Biology
<p>The glp-1 and lin-12 genes encode homologous transmembrane proteins that may act as receptors for cell interactions during development. The glp-1 product is required for induction of germ-line proliferation and for embryogenesis. By contrast, lin-12 mediates somatic cell interactions, including those between the precursor cells that form the vulval hypodermis (VPCs). Here we analyse an unusual allele of glp-1, glp-1(q35), which displays a semidominant multivulva phenotype (Muv), as well as the typical recessive, loss-of-function Glp phenotypes (sterility and embryonic lethality). We find that the effects of glp-1(q35) on VPC development mimic those of dominant lin-12 mutations, even in the absence of lin-12 activity. The glp-1(q35) gene bears a nonsense mutation predicted to eliminate the 122 C-terminal amino acids, including a ProGluSerThr (PEST) sequence thought to destabilize proteins. We suggest that the carboxy terminus bears a negative regulatory domain which normally inactivates glp-1 in the VPCs. We propose that inappropriate glp-1(q35) activity can substitute for lin-12 to determine vulval fate, perhaps by driving the VPCs to proliferate.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/14
oai:surface.syr.edu:bio-1017
2012-09-21T13:39:51Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Regulation of Heterochromatin Assembly on Unpaired Chromosomes during Caenorhabditis elegans Meiosis by Components of a Small RNA-Mediated Pathway
She, Xingyu
Xu, Xia
Fedotov, Alexander
Kelly, William G.
Maine, Eleanor M
Article
2009-01-01T08:00:00Z
Caenorhabditis elegans
Caenorhabditis elegans proteins
Chromosomes
Genetics
Germ cells
Heterochromatin and genetics
Heterochromatin and metabolism
Meiosis
RNA
Biology
<p>Many organisms have a mechanism for down regulating the expression of non-synapsed chromosomes and chromosomal regions during meiosis. This phenomenon is thought to function in genome defense. During early meiosis in Caenorhabditis elegans, unpaired chromosomes (e.g., the male X chromosome) become enriched for a modification associated with heterochromatin and transcriptional repression, dimethylation of histone H3 on lysine 9 (H3K9me2). This enrichment requires activity of the cellular RNA-directed RNA polymerase, EGO-1. Here we use genetic mutation, RNA interference, immunofluorescence microscopy, fluorescence in situ hybridization, and molecular cloning methods to identify and analyze three additional regulators of meiotic H3K9me2 distribution: CSR-1 (a Piwi/PAZ/Argonaute protein), EKL-1 (a Tudor domain protein), and DRH-3 (a DEAH/D-box helicase). In csr-1, ekl-1, and drh-3 mutant males, we observed a reduction in H3K9me2 accumulation on the unpaired X chromosome and an increase in H3K9me2 accumulation on paired autosomes relative to controls. We observed a similar shift in H3K9me2 pattern in hermaphrodites that carry unpaired chromosomes. Based on several assays, we conclude that ectopic H3K9me2 accumulates on paired and synapsed chromosomes in these mutants. We propose alternative models for how a small RNA-mediated pathway may regulate H3K9me2 accumulation during meiosis. We also describe the germline phenotypes of csr-1, ekl-1, and drh-3 mutants. Our genetic data suggest that these factors, together with EGO-1, participate in a regulatory network to promote diverse aspects of development.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/19
oai:surface.syr.edu:bio-1018
2012-09-21T13:40:18Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Studying gene function in Caenorhabditis elegans using RNA-mediated interference
Maine, Eleanor M
Article
2008-01-01T08:00:00Z
Caenorhabditis elegans
RNA
functional genomics
gene networks
gene interaction
Biology
<p>The RNA interference (RNAi) method for targeted gene silencing is widely used in Caenorhabditis elegans for large-scale functional genomic studies, analysis of limited gene sets and detailed analysis of individual gene function. The application of RNAi has identified genes that participate in various aspects of development, physiology and cell biology. In addition, RNAi has been used to identify interacting genes and to study functionally redundant genes. This review discusses the various applications of RNAi in C. elegans, focusing particularly on the analysis of developmental processes.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/18
oai:surface.syr.edu:bio-1019
2012-09-21T14:06:18Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Genetics of intercellular signalling in C. elegans
Austin, Judith
Maine, Eleanor M
Kimble, Judith
Article
1989-01-01T08:00:00Z
Caenorhabditis elegans
cell-cell interaction
cell fate
glp-1
lin-12
Biology
<p>Cell-cell interactions play a significant role in controlling cell fate during development of the nematode Caenorhabditis elegans. It has been found that two genes, glp-1 and lin-12, are required for many of these decisions. glp-1 is required for induction of mitotic proliferation in the germline by the somatic distal tip cell and for induction of the anterior pharynx early in embryogenesis. lin-12 is required for the interactions between cells of equivalent developmental potential, which allow them to take on different fates. Comparison of these two genes on a molecular level indicates that they are similar in sequence and organization, suggesting that the mechanisms of these two different sets of cell-cell interactions are similar.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/23
oai:surface.syr.edu:bio-1020
2012-09-21T14:05:05Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Identification of Genes That Interact with glp-1, a Gene Required for Inductive Cell Interactions in Caenorhabditis elegans
Maine, Eleanor M.
Kimble, Judith
Article
1989-01-01T08:00:00Z
Caenorhabditis elegans
extragenie suppressor glp-1
Cell interaction
Temperature-sensitive mutant
Biochemistry
Molecular Biology
<p>The glp-1 gene functions in two inductive cellular interactions and in development of the embryonic hypodermis of C. elegans. We have isolated six mutations as recessive suppressors of temperature-sensitive (ts) mutations of glp-1. By mapping and complementation tests, we found that these suppressors are mutations of known dumpy (dpy) genes; dpy genes are required for development of normal body shape. Based on this result, we asked whether mutations previously isolated in screens for mutants defective in body shape could also suppress glp-1(ts). From these tests, we learned that unselected mutations of eight genes required for normal C. elegans morphogenesis, including the four already identified, suppress glp-1(ts). All of these suppressors rescue all three mutant phenotypes of glp-1(ts) (defects in embryonic induction of pharyngeal tissue, in embryonic hypodermis development, and in induction of germline proliferation). However, they do not rescue putative glp-1 null mutants and therefore do not bypass the requirement for glp-1 in development. In the light of current ideas about the molecular nature of the glp-1 and suppressor gene products, we propose an interaction between the glp-1 protein and components of the extracellular matrix and speculate that this interaction may impose spatial constraints on the decision between mitosis and meiosis in the germline.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/22
oai:surface.syr.edu:bio-1021
2012-09-21T14:05:35Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
The Drosophila Female-Specific Sex-Determination Gene, Sex-Lethal, Has Stage-, Tissue-, and Sex-Specific RNAs Suggesting Multiple Modes of Regulation
Salz, Helen K.
Maine, Eleanor M.
Keyes, Linda N.
Samuels, Mark E.
Cline, Thomas W.
Schedl, Paul
Article
1989-01-01T08:00:00Z
Drosophila melanogaster
Sex-lethal
sex determination
dosage compensation
sex-specific transcripts
Biology
<p>For proper sexual development of females, the Sex-lethal (Sxl) gene must be activated early in development and remain on during the rest of the life cycle. Conversely, in males, Sxl must remain functionally off through development. Here, we show that the Sxl transcription unit spans a DNA segment of greater than 20 kb and encodes at least 10 distinct, but overlapping, RNA species. These RNAs range in size from 4.4 to 1.7 kb and exhibit sex, stage, and tissue specificity. Six RNAs, three female specific and three male specific, are first detected by midembryogenesis and persist through the adult stage: Their expression reflects the on/off regulation of Sxl's activity at the level of sex-specific alternate splicing. Four Sxl RNAs are found in adult females. Two of these RNAs are dependent on the presence of a functional germ line and may be relevant to Sxl's role in adult germ-line development. All four are present in unfertilized eggs. Finally, three Sxl RNAs are found only transiently during very early embryogenesis; we suggest that the expression of these RNAs may reflect an early regulation of Sxl at the level of transcription and that these transcripts are involved in the initial selection of the Sxl activity state in response to the primary sex-determination signal, the X/A ratio.</p>
http://creativecommons.org/licenses/by/3.0/
https://surface.syr.edu/bio/21
oai:surface.syr.edu:bio_etd-1023
2010-09-30T17:28:36Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Enzyme responses of Serengeti grasses to defoliation: Coupling plant cellular processes and Serengeti ecosystem processes
Dong, Yan
2005-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Samuel J. McNaughton
Grasses
Defoliation
Ecosystem processes
Sporobolus kentrophyllus
Themeda triandra
Digitaria macroblephara
Botany
Ecology and Evolutionary Biology
Life Sciences
Plant Sciences
Grazing can cause grass compensatory growth (McNaughton 1983) and compensatory photosynthesis (Detling et al. 1979). However, different species have different tolerance levels toward grazing (Oesterheld and McNaughton 1988, 1991). Responses of grasses after grazing depend partially on photosynthesis and N assimilation rates. Three Serengeti C4 grasses, Sporobolus kentrophyllus, Themeda triandra and Digitaria macroblephara , were examined for their enzyme responses to defoliation, light conditions, nitrogen fertilization and soil texture. Assays in cell-free extracts and gel densitometry of proteins were conducted. Four key enzymes were studied, including two enzymes crucial to carbon assimilation, phosphoenolpyruvate carboxylase (PEPc) and pyruvate, orthophosphate dikinase (PPDK), and two crucial to nitrogen assimilation, nitrate reductase (NR) and glutamine synthetase (GS).
In a comparison study with S. kentrophyllus and T. triandra , all four enzymes and total soluble protein content in the former species responded positively to defoliation; but in the latter species, none of the four enzymes responded positively to defoliation. Positive enzymatic responses of S. kentrophyllus to defoliation happens on both high light level and low light level, suggesting that positive enzymatic responses to defoliation did not depend on light conditions. Increased PEPc activity after defoliation was partially due to increased PEPc protein synthesis. Enzyme responses of S. kentrophyllus required N fertilization to be above a critical level. A comparison study showed that D. macroblephara responded positively to defoliation and sandy soil; while T. triandra did not respond to defoliation and responded negatively to sandy soil.
These results showed that survival and growth of grasses under grazing depend on their enzymatic responses to defoliation. Enzymes and proteins of grasses also respond promptly to environmental changes of nutrient level, light conditions and soil texture. Photosynthetic enzymes and N assimilating enzymes sometimes are associated in their responses to environmental factors. This reflects the interaction and relation between carbon metabolism and N metabolism in grasses. Differential enzyme responses of grasses to defoliation and soil texture may be part of the adaptation of the grasses to grazing conditions. Therefore enzymatic responses contribute to the spatial distribution of species through grass-herbivore interaction in the Serengeti.
https://surface.syr.edu/bio_etd/25
oai:surface.syr.edu:bio_etd-1024
2010-09-30T20:01:18Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The effects of mineral nitrogen on embryonic and larval amphibians
Griffis-Kyle, Kerry L.
2005-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Mark E. Ritchie
Nitrogen
Embryonic
Larval
Amphibians
Ecology and Evolutionary Biology
Forest Sciences
Life Sciences
Many amphibian populations have declined over the past several decades, often in response to anthropogenic stressors, one of which may be mineral nitrogen (N). Increasing N enrichment of the environment has become one of the largest and most pervasive environmental threats. Over the last century, increased fossil fuel consumption, fixation of previously unavailable gaseous N 2 for fertilizer, as well as other anthropogenic activities have doubled the amount of available N in the environment. This mineral N can have both direct and indirect toxic effects on aquatic organisms.
I used a series of laboratory experiments to demonstrate that nitrite exposure can have large impacts on amphibians. In aquatic systems nitrite can accumulate if there is an asymmetry in chemical transformations in either nitrification or denitrification. Only two previous studies have examined the effects of nitrite on amphibian mortality. I showed that early nitrite exposure can not only cause mortality, but can also have a delayed effect on later larval survival. This is particularly important because large fluxes of N during early spring arrive coincident with embryo and early larval development and can cause mortality later in ontogeny. I also demonstrated that nitrite slows growth and development throughout embryonic and larval development. I discussed the implications of slowed growth and development from nitrite toxicity in ephemeral environments where many amphibians breed.
In a whole-pond fertilization experiment in a series of ponds, I demonstrated that mineral N can reduce survival and growth of amphibians in the wild, in addition to causing decreases in the overall number of frog tadpoles in the ponds. I have demonstrated that ammonium and nitrate plus nitrite can remain high enough in surface waters to cause toxic reactions in amphibians that manifest on both population and community levels.
Overall, my research demonstrates that mineral N can increase amphibian mortality and slow growth and development in embryonic and larval stages and is important to consider when managing amphibian populations, especially in amphibians that breed in ephemeral pools where mineral N concentrations can be elevated into the summer and delays in development can lead to catastrophic failures in reproduction.
https://surface.syr.edu/bio_etd/24
oai:surface.syr.edu:bio_etd-1025
2010-10-01T14:09:44Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The spatial and temporal regulation of morphogenesis in the budding yeast Saccharomyces cerevisiae
Mackin, Nancy A.
2006-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Scott E. Erdman
Morphogenesis
Cell polarity
Paxillin
Calcium
Cell and Developmental Biology
Molecular Biology
Contributing to a number of cellular responses including fibroblast motility and axonal guidance, morphogenesis, more commonly referred to as polarized growth, is fundamental to many aspects of cell and developmental biology. An important goal for understanding the mechanisms of polarized growth is to determine the full complement of polarity proteins functioning in the establishment and maintenance of polarized growth sites as well as their dynamics of assembly and disassembly. To elucidate these mechanisms, we employed the model eukaryote budding yeast or Saccharomyces cerevisiae . Given that S. cerevisiae exhibits polarized growth in at least two facets of its life cycle: vegetative growth (or budding) and mating, we focused our investigations on the roles of two genes, PXL1 and FIG1 , with putative functions during these aspects of polarized growth in budding yeast.
Sequence analysis of Pxl1p, or Paxillin-like protein 1, revealed the presence of two C-terminal LIM domains that bear homology to those of the mammalian scaffolding protein paxillin. In higher eukaryotes, paxillin is targeted to polarized growth sites via its LIM domains where it has been shown to function in the assembly and maintenance of polarization sites through its associations with various signaling and actin cytoskeletal organization components. Similar to paxillin, localization of Pxl1p to polarized growth sites is dependent on the function of its individual LIM domains. At these sites, Pxl1p participates in such polarized growth events as budding and mating projection formation, possibly as a modulator of Rho GTPase signaling. These findings demonstrate an evolutionarily conserved role for LIM domain proteins as mediators of cell signaling and cytoskeletal organization during polarized growth.
The discovery of FIG1 , or F actor- i nduced g ene 1 , in a screen for pheromone-regulated genes suggested Fig1p functions exclusively at the time of mating. In this collaborative study, Fig1p has been shown to act as a novel component or regulator of the calcium influx system LACS. Our studies demonstrate LACS activities function cooperatively with those of HACS to positively regulate cell fusion via the influx of calcium ions at the time of mating, providing the first direct evidence for calcium signals in promoting cell-cell fusion during yeast mating.
https://surface.syr.edu/bio_etd/26
oai:surface.syr.edu:bio_etd-1028
2010-10-01T19:59:15Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Molecular evolution of visual pigments of the Tokay gecko and bluefin killifish
Blow, Nathan S.
2004-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
John Belote
Visual pigments
Tokay gecko
Bluefin killifish
Gekko gekko
Lucania goodei
Animal Sciences
Biochemistry, Biophysics, and Structural Biology
Life Sciences
Molecular Biology
Zoology
Visual pigments are proteins which absorb photons and convert light into neuronal signals which are interpreted by the visual cortex of the brain. Functionally, the visual pigment has two major roles: absorb a photon within a specific spectral range and initiate a G-protein signaling cascade. In nature, visual pigments absorb light of many different wavelengths, from 360 nm to 635 nm. Previous studies have shown that changing specific amino acid residues in visual pigments alters maximal light absorbance (λmax), and that this may enhance vision under various environmental lighting conditions. The goal of this dissertation was to study visual pigment function and evolution in two species, the Tokay gecko and the Bluefin killifish, from two distinct lighting environments, a nocturnal habitat and an aquatic habitat. First, in Chapter 2, the vision of the nocturnal Tokay gecko was studied. It was determined that this species relies on only three cone visual pigments for vision (SWS1, RH2, and MWS) and two of these cone visual pigments are dramatically blue-shifted in λmax (RH2 and MWS). Secondly, in Chapters 3 and 4, the molecular evolution of visual pigments in the bluefin killifish were examined. Eight opsin cDNA's were identified and characterized for this species. It was demonstrated that for short wavelength vision, improved chromatic discrimination arose in the bluefin killifish as the result of a gene duplication of the SWS2 class opsin and amino acid changes at sites 44, 94, 118, and 265 to blue-shift the function of one copy, changing the function from blue light absorbance to violet light absorbance.
https://surface.syr.edu/bio_etd/27
oai:surface.syr.edu:bio_etd-1026
2010-10-01T19:01:53Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Determinants of plant species diversity across spatial scales in Serengeti National Park, Tanzania
Anderson, T. Michael
2004-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Samuel J. McNaughton
Serengeti National Park
Tanzania
Plant diversity
Soil nitrate
Botany
Ecology and Evolutionary Biology
Life Sciences
Plant Sciences
The determinants of plant species and functional-type diversity were studied in savanna-grasslands of Serengeti National Park, Tanzania. Experiments and observational studies spanned multiple spatial scales, from 1 m 2 plots to large areas of similar climate, topography, and geomorphology classified as Land Regions. Scale-dependence was a universal and important ecological property of Serengeti grasslands; small scale diversity (1 m 2 ) was associated with climate factors while larger scale diversity (1000 m 2 ) was associated with landscape factors. A measure of among-plot similarity in species composition vs. distance varied among Land Regions, with eastern regions having no variation among plot similarity with distance and western plots showing a strong decay in compositional similarity as plot distance increased.
The spatial distribution of soil nitrate was related to plant species diversity and plant size distributions. Small-scale variation in soil nitrate was positively correlated with 1 m 2 plant species diversity (H ' ), while 1000 m 2 plant species richness was a log normal function of soil nitrate patch size. Moreover, between scales of 10 m 2 and 1000 m 2 , where the spatial distribution of soil nitrate was fractal-like, plant species diversity was greater and plant size distributions were more left-skewed than where the spatial distribution of soil nitrate was random. Computer simulation models suggested that the fractal dimension of the spatial distribution of resources affects plant functional-type diversity when plants are of different sizes and their nutrient requirements are size-specific. Plant diversity in simulated landscapes was greater when the fractal dimension of the resource was lower (more aggregated and patchy) compared to when resources had a higher fractal dimension (more diffuse and grainy).
Finally, a series of laboratory and field experiments conducted at multiple spatial scales suggested that the dominant plant species in Serengeti, Digitaria macroblephara and Themeda triandra , coexist by exploiting different topographic hillslopes positions that differ in water availability and grazing frequency. Isotopic tracer studies conducted in the field and physiological studies conducted in the laboratory showed that the species have opposite responses to water availability and grazing, but that grazing may promote species coexistence by producing different responses to water availability.
https://surface.syr.edu/bio_etd/29
oai:surface.syr.edu:bio_etd-1027
2010-10-01T19:57:22Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Molecular genetic analysis of TART (telomere-associated retrotransposon) in Drosophila melanogaster
Maxwell, Patrick
2004-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
John Belote
Telomere-associated retrotransposon
Retrotransposons
RNA interference
Biochemistry, Biophysics, and Structural Biology
Life Sciences
Molecular Biology
Telomeres are the ends of linear chromosomes, and Drosophila melanogaster telomeres are exceptional in that they consist of tandem arrays of two families of non-LTR (long terminal repeat) retrotransposons, HeT-A and TART. Maintenance of D. melanogaster telomeric DNA is accomplished by repeated retrotransposition of HeT-A and TART specifically to telomeres, in contrast to the maintenance of tandem arrays of species-specific simple DNA repeats at telomeres by telomerase in many eukaryotic organisms. R[barbelow]apid a[barbelow]mplification of c[barbelow]DNA e[barbelow]nds (RACE) was used to analyze the 5 ' and 3 ' ends of TART transcripts and antisera that recognize TART ORF1p, one of two proteins encoded by TART, were generated and used to examine the expression pattern of ORF1p to better characterize TART elements. The transcript analysis was consistent with TART producing an array of heterogeneous sense and antisense transcripts. The identification of transcription initiation sites in the direct repeats of TART, and sequence analysis of a major sense transcription initiation site suggest a parallel between transcription of TART and Drosophila LTR retrotransposons. Also, intron sequences were identified in antisense transcripts. The specificity of antisera against TART ORF1p was determined by inducing RNA interference (RNAi) against TART. Western blotting experiments with these antisera revealed that TART ORF1p accumulates throughout much of development, especially in larval and pupal stages. ORF1p was detected in somatic cells, and notably in larval imaginal discs and brains, but was not detected in germline cells. Temperature-dependent pupal lethality observed in some strains in which RNAi was induced raises the possibility that induction of RNAi might lead to nonspecific lethality under certain circumstances in Drosophila .
https://surface.syr.edu/bio_etd/28
oai:surface.syr.edu:bio_etd-1029
2010-10-04T13:00:08Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Trafficking and turnover of the calcium sensing receptor
Huang, Ying
2006-12-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Gerda E. Breitwieser
Calcium sensing
Endoplasmic reticulum
Ubiquitination
Molecular Biology
The calcium sensing receptor (CaR), a member of G protein-coupled receptor family C, regulates systemic calcium homeostasis by activating G q - and G i -linked signaling in the parathyroid, kidney and intestine.
To examine agonist-mediated cell surface trafficking of CaR, we quantified CaR by ELISA (enzyme-linked immunosorbent assay), and found that upon stimulation by extracellular Ca 2+ , CaR undergoes multiphasic internalization that is dependent on changes in intracellular Ca 2+ .
To define the importance of the intracellular C-terminus of CaR in trafficking and targeting, we characterized amino acids/regions potentially critical for receptor localization. An RKR motif (from amino acid 896 to 898) within the proximal region of the C-terminus of CaR was demonstrated to be an ER retention/retrieval signal that becomes functional when exposed. By grafting the RKR motif to ectopic sites within the C-terminus, we found that the RKR motif functions in a distance-sensitive manner, being more potent at distal locations within the C-terminus.
To further characterize the contribution of the C-terminus of CaR to trafficking and turnover, we carried out yeast two-hybrid library screening to identify potential interactors for the intracellular C-terminus of CaR. Screening both human kidney and brain cDNA libraries, we identified nineteen interacting proteins, one of which is an E3 ubiquitin ligase named dorfin ( do uble R ING- f inger prote in ). Biochemical analyses demonstrated that CaR and dorfin interact in mammalian cells; ubiquitination of CaR is observed in the presence of proteasomal inhibitor MG132; and ubiquitination and degradation of CaR are mediated by dorfin via ERAD ( e ndoplasmic r eticulum- a ssociated d egradation) pathway.
To begin to characterize the biogenesis of CaR, we examined the structural and functional checkpoints in the ER ( e ndoplasmic r eticulum) quality control mechanism for CaR. Impaired N-linked glycosylation or disulfide bond formation of CaR results in ER retention and degradation by ERAD, suggesting that glycosylation and disulfide bond formation serve as structural checkpoints in the biogenesis of CaR. Interestingly, we identified a potential conformation-dependent checkpoint in the biogenesis of the CaR, using both receptor mutants and allosteric drugs.
https://surface.syr.edu/bio_etd/30
oai:surface.syr.edu:bio_etd-1030
2010-10-06T14:04:36Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Ecological and social correlates of foraging decisions in a social forager, the bonnet macaque, Macaca radiata diluta
Ratnam, Jayashree
2002-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Larry L. Wolf
Macaca radiata diluta
Foraging
Social forager
Bonnet macaque
Behavior and Ethology
Zoology
This study investigated ecological and social factors determining social foraging behavior in a diurnal, group-foraging primate, the southern Indian bonnet macaque, Macaca radiata diluta . It set out to ask how ecological and social variables interacted to influence the individual forager and whether either subset of the environment, ecological or social was more important in determining an individual's response. A series of foraging decisions was analyzed in response to experimentally induced variation in (1) ecological variables, including patch number, patch size, inter-patch distances, food search time, food quality and potential predation risk in habitat and (2) social variables, including social dominance rank, feeding group sizes, feeding group compositions and intra-group levels of aggression.
Social dominance emerged as an important organizing factor and determined which individuals fed at a given time in experimental trials. As long as habitats were relatively free from predation, dominant males, dominant females and sub-adult classes were well represented. Subordinate females were somewhat less represented while subordinate males were severely constrained and did not feed in most trials.
Similarly, social rank affected the spatial distribution of individuals across patches. Dominant males typically came into a feeding area and spread out uniformly over food patches. In sharp contrast, sub-adults aggregated into one or two patches. Dominant females were randomly distributed across patches in most experimental trials.
Social rank also affected an individual's feeding rate. Dominant males fed in a slow 'leisurely' fashion in almost all conditions, were unaffected to a great degree by the individuals around them, and showed little variation in their foraging returns across a wide range of conditions. In contrast, individuals of other social ranks fed much faster and showed more variation in their foraging returns, which were affected to varying degrees by interactions with other individuals around them.
Interference among foragers was an important underlying factor that drove the above patterns. The presence and distribution of subordinate classes were best explained as strategies of avoidance of adult males who engage in frequent and ritualized aggressions. In response to this, subordinate adult males avoided areas with dominant males, while sub-adults traveled in groups and aggregated in space, effectively reducing per-capita levels of aggression upon themselves. The behaviors of females as a class were more ambiguous and did not show easily discerned patterns. It is hypothesized that high levels of relatedness in this class skewed interference costs for this class in unknown ways.
A simple functional model for this system is proposed and it is shown that the relative importance of ecological versus social factors is mediated by social rank and interference.
https://surface.syr.edu/bio_etd/33
oai:surface.syr.edu:bio_etd-1031
2010-10-07T18:46:29Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
A molecular and genetic analysis of the SNR1 subunit of the Brahma chromatin remodeling complex
Marenda, Daniel Raymond
2003-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Andrew K. Dingwall
SNR1
Brahma
Chromatin
Brm
Biochemistry, Biophysics, and Structural Biology
Genetics and Genomics
Life Sciences
Molecular Biology
The Drosophila melanogaster Brahma (Brm) complex, a counterpart of the Saccharomyces cerevisiae SWI/SNF ATP-dependent chromatin remodeling complex, is a multi-protein complex important for proper development by maintaining specific gene expression patterns. The snr1 gene of Drosophila melanogaster encodes a conserved component of Brm complex. Loss-of-function and null mutations in the snr1 gene reveal its essential role in Drosophila development, while specific mutations in Ini1 , the human snr1 homolog, are associated with aggressive cancers, underscoring the importance of this particular subunit in development. This thesis focuses on a genetic and molecular analysis of the SNR1 subunit during Drosophila development, centering around the characterization and analysis of a temperature-sensitive allele of the snr1 gene, snr1 E1 . Genetic analyses of snr1 E1 reveal that it functions as an antimorph and that snr1 has critical roles in tissue patterning and growth control. Importantly, during wing vein formation in Drosophila , the SNR1 subunit appears to constrain the ATPase activities of the BRM subunit in maintaining or activating the expression of particular genes critical for proper vein formation to occur. Analysis of the SNR1 E1 protein in the context of Drosophila vein formation has therefore revealed an important regulatory function of SNR1 to activate or repress gene expression through specific associations with other members of the Brm complex.
https://surface.syr.edu/bio_etd/32
oai:surface.syr.edu:bio_etd-1032
2010-10-07T20:15:49Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Ecological and evolutionary variation in heat-shock proteins
Barua, Deepak
2003-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Scott A. Heckathorn
Stress response
Photosynthesis
Thermotolerance
Chenopodium album
Solidago altissima
HSP
Botany
Ecology and Evolutionary Biology
Life Sciences
Plant Sciences
In spite of the ubiquitous nature of heat-shock proteins (Hsps), and their central role in thermotolerance, there is variation in the patterns of expression of Hsps. While much is now known about Hsp function, it remains unclear why we see such variation. A primary objective of my work was to understand variation in the heat-shock response (HSR), and examine the relationship of the HSR to the thermal environment. In particular, I focussed on the temperature set-points of the HSR and magnitude of Hsp expression. In a comprehensive analysis of the HSR, I demonstrate a strong relationship between the set-points of the HSR and growth temperature in diverse species, across a broad range of temperatures. Deviation from this relationship was related to the ecological and evolutionary history of organisms. The HSR also exhibits plasticity in response to changing growth temperatures, and I describe general patterns of plasticity in the HSR. In examining variation in Hsp content in Chenopodium album , I show that contrary to expectations, populations from more stressful habitats had lower Hsp content and induced thermotolerance. Among populations, acclimation to higher growth temperatures decreased Hsp content. Organisms from stressful habitats that require frequent induction of the HSR may down-regulate the HSR, and instead rely on basal mechanisms of thermotolerance. An in vitro assay was used to explicitly determine the adaptive significance of natural variation in Hsps. I demonstrate natural variation in protection of photosynthetic electron transport by chloroplast small Hsps (csHsp), and show that csHsps can account for most of the induced thermotolerance observed. In the final study, I examine the influence of light on Hsp expression in the field. I show that Hsp content is greater in Solidago altissima plants in the sun, than in the shade. In accordance with these results, I demonstrate a significant effect of light and temperature on Hsp accumulation in the laboratory. These results add to the surprisingly small number of studies that have examined Hsp accumulation in field plants, confirm previous results on the effect of light, and importantly, show that light influences Hsp accumulation in plants in their natural habitat.
https://surface.syr.edu/bio_etd/31
oai:surface.syr.edu:bio_etd-1033
2010-10-08T13:13:23Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Genetic and molecular characterization of nutritional signaling pathways regulating meiosis in Saccharomyces cerevisiae
Lee, Rita H.
2002-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Saul Honigberg
Nutritional
Meiosis
Glucose
Molecular Genetics
Environmental signals regulate many cell differentiation processes in eukaryotes. In the yeast Saccharomyces cerevisiae , environmental signals regulate meiosis. Initiation of meiosis requires the expression of IME1 , an essential gene for meiosis. Expression of the IME1 is blocked by glucose. Previous work suggested that in low concentrations of glucose, overexpression of IME1 was sufficient to induce high levels of recombination. My work demonstrates that these cells undergo the early meiotic events including DNA replication, commitment to recombination, and synaptonemal complex formation and dissolution. In contrast, the late meiotic events including chromosome segregation, commitment to meiosis, and spore formation do not occur. These cells are stably arrested and when transferred to sporulation medium, are able to complete meiosis. This implies that extracellular nutrients regulate meiosis in yeast at both early and late stages of meiosis.
Specifically, glucose was shown to regulate meiosis at multiple stages ( IME1 transcription, IME2 transcription, and entry into late stages of meiosis). Therefore, since many cellular responses to glucose in yeast are mediated through the glucose repression pathway, the central component of this pathway was analyzed for its role in meiosis.
My work confirmed that SNF1 is required both prior to and after DNA replication. Given this finding, I constructed deletion mutants of other components of the glucose repression pathway ( SNF3, RGT2, RGT1, CAT8 , and SIP4 ) to determine their role in glucose repression of meiosis. My results suggest that SNF3 and RGT2 are involved, in part, in glucose repression of meiosis. Furthermore, while the downstream component RGT1 is not involved in glucose repression, cells lacking RGT1 were mildly defective in meiosis. Candidates for Snf1 targets were also tested. Deletion of the transcriptional repressors MG1 and YHP1 fails to suppress the meiotic defects of a snflΔ mutant. Deletion of CAT8 , but not SIP4 , caused severe defects in meiosis. Therefore, this works demonstrates that only a subset of the glucose repression pathway components are involved in regulation of meiosis.
https://surface.syr.edu/bio_etd/34
oai:surface.syr.edu:bio-1022
2010-10-08T15:32:19Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
XanthusBase: Adapting Wikipedia Principles to a Model Organism Database
Arshinoff, Bradley I.
Suen, Garret
Just, Eric M.
Merchant, Sohel M.
Kibbe, Warren A.
Chisholm, Rex L.
Welch, Roy D.
Article
2007-01-01T08:00:00Z
Myxococcus xanthus ; xanthusBase ; Generic Model Organism Database ; MediaWiki ; dictyBase ; Myxobacterial research ; Open-source software
Biology
xanthusBase (http://www.xanthusbase.org) is the official model organism database (MOD) for the social bacterium Myxococcus xanthus. In many respects, M.xanthus represents the pioneer model organism (MO) for studying the genetic, biochemical, and mechanistic basis of prokaryotic multicellularity, a topic that has garnered considerable attention due to the significance of biofilms in both basic and applied microbiology research. To facilitate its utility, the design of xanthusBase incorporates open-source software, leveraging the cumulative experience made available through the Generic Model Organism Database (GMOD) project, MediaWiki (http://www.mediawiki.org), and dictyBase (http://www.dictybase.org), to create a MOD that is both highly useful and easily navigable. In addition, we have incorporated a unique Wikipedia-style curation model which exploits the internet’s inherent interactivity, thus enabling M.xanthus and other myxobacterial researchers to contribute directly toward the ongoing genome annotation.
https://surface.syr.edu/bio/20
oai:surface.syr.edu:bio_etd-1034
2010-10-11T12:57:22Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Large herbivores and process dynamics in a managed savanna ecosystem
Augustine, David J.
2002-12-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Samuel J. McNaughton
Nitrogen cycling
Savanna
Ungulates
Phosphorus
Biogeochemistry
Physical and Environmental Geography
Systems Biology
Terrestrial and Aquatic Ecology
Feedbacks between soil nutrients, plant communities, and large mammalian herbivores were studied at the Mpala Ranch and Research Centre in Laikipia, Kenya. The landscape consists of short-grass glades, typically 0.5-1.0 ha in size, dispersed throughout the dominant Acacia bushland vegetation. I examined (1) whether glades are created through the use and abandonment of overnight cattle corrals or 'bomas', (2) the importance of these glades as habitat for native and domestic ungulates, (3) feedback effects of herbivores on aboveground herbaceous productivity (ANPP) and soil nutrient dynamics, and (4) effects of climatic variability on herbivore abundance and soil-grass-grazer interactions.
All soil characteristics measured across a boma-glade chronosequence indicate glades are indeed derived from abandoned bomas. In particular, soil N, P and organic matter quality in the surface (0-15 cm) layer were similar for glades and 30-39 year old bomas, but were enriched relative to surrounding bushland. Soil texture was similar for bomas, glades, and bushland, indicating glades were not derived from a unique parent material. Cynodon leaves from bomas and glades were highly enriched in P, Ca and N relative to Cynodon from nearby bushland sites. Local abundance of impala, zebra and eland was closely tied to the distribution of nutrient-rich glades. Seasonal analyses of impala habitat selection suggested that selection for glade habitat was related both to predation risk and the availability of mineral-rich forage. In particular, P in boma and glade grass was above recommended levels for growing and lactating ruminants, while P content of bushland grass was lower than recommended levels.
Across a soil nutrient gradient, large herbivore consumption rates were linearly related to ANPP. A fertilization experiment and analyses of grass N:P ratios indicate that N and P co-limit productivity on the most nutrient-poor sites. Grazing pressure was consistently high (>60% of ANPP) at all but one site in a dry year (1999), and was greater in nutrient-rich glades (73.0 ± 4.2% of ANPP) compared to nutrient-poor bushland sites (42.7 ± 6.7% of ANPP) in a wet year (2001). Nitrogen budgets constructed for nutrient-rich and nutrient-poor sites showed that large herbivores themselves caused a net N input to the former and a net N loss from the latter. During short (1 month) growing seasons, grazers reduced aboveground productivity regardless of soil nutrient availability. However, over a 5 month growing season, grazers increased ANPP on glades and suppressed ANPP on bushland sites. Grazers increased the size of the inorganic N pool available to plants at onset of the growing season, but also decreased N mineralization rates at all sites early in the growing season. N availability measured via ion-exchange resin bags showed that the net effect of grazers was to increase in N availability at glade sites, but to decrease in N availability at bushland sites. This net effect on N availability mirrored grazer effects on ANPP in the high-rainfall year. Overall, results indicate that grazer effects on N balance and N availability in ecosystems are closely linked to effects on productivity and resilience to drought, and that both soil fertility and climatic variation mediate grazer effects on ecosystem processes.
https://surface.syr.edu/bio_etd/38
oai:surface.syr.edu:bio_etd-1035
2010-10-11T15:41:58Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Molecular genetics and evolution of red-green vision in vertebrates
Radlwimmer, Friedrich Bernhard
2002-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Scott Pitnick
Shozo Yokoyama
Evolution
Visual pigments
Red-green vision
M/LWS
Vertebrates
Evolution
Genetics
Molecular Biology
To investigate the molecular mechanism and evolution of red-green color vision, we cloned, sequenced and expressed in vitro the red-green sensitive type pigment genes of 8 mammals, goat ( Capra hircus ), rabbit ( Oryctolagus cuniculus ), rat ( Rattus norvegicus ), cat ( Felis catus ), horse ( Equus caballus ), deer ( Odocoileus virginianus ), guinea pig ( Cavia porcellus ) and squirrel ( Sciurus carolinensis ), and the goldfish ( Carassius auratus ). Additionaly, red-green pigments of three more vertebrates, chicken ( Gallus gallus ), frog ( Xenopus laevis ) and cave fish ( Astyanax mexicanus ), that previously had been isolated by other investigators, were expressed in vitro. The absorption spectra of the purified goat, rabbit, rat, cat, horse, deer, guinea pig, squirrel, chicken, frog, cave fish red, cave fish green and goldfish pigments had wavelengths of maximal absorption (λ max ) of 553 ± 1, 509 ± 1, 509 ± 1, 553± 1, 545 ± 1, 531 ± 1, 516 ± 1, 532 ± 1, 561 ± 2, 558 ± 2, 559 ± 2, 530 ± 2 and 559 ± 4 nm, respectively. Multiple linear regression analyses showed that differences between the λ max values of vertebrate red-green sensitive visual pigments estimated in vitro can be fully explained by the effects of amino acid differences at positions 180, 197, 277, 285, 308, and the pair-wise interaction of positions 180 and 197. Amino acid substitutions S180A, H197Y, Y277F, T285A and A308S and interaction the interaction between sites 180 and 197 were estimated to shift λ max value by about -7, -28, -8, -15 and -27 and +11 nm, respectively. This model was corroborated experimentally by site-directed mutagenesis and in vitro expression of constructed visual pigments. Furthermore, the evolution of vertebrate red-green pigments was experimentally retraced by construction and in vitro expression of inferred vertebrate ancestral pigments. The results of this analysis suggest that extant red-green pigments evolved from a red-sensitive ancestral pigment with a λ max value of about 560 nm.
https://surface.syr.edu/bio_etd/37
oai:surface.syr.edu:bio_etd-1036
2010-10-11T15:47:08Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Molecular genetics and evolution of UV vision in vertebrates
Shi, Yongsheng
2002-09-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Shozo Yokoyama
Evolution
UV
Vision
Spectral tuning
Evolution
Genetics
Molecular Biology
Many vertebrates achieve UV vision through a class of photoreceptors in the retina containing UV-sensitive visual pigments with optimal light absorption (λ max ) at 360∼370 nm. UV vision has been used for such basic behaviors as communication, foraging, and mating. Despite its biological importance, the molecular bases and the evolution of UV vision are not well understood. In this study, we first identified the major amino acid residues that determine the spectral sensitivity of UV pigments in vertebrates. We also provided strong supportive evidence that UV pigments, unlike all the other visual pigments, are based on an unprotonated Schiff base chromophore. We then inferred the amino acid sequences of the ancestral pigments of UV/violet pigments in vertebrates. These ancestral pigments were then reconstructed by introducing the necessary mutations into the contemporary pigments and their absorption spectra evaluated by using an in vitro assay. Our results demonstrated that the common ancestor of vertebrates and most other ancestors had UV-sensitive pigments, and most contemporary UV pigments maintained the ancestral function. The ancestral pigments of birds achieved violet sensitivity by four amino acid substitutions, and some extant avian species regained UV vision by a change at another amino acid site. In conclusion, this study not only revealed the molecular mechanisms for spectral tuning in UV pigments, but also elucidated the evolutionary history of UV vision in vertebrates.
https://surface.syr.edu/bio_etd/36
oai:surface.syr.edu:bio_etd-1037
2010-10-11T19:35:30Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The community dynamics of protein phosphatase 2A subunits in Saccharomyces cerevisiae
Gentry, Matthew Shawn
2003-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Richard L. Hallberg
Protein phosphatase 2A
Holoenzymes
Cell cycle
Biochemistry, Biophysics, and Structural Biology
Cell and Developmental Biology
Cell Biology
Genetics and Genomics
Life Sciences
Molecular Biology
Protein phosphatase 2A (PP2A) regulates a broad spectrum of cellular processes. This holoenzyme is a collection of varied heterotrimeric complexes, composed of a catalytic (C) and regulatory (B) subunit bound together by a structural (A) subunit.
To understand the cell cycle dynamics of this enzyme population, we carried out quantitative and qualitative analyses of PP2A subunits in Saccharomyces cerevisiae . We found: the level of each subunit remained constant throughout the cell-cycle; there is at least ten times more of one of the regulatory subunits (Rts1p) than the other (Cdc55p); Tpd3p, the structural subunit, is limiting for both catalytic and regulatory subunit binding; the two regulatory subunits display distinctly different dynamic localization patterns that overlap with the A and C subunits at the bud tip, kinetochore, bud neck and nucleus. Using strains null for single subunit genes, we confirmed the hypothesis that regulatory subunits determine sites of PP2A accumulation. Additionally, we identified some cis- and trans-acting machinery involved in PP2A Cdc55p and PP2A Rts1p localizations.
In vitro binding assays demonstrated that C subunit post-translational modification(s) affects PP2A heterotrimer stabilities. However, the degree to which different types of heterotrimers are affected in vivo unknown. Using subunit localization to assess in vivo binding stabilities, we found that Pph21p C-terminal mutants that are not efficiently modified and cells lacking the C subunit's methyltransferase, PPM1 , both show reduced PP2A localization, presumably because the formation of stable PP2A heterotrimers is decreased. Furthermore, Cdc55p-directed PP2A localizations are reduced to a greater degree than Rts1p-directed localizations. Thus, the formation of stable PP2A heterotrimers is dependent on C subunit modification and PP2A Cdc55p stability is more dependent on this than is PP2A Rtrs1p .
In vitro data also suggest that specific regions of the A subunit are necessary for heterotrimer formation. However, there is little to no in vivo data to support this finding. Thus, we determined the role of various A subunit regions in in vivo heterotrimeric makeup. Overall, we found that PP2A heterotrimers are more stable in vivo than they are in vitro . Using PP2A mutant phenotypes, we found that PP2A Rts1p is more resistant to A subunit mutations than is PP2A Cdc55p .
https://surface.syr.edu/bio_etd/35
oai:surface.syr.edu:bio_etd-1038
2010-10-12T14:14:12Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Predictions and quantitative tests of optimal time and temperature allocation during intermittent incubation
Voss, Margaret A.
2002-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
F. Reed Hainsworth
Incubation
Eggs
Foraging
Ecology and Evolutionary Biology
Physiology
Poultry or Avian Science
Tests of optimization models are often criticized when the qualitative predictions of general models cannot be falsified. However, adequate tests of specific models require both quantitative predictions and precise data. Precise modeling techniques for thermal transients permit prediction of the egg temperature ( T Eexit ) at which a bird should leave the nest to maximize percent time foraging ( P ) when constrained by incubation. Within an incubation cycle, [Special characters omitted.] , when t cool , t heat , and t equil are times for egg cooling, heating, and maintenance at a constant temperature, and τ is travel time. T Eexit values for 4 bird species were compared with those predicted by the model. Observed T Eexit values for approximately half of all incubation cycles did not maximize P (N = 243; 4 species combined). Variation in incubation patterns produced slightly different average egg temperatures for each species, possibly producing different embryo development times. However, average predicted T Eexit across entire incubation periods (egg laying to hatching), were the same as observed. Thus, birds may maximize long-term P by combining incubation cycles with variation in time components to compensate for non-optimal behavior. Time required for incubation reduced P from a theoretical maximum of 100% to 19.77% for Black-capped Chickadees, 28.06% for Yellow-eyed Juncos, 34.27% for Tree Swallows, and 39.4% for House Wrens. Additional optimization criteria were also considered. T Eexit to maximize the rate of net energy gain ( RNEG ) differed from observed for individual junco, wren, and swallow incubation cycles and entire incubation periods. T Eexit to maximize RNEG for chickadee incubation cycles differed from observed, but average optimal T Eexit was the same as observed for the entire incubation period. The chickadees' ability to achieve the predicted average optimal T Eexist is linked to superior nest insulation and food caching behavior. T Eexist to maximize foraging efficiency differed from observed for all species. Although tests for all criteria could be falsified for individual cycles, predictions for P for entire incubation periods could not be falsified. Critics of optimization modeling would consider this a panglossian, and thus flawed, result. However, it appears valid and is likely the result of attempts to balance reproductive effort with self-maintenance over long time periods.
https://surface.syr.edu/bio_etd/39
oai:surface.syr.edu:bio_etd-1039
2010-10-15T17:56:53Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Disturbance, diversity and community dynamics in a southern Indian savanna-grassland ecosystem
Sankaran, Mahesh
2001-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
S. J. McNaughton
Disturbance
Diversity
Indian
Savanna-grassland
Ecology and Evolutionary Biology
Life Sciences
Factors influencing structure and dynamics of savanna-grassland ecosystems were investigated at the Kalakad-Mundanthurai Tiger Reserve (KMTR), India, across spatial scales ranging from a few square meters to the entire reserve (900 km 2 ). Savanna habitats comprise a third of the reserve area, and are characterized by dominance of a few widespread species that turn-over across an elevation gradient. Most mid- and low-elevation grasslands are anthropogenic in origin, maintained by recurrent fires. Tall grasses dominate most low and mid elevations, short-grasses characterize high elevations. Plant richness is greatest at low elevations, and both point richness and pattern diversity are important determinants of site richness in the reserve. Herbivores primarily utilize short grasslands at low and high elevations, and avoid tall-grasslands due to the unpalatable nature of dominant plants.
In recent years, fire frequencies have increased and herbivore populations have decreased in the reserve. To understand causes, and potential consequences, of these changes, experimental manipulations were carried out to determine how burning and grazing influence plant and nutrient dynamics in three low-elevation grassland communities. Low diversity communities were more compositionally stable than high diversity communities to perturbations characteristic of their native environment, but less so when perturbations were novel. Grazing effects were more pronounced than burning. Increased colonization by annual species following high rainfall during the study period also induced compositional shifts in communities. Such shifts were greater in high diversity communities because of more free-space for colonization in these communities. When considering treatment effects on soil nitrogen dynamics, responses were complex and contingent on community type, disturbance history and season.
Herbivore densities at KMTR are low relative to other similar protected areas in India. It is hypothesized that herbivore declines and spread of tall-grasses at KMTR has resulted from disruption of facilitatory interactions between different body-sized herbivores. Reduction in large-bodied herbivores, the control agents for coarse tall grasses, has favored the spread of competitively superior tallgrasses in an environment where light competition is critical. High litter production by these species has increased fire frequencies and suppressed smaller palatable plants, subsequently reducing densities of selectively feeding smaller-bodied herbivores. This hypothesis awaits experimental testing.
https://surface.syr.edu/bio_etd/40
oai:surface.syr.edu:bio_etd-1040
2010-10-18T17:49:12Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Functional study of RTS1 and CDC55, two genes encoding regulatory subunits of protein phosphatase 2A in Saccharomyces cerevisiae
Yang, Haifeng
2001-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Richard L. Hallberg
John Russel
RTS1
CDC55
Encoding regulatory subunits
Protein phosphatase 2A
Saccharomyces cerevisiae
Biochemistry, Biophysics, and Structural Biology
Life Sciences
Molecular Biology
Protein phosphatase 2A (PP2A) is a major serine/threonine phosphatase in eukaryotic cells. Its activity has been implicated in the control of many cellular processes. It is a heterotrimeric complex comprised of a catalytic, a structural and a regulatory subunit.
In Saccharomyces cerevisiae, RTS1 encodes a regulatory subunit of PP2A. rts1Δ cells are temperature sensitive for growth and have a global stress response defect. In order to find out what other cellular functions require RTS1 , a search for multicopy suppressors of the temperature sensitive rts1 -null cells was carried out. The genes CLB2, PIR2, MSB1, PH085 , YLR426W and CAK1 were found to be multicopy suppressors. As indicated by the suppressors' functions, rts1 -null cells were found to have a G2/M delay at 37°C, weakened cell walls at 37°C, and a possible budding defect at 39°C.
The mechanism of ts suppression achieved by overproducing CLB2 was further investigated. We found that Clb2p levels in cells arrested in S phase was decreased relative to that seen in wild type cells, and this decrease was shown to be caused by the anaphase-promoting complex (APC)-dependent ectopic degradation. In investigating the role of two APC regulators, we also found that the ectopic degradation of Clb2p in rts1 -null cells was Cdc20p-dependent but Cdh1p-independent.
Loss of CDC55 , another regulatory subunit of PP2A, causes the cells to exhibit a failure of cytokinesis and the production of abnormally elongated buds at low temperature. As cdc55 -null cells exhibit a hyperphosphorylation of Y19 on the cyclin-dependent kinase Cdc28, we examined wild type and cdc55 -null cells for the levels and activities of the kinase (Swe1p) and phosphatase (Mih1p) that normally regulate the extent of this phosphorylation. Our results showed that Mih1p contributed little to this hyperphosphorylation. By contrast, Swe1 kinase was significantly elevated in mitosis arrested cdc55 -null cells. This excess buildup of Swe1p in cdc55 -null cells is the result of ectopic stabilization of this protein during G2/M phase. Present evidence indicating that this is a result of a gain-of-function of PP2A complexes lacking Cdc55p. My work shows that different PP2A regulatory subunits are required for the proper function of distinctly different cell cycle related pathways.
https://surface.syr.edu/bio_etd/44
oai:surface.syr.edu:bio_etd-1041
2010-10-18T18:23:51Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Identification, characterization and functional study of par1 and par2, two genes encoding B' subunits of protein phosphatase 2A in Schizosaccharomyces pombe
Jiang, Wei
2001-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Richard L. Hallberg
Protein phosphatase 2A
Schizosaccharomyces pombe
par1
par2
Cytokinesis
Biochemistry, Biophysics, and Structural Biology
Cell and Developmental Biology
Cell Biology
Genetics and Genomics
Life Sciences
Molecular Biology
Protein phosphatase 2A (PP2A) is a major serine/threonine phosphatase found in all eukaryotic cells. The PP2A holoenzyme is comprised of a catalytic C subunit, a structural A subunit, and a regulatory B subunit.
In my thesis work, I identified and characterized two B ' subunit genes of PP2A in Schizosaccharomyces pombe , namely, par1 and par2 . Both genes share high homology with other known B ' genes. Neither is essential but together they are required for growth at both high and low temperatures, growth under a number of physiologically stressful conditions, and septum positioning and cytokinesis. Cross-organism experiments showed that both par1 and par2 could complement a deletion of Saccharomyces cerevisiae RTS1 , a gene encoding a B ' homolog. Furthermore, both Par1p and Par2p physically associate with the catalytic subunits of PP2A, confirming that both genes encode S. pombe PP2A B ' subunits.
In some par mutant cells, the septum is not centrally located. Fluorescence microscopy revealed that the mitotic actin ring and Mid1p are also eccentrically located in par1Δpar2Δ cells, indicating that par genes are involved in regulating Mid1p and actin ring localization to ensure correct septum positioning. We also found that in par mutants, interphase Mid1p nuclear staining is much weaker than that seen in wild type cells, and Western blots showed that this might be caused by a reduced Mid1p abundance.
par1Δpar2Δ cells also show a multi-septation phenotype, very similar to that seen in hyperactive SIN (septation initiation network) mutants. I examined the genetic interactions between par deletions and SIN mutants, and found that par1Δpar2Δ suppressed Byr4-OP defects, and rescued a loss-of-function allele of spg1 , but could not suppress any mutations in genes downstream of spg1 . I showed further that par deletions suppressed the lethality of the spg1 allele through restoration of the correct localization of Cdc7p to the spindle pole body (SPB). The fact that par mutants themselves exhibited a symmetric localization of Cdc7p to SPBs indicated a hyperactivity of SIN in such cells. I concluded that par genes normally negatively regulate SIN at or upstream of cdc7 insuring that multiple rounds of septation do not occur.
https://surface.syr.edu/bio_etd/43
oai:surface.syr.edu:bio_etd-1042
2015-01-12T15:44:36Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Influence of trophic factors on the intra-plant distribution of aphids
Gould, Georgianna Grimshaw
2001-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Larry L. Wolf
Trophic factors
Aphids
Chaitophorous populicola
Populus deltoides
Botany
Ecology and Evolutionary Biology
Entomology
Life Sciences
Plant Sciences
<p>Feeding site selection behavior of the aphid Chaitophorous populicola on cottonwood ( Populus deltoides ) was studied to determine whether bottom-up or top-down trophic factors better explain intra-plant feeding location. C. populicola are often found on stems and lamina of new and rapidly expanding leaves, and on petioles of senescing leaves. Bottom-up factors associated with leaf developmental stage were examined in cottonwood phloem sap collected from cut petioles using EDTA-enhanced exudation. Sap exudation rate was relatively constant, as indicated by sucrose concentrations. HPLC analysis revealed that new leaves have a higher content of the phenolic glycoside salicin and aspartic acid than other developmental stages (20 amino acids were analyzed). Mature leaves have higher concentrations of gamma-amino butyric acid (GABA) and are least preferred by C. populicola . Also, leaf toughness, measured by vascular bundle lignin density and distance from phytodermal surface to vascular bundle, increased with leaf age and could contribute to fewer aphids selecting mature leaves. Conversely, top-down factors also contribute to aphid feeding distribution patterns. Ant mutualists are associated with increased aphid population sizes and occurrence of aphids on mature leaves. Ants prefer sucrose solutions containing asparagine, GABA, salicin and salicortin over sucrose alone, suggesting that these may attract tending ants if excreted in aphid honeydew. Ladybugs reduced aphid populations in localized areas and did not initiate aphid migration within plants. Phloem sap phytochemicals that vary among leaves of different developmental stages may serve as cues for the aphid C. populicola feeding on cottonwood, and for their associated mutualistic ants. Honeydew collected from aphids on leaves of different developmental stages was analyzed by HPLC. The phytochemicals studied did not vary qualitatively or quantitatively in the honeydew between leaves of different developmental stages, indicating that they are not transmitted as cues to the third trophic level. However, aphids on mature leaves produced more honeydew than on other leaf stages. Ants tending aphids may therefore respond to increases in honeydew quantity as opposed to elements of honeydew quality. Thus, aphid intra-plant feeding location is directly and indirectly driven by plant-mediated (bottom-up) factors, and influenced directly by ant mutualists to a lesser extent.</p>
https://surface.syr.edu/bio_etd/42
oai:surface.syr.edu:bio_etd-1043
2010-10-18T20:29:39Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The alpha3T gene of Drosophila melanogaster encodes a spermatogenesis-specific proteasome subunit
Ma, Jing
2001-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
John M. Belote
Drosophila melanogaster
alpha3T
Proteasome subunit
Spermatogenesis
Biochemistry, Biophysics, and Structural Biology
Life Sciences
Molecular Biology
The proteasome is responsible for degradation of substrates of the ubiquitin pathway. 20S proteasomes are cylindrical particles with subunits arranged in a stack of four heptameric rings. The outer rings are composed of a subunits, and the inner rings are composed of β subunits. One major question in proteasome research concerns the extent and nature of proteasome structural heterogeneity. To investigate this topic, a molecular and genetic study of Drosophila proteasomes has been undertaken, with a goal of identifying proteasome subunit isoforms exhibiting developmental or cell-type specificity. Previous work led to the isolation of two proteasome subunit genes, encoding α4 isoforms that exhibit testis-specific expression (Yuan, et al., 1996). Here, I report the identification and characterization of a gene, α 3T , encoding a testis-specific isoform of another 20S proteasome subunit, α3.
The α 3T and α3 genes are unlinked, and they encode proteins that are 58% identical. Northern blot analyses reveal that α3 is expressed throughout development in both sexes, while α 3T is expressed primarily during the pupal and adult stages, and only in males. The tissue-specific expression pattern was examined in transgenic flies expressing GFP fusion proteins, revealing that the testis-specific α 3T -GFP only becomes prominent during and after meiosis, accumulates in the nucleus at later stages of spermatogenesis and is present in the head of mature, motile sperm. This pattern is in contrast to that of α3-GFP, which is expressed in all tissues examined. In testes, α3-GFP is prominently expressed in early stages of spermatogenesis in the cytosol and nucleus, but it fades from the elongating spermatid nuclei as the α 3T subunit accumulates. α 3T can functionally replace its yeast ortholog and, thus, encodes a functional proteasome subunit. Ectopic constitutive expression of α 3T in flies causes dominant pupal lethality, while expressing the α3 gene in the same way has no detectable consequence, suggesting that the α 3T subunit is functionally distinct from α3. In an attempt to silence α 3T gene expression, an inverted repeat of a portion of the α 3T gene (α 3T-IR ) was expressed in flies using the UAS/GAL4 system. The expressed α 3T-IR caused a dominant pupal lethality, instead of the anticipated male sterility phenotype.
https://surface.syr.edu/bio_etd/41
oai:surface.syr.edu:bio-1023
2010-12-17T15:32:01Z
publication:dbio
publication:coscde
publication:cas
publication:biopub
publication:bio
Transport of ER Vesicles on Actin Filaments in Neurons by Myosin V
Langford, George M
Tabb, Joel S
Molyneaux, Bradley J
Cohen, Darien L
Kuznetsov, Sergei A
Article
1998-10-14T07:00:00Z
Myosin V
ER transport
Squid giant axon
Organelle/vesicle movement
Actin filament
Axonal transport
Biology
Cell Biology
Axoplasmic organelles in the giant axon of the squid have been shown to move on both actin filaments and microtubules and to switch between actin filaments and microtubules during fast axonal transport. The objectives of this investigation were to identify the specific classes of axoplasmic organelles that move on actin filaments and the myosin motors involved. We developed a procedure to isolate endoplasmic reticulum (ER) from extruded axoplasm and to reconstitute its movement in vitro. The isolated ER vesicles moved on exogenous actin filaments
adsorbed to coverslips in an ATP-dependent manner without the addition of soluble factors. Therefore myosin was tightly bound and not extracted during isolation. These vesicles were identified as smooth ER by use of an antibody
to an ER-resident protein, ERcalcistorin/protein disulfide isomerase (EcaSt/PDI). Furthermore, an antibody to squid myosin V was used in immunogold EM studies to show that myosin V localized to these vesicles. The antibody was
generated to a squid brain myosin (p196) that was classified as myosin V based on comparisons of amino acid sequences of tryptic peptides of this myosin with those of other known members of the myosin V family. Dual labeling with the squid myosin V antibody and a kinesin heavy chain
antibody showed that the two motors colocalized on the same vesicles. Finally, antibody inhibition experiments were performed with two myosin V-specific antibodies to show that myosin V motor activity is required for transport of vesicles on actin filaments in axoplasm. One antibody was made to a peptide in the globular tail domain and the other to the globular head fragment of myosin V. Both antibodies inhibited vesicle transport on actin filaments by greater than 90% compared to controls. These studies provide the first direct evidence that ER vesicles are transported on actin filaments by myosin V. These data confirm the role of actin filaments in fast axonal transport and provide support for the dual filament model of vesicle transport.
Work done while GM Langford was a member of the Department of Biological Sciences, Dartmouth College, Hanover, NH 03755-3576, USA.
Other authors (Tabb, Cohen, Molyneaux) were also at Dartmouth. Kuznetsov was at Universität Rostock.
Langford is the author for correspondence.
https://surface.syr.edu/bio/24
oai:surface.syr.edu:bio_etd-1044
2010-10-25T19:52:20Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Genetic and molecular characterization of genes that interact withglp-1 in Caenorhabditis elegans germline development
Smardon, Anne Marie
1999-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Richard Levy
Cell signaling
Glp-1
Caenorhabditis elegans
Germline development
Cell and Developmental Biology
Genetics and Genomics
Life Sciences
Signaling between cells is required for the proper specification of cell fate during the development of multicellular organisms. In Caenorhabditis elegans an inductive interaction between the somatic distal tip cells (DTCs) and the distal germline is essential in order to maintain proliferation in the developing germline. This cell signaling relies on a conserved signaling pathway with three known components: the LAG-2 ligand, GLP-1 receptor and LAG-1 effector. These proteins are homologous to members of "Notch" signaling pathways which function in multiple aspects of Drosophila development as well as the development of a number of vertebrate species. In order to identify other components of the LAG-2/GLP-1/LAG-1 mediated signaling pathway, extragenic suppressors ( sog mutations) and enhancers ( ego mutations) of a weak glp-1 loss-of-function ( lf ) phenotype in the germline have been isolated. The studies in this thesis focus on three genes that interact with glp-1 in germline development: a suppressor, sog-10 , and two enhancers, ego-2 and ego-1 . The sog-10 mutation suppresses glp-1 ( lf ) germline defects and feminizes XX germlines. Genetic studies of sog-10 have narrowed the map position of the sog-10 gene to a well defined region of chromosome III, and phenotypic analyses of the sog-10 mutant phenotype indicates involvement of sog-10 in both germline proliferation and sex determination. An analysis of the phenotype of the ego-2 mutation has identified it as a strong enhancer of both the germline defect and the maternal effect lethality of glp-1 ( lf ) mutations. Mutations in ego-1 are able to enhance the germline phenotypes of both glp-1 ( lf ) and lag-1 ( lf ) mutations (E. Maine, unpublished). In addition, ego-1 mutants exhibit various germline abnormalities in a glp-1 wildtype background which indicate defects in germline mitosis and/or meiotic prophase. The gene responsible for the ego-1 germline phenotype was first detected as a small deletion on chromosome I associated with a UV-induced allele, ego-1 ( om84 ). The ego-1 gene was sequenced and confirmed to be the one responsible for the phenotypes in ego-1 mutants by PCR amplifying and sequencing this gene region from three strains containing ego-1 mutations, om84, om97 , and om71 . The predicted EGO-1 protein contains 1632 amino acids. Northern analysis identified a transcript of ∼5.2 kb which is developmentally regulated and primarily germline specific. ego-1 mRNA is detected in later larval and adult stages which is consistent with the germline specific phenotype of the ego-1 mutants. Sequencing of the ego-1 gene region revealed a second ego-1 related gene which is immediately adjacent to ego-1 on chromosome I. In addition, sequence comparison searches for EGO-1 homologs have identified two other proteins in C. elegans ; five proteins which have been identified as RNA-directed RNA polymerases (RdRP) in tomato, wheat, tobacco, petunia and Arabidopsis; and two uncharacterized proteins, one in Arabidopsis and one in the yeast Schizosaccharomyces pombe .
https://surface.syr.edu/bio_etd/45
oai:surface.syr.edu:bio_etd-1045
2010-10-26T13:00:36Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The oviposition response of the imported willow leaf beetle (Plagiodera versicolora): Does mother really know best?
Gross, Sherri Lynn
2000-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
James S. Coleman
Larry L. Wolf
Plagiodera versicolora
Oviposition
Willow leaf beetle
Ecology and Evolutionary Biology
Entomology
This dissertation addresses whether the oviposition responses of the female imported willow leaf beetles ( Plagiodera versicolora ) maximizes individual lifetime fitness. Rather than addressing this question on the scale of across habitats, species, or individuals, I chose to investigate the female's oviposition response on a plant as a function of different leaf stages differing in suitability as oviposition and larval growth sites.
The first series of observations examined the distribution of eggs in the field in relation to availability of leaf developmental stages. Females place a disproportionate number of eggs on the mature leaves compared to the younger ones. In the laboratory, larvae fed younger leaves grow faster and bigger than those fed mature leaves. This presents an interesting anomaly.
Experiments in the field examined the success of eggs placed on different leaf developmental stages. Traditionally, some measure of time, size, and/or survival is utilized to approximate the offspring's ultimate success and the mother's fitness. Using all these variables, I conclude that offspring success is most influenced by oviposition site in the egg stage. Eggs placed on mature and older leaves have a higher probability of hatching than those placed on other leaf stages. This higher hatching success more than compensates for the below average growth rate and survival experienced by early instars in comparison to their growth rate on younger leaf stages.
The final series of experiments examined factors that influence the probability of an egg hatching and attempted to quantify the advantage gained by eggs on mature leaves over other stages. Egg predators, Podisus spp., concentrate searches on the upper leaves, therefore mature leaves are less likely to be searched. Upper leaves experience lower humidities than those at the base of the plant. Eggs have a higher probability of hatching at higher humidities (>67%).
The results of this study question the assumptions upon which most oviposition studies are based. Questions include the validity of some estimates of fitness such as adult or larval size, testing of just the larval stage as an indicator of offspring success, and the focus on nutritional value of the host with the exclusion of other possible selective factors. These assumptions are shown not to hold true, bringing into question some of the conclusions from the existing body of literature.
https://surface.syr.edu/bio_etd/49
oai:surface.syr.edu:bio_etd-1046
2010-10-26T17:21:34Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Insulin-like growth factor stimulation of myoblast proliferation is specified by a serum activity, mitogenic competence factor
McWade, Francis Joseph
1998-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
James R. Florini
Richard Levy
Rapamycin
Pertussis
Mitogenic
Insulin-like growth factor
Myoblast proliferation
Biology
The stimulation of both myoblast proliferation and differentiation by Insulin-like growth factors (IGF) is well documented, but how each response is specified remains unclear. IGF stimulation of myoblast proliferation is transient, only lasting 36 hours. Cessation of proliferation does not require cell contact and does not involve a loss in the mitogenic activity of the IGF in the extracellular medium. IGF stimulation of proliferation can be restored by the addition of horse serum. Thus, a serum component serves a competence function for IGF stimulation of proliferation. The mitogenic competence activity of serum cannot be replaced by PDGF, bFGF, TGF-$\beta,$ HGF, transferrin, or fetuin and is not one of the known IGF binding proteins. Fractionation of horse serum by ammonium sulfate precipitation reveals that the proliferation competence and differentiation suppressing effects are both recovered in the 60-80% ammonium sulfate fraction. The 60-80% fraction has little or no mitogenic activity alone and thus functions as a competence factor or cofactor for IGF in stimulating myablast proliferation. I have named this activity Mitogenic Competence Factor (MCF).
To explore how MCF specifies IGF stimulation of myoblast proliferation, the effect of IGF on mediators of cell cycle progression and myogenesis in the absence and presence of MCF was tested. IGF and MCF were both required for phosphorylation of the retinoblastoma (Rb) protein, but IGF alone was sufficient to maintain levels of cyclin D and decrease levels the the cyclin dependent kinase (cdk) inhibitor p27$\rm\sp{Kip1}.$ The Extracellular Regulated Kinases (ERKs), and p70 S6 kinase (p70 S6K), have been implicated in mediating stimulation of proliferation by several extracellular factors. IGF and MCF were co-required to increase the activities of ERK1 and ERK2 kinases and p70 S6 kinase. Proliferation stimulated by IGF-I and MCF was inhibited in the presence of PD098059 and rapamycin, inhibitors of ERK and p70 S6 kinase pathways, respectively. Proliferation was also inhibited by the G protein inhibitor pertussis toxin and the PI-3 kinase inhibitor LY294002. IGF and MCF stimulation of Rb phosphorylation was almost completely inhibited by PD098059 and partially inhibited by pertussis toxin. Rapamycin and LY294002 inhibited Rb phosphorylation very slightly. Although all four compounds inhibited proliferation, myoblasts treated with PD098059 and pertussis toxin fused into myotubes, although to different extents. Thus, both the ERK or p70 S6K pathways appear to be involved in L6A1 myoblast proliferation stimulated by IGF-I and MCF but the ERK pathway may be important in dictating the choice between proliferation and differentiation, which may be related to its involvement in Rb phosphorylation.
https://surface.syr.edu/bio_etd/48
oai:surface.syr.edu:bio_etd-1047
2010-10-26T18:00:21Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Na stress and grazing: Molecular, physiological and growth responses on four Serengeti C4 grasses
Hamilton, Eugene William, III
1999-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Samuel J. McNaughton
Grazing
Growth responses
Serengeti
Soil sodium
Andropogan greenwayi
Sporobulus
Botany
Ecology and Evolutionary Biology
Life Sciences
Plant Sciences
The concentration of soil sodium (Na) is an important factor that influences species distribution in the Serengeti short-grass plains, Tanzania. Experiments involving soil Na treatments were conducted to detect physiological (photosynthetic and water relations) and molecular (heat shock proteins and organic solutes) adaptations. The species tested were Andropogon greenwayi and three species of Sporobulus, S. ioclados, S. kentrophyllus and S. spicatus . Each species grows on different concentrations of soil Na that vary over two orders of magnitude. Clipping experiments were conducted to investigate the trophic interaction of grasses and herbivores with respect to Na, a required micronutrient in C4 grasses, and an essential nutrient for herbivores.
Comparisons of all four species detected short-term physiological and molecular responses to Na treatments (0, 100 and 400 m.M Na) that correlated with their field soil Na concentrations. S. kentrophyllus and S. spicatus exhibited rapid molecular induction of heat-shock proteins (Hsp) in response to experimental soil Na treatments (24 hr). Photosynthetic tolerance to Na was positively correlated with field soil Na concentrations, and Hsp induction was clearly associated with photosynthetic tolerance.
Long term (6 weeks) responses of the four species to Na treatment and clipping supported trends observed in the short term responses to Na. Species that occur on low soil salinity in the field did not survive past week one when treated with 400 m.M Na and exhibited significant reductions in biomass when treated with 100 MM Na. Reduced biomass, increased shoot tissue Na concentrations and Na tolerance correlated with the Na concentrations found in field soil Na concentrations. Analysis of rinse samples from leaf tissue showed that S. spicatus exudes Na salts. Clipping reduced the amount of root and shoot produced by S. spicatus and increased the total amount of Na available for herbivore consumption in all species. The results suggest that Na tolerant specie exhibit molecular responses that protect physiological processes and therefore sustain growth. Soil Na and grazing influence each species differently and these differences maintain the observed species distribution. This suggests that Na and grazing tolerance may have coevolved in the Serengeti ecosystem through a grass-grazer trophic interaction.
https://surface.syr.edu/bio_etd/47
oai:surface.syr.edu:bio_etd-1048
2010-10-26T18:46:26Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Mate choice in the presence of sperm parasites
Wisniewski, Timothy James
1998-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Larry L. Wolf
Sexual imprinting
Poeciliopsis
Mate choice
Sperm parasites
Biology
Zoology
The present study looks at the interaction between species recognition and mate choice within a set of sympatric species. In Mexico, a complex of species of internally fertilizing, viviparous fishes (Genus Poeciliopsis ) live sympatrically with each other and with an array of clonal females. The clones depend on inseminations from males of the various species to initiate embryogenesis. The clones are considered "sperm parasites" since none of the DNA from the males is incorporated into the offspring. Considering the costs incurred by males in time, energy, inter-male aggression, exposure to predation and, of course, the wasted gametic material, the question investigated was why males from these species court and mate with clonal females? Many hypotheses were considered, including whether males are allocating mating effort in proportion to the probability of achieving sexual offspring, or whether males are deriving other fitness benefits from clonal inseminations, such as discarding old sperm, gaining valuable practice or increasing their attractiveness to sexual females. The hypothesis found to be most consistent with the evidence was the one that tested whether males must interact and mate with clonal females while learning to discriminate between clonal and conspecific females. It was demonstrated that males can learn clonal discrimination only if they are given extensive experiences with a sexual female first and then given extensive experience with a clonal female. The reverse order of experiences did not result in males exhibiting clonal discrimination. Further, it was shown that males more rapidly learned cues associated with sexual females than they learned cues associated with clonal discrimination. Finally, a model of the proximate mechanism involved in learning clonal discrimination was proposed.
https://surface.syr.edu/bio_etd/46
oai:surface.syr.edu:bio_etd-1049
2010-10-28T14:16:50Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Structural and functional studies on the catalytic mechanism of glucose 6-phosphate dehydrogenase from Leuconostoc mesenteroides
Cosgrove, Michael Scott
1998-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
H. Richard Levy
Catalytic
Glucose 6-phosphate dehydrogenase
Phosphate dehydrogenase
Leuconostoc mesenteroides
Biochemistry
Biophysics
A protein structure-function-based approach was used to study the catalytic mechanism of glucose 6-phosphate dehydrogenase from Leuconostoc mesenteroides . Site directed mutagenesis was used to replace Asp-177, His-178, and His-240 with asparagine, and the structural and functional characteristics of the mutated enzymes were assessed. The results show that His-240 is the general base that abstracts a proton from the C1-OH of glucose 6-phosphate (G6P), and that the charge supplied by Asp-177 functions to make His-240 a better general base by affecting its pK a , forming a His-Asp catalytic dyad. The function of His-178 is to bind the phosphate moiety of the G6P.
The three-dimensional structure of the first ternary complex for G6PD is reported. The results indicate that upon formation of the ternary complex, the enzyme becomes more compact bringing several residues into close contact with the substrate presumably to provide additional binding energy that is used to help stabilize the transition state. Additionally, the binding site for a ternary complex-specific water molecule is observed that may help to regenerate the enzyme after oxidation of substrate. Binding of G6P and ordering His-178 appears to be necessary in forming this water-binding site.
Finally, the 500 MHz 1 H-NMR chemical shifts of the C[varepsilon] and Cδ protons of His-240 were assigned by comparing spectra of the wild-type and H240N G6PDs. This information was used to determine the pK a of His-240 within the three dimensional structure of this 109 kD enzyme. The results indicate that 1 H-NMR can be a useful tool for functional analysis of some moderate-to-large sized proteins.
https://surface.syr.edu/bio_etd/51
oai:surface.syr.edu:bio_etd-1050
2010-10-28T16:59:17Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Three-dimensional structure determination of a subunit of the HIV-1 packaging signal
Pappalardo, Lucia
1998-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Philip N. Borer
Immune deficiency
SL3
Nucleocapsid
HIV-1
Packaging signal
Biophysics
The Human Immunodeficiency Viruses (HIV), responsible for the Acquired Immunodeficiency Syndrome (AIDS), are members of the retroviral family. The RNA genomic material is packaged in new infective virions during the last stage of the retroviral life cycle, a process common to all retroviruses. Only full-length retroviral RNA genome is packaged efficiently. A relatively small region of the HIV-1 genome has been found to be critical for efficient packaging of the virus. This region, known as the "packaging signal", $\Psi$, folds in a secondary structure composed of three to four stem-loop hairpins, named SL1 to SL4. No X-ray or NMR three-dimensional structures have been reported for retroviral packaging signals or for their subunits. Structural studies of the "packaging signal" may provide clues about the HIV packaging mechanism and suggest tools to defeat AIDS.
This thesis is based upon the three-dimensional structure determination of a 20-nucleotide stem-loop RNA SL3 subunit of the HIV-1 packaging signal, using Nuclear Magnetic Resonance techniques. NMR is currently the only technique that allows the determination of high resolution three-dimensional structures of biomolecules in solution under nearly physiological conditions.
SL3 RNA was synthesized by transcription of a synthetic DNA template by the enzyme T7 RNA polymerase. UV melting curves, native gel electrophoresis and 1D H$\sb2$O imino proton spectra were analyzed to confirm the secondary structure of the SL3 molecule. Two-dimensional COSY-type, NOESY, heteronuclear $\sp1$H-$\sp{31}$P-COSY NMR experiments were acquired, processed and analyzed to assign resonances and determine distance and dihedral angle constraints. The NMR constraints were used to refine the three-dimensional structure of SL3 by running restrained Molecular Dynamics calculations.
The final structure of SL3 has revealed an A-form stem and a quite flexible GGAG tetraloop with the second (G10) and fourth (G12) bases extruded from the normal stacking arrangement. The H-bonding loci of G10, A11, and G12 are unoccupied in the free RNA structure and exposed to the solvent available to be easily recognized by possible binding agents. In collaboration with Dr. Summers (UMBC) we have shown that SL3 forms a stable 1:1 complex with the nucleocapsid NCp7, and determined the three-dimensional structure of the SL3-NCp7 complex.
https://surface.syr.edu/bio_etd/50
oai:surface.syr.edu:bio_etd-1051
2010-10-29T16:15:46Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Effects of elevated CO(2) on plant-grazer interactions
Wilsey, Brian John
1995-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Samuel J. McNaughton
carbon dioxide
global change
herbivory
Ecology
Range management
Botany
Ecology and Evolutionary Biology
Atmospheric carbon dioxide levels have been increasing since the beginning of the industrial age, and are expected to reach 700 ppm during the mid- to late 21st century. Most research on plant response to elevated C02 has centered on plant processes in isolation from effects of higher trophic levels, and studies on the effects of elevated CO$\sb2$ on plant-herbivore interaction have concentrated on insects. The objectives of my dissertation were (1) to evaluate the possible effects of elevated CO$\sb2$ on plant-mammal interactions, and (2) to determine whether species from Yellowstone National Park, Flooding Pampa of Argentina, and Serengeti of Tanzania responded similarly to elevated CO$\sb2$ and whether responses were dependent on the mode of photosynthesis (C$\sb3$ or C$\sb4$) of dominant species. To test these objectives, plant species were grown in growth chambers at Syracuse University under ambient (350-370 ppm) and elevated (700 ppm) atmospheric CO$\sb2$, and under clipped and unclipped conditions.
Species from the 3 ecosystems responded differently to elevated CO$\sb2$. Significant increases in biomass occurred only in Yellowstone species (all C$\sb3$ plants), and increases occurred mostly in crowns and roots (storage organs). Aboveground biomass, which is the portion that is consumed by grazing mammals, was not affected by elevated CO$\sb2$. There was, in general, no interaction between clipping and CO$\sb2$, and this suggests that defoliation will not affect how plants respond to elevated CO$\sb2$. Reductions in leaf N (an index of forage quality) occurred only in Yellowstone and Flooding Pampas species; no reduction in biomass and nutrient concentration was found in Serengeti species. Since ungulates can not regulate time spent feeding (as insects do), and since passage time is inversely related to food quality, grazing mammals in temperate grasslands may be negatively affected by elevated CO$\sb2$.
Data on plant response to elevated CO$\sb2$ was also compiled from the literature, and responses were compared among biomes. Grassland species had smaller responses than forest species. Biome origin accounted for more of the variation in plant response than did mode of photosynthesis or growth form. Thus, biome origin was also a good predictor of plant species response to elevated CO$\sb2$.
https://surface.syr.edu/bio_etd/52
oai:surface.syr.edu:bio_etd-1052
2010-11-03T13:45:21Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Fire effects in the grasslands of Yellowstone National Park
Tracy, Benjamin Franklin
1996-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Samuel J. McNaughton
nutrient cycling
Lupinus sericeus
ungulate
grazing
ecology
Cervus elaphus
Ecology and Evolutionary Biology
Fires burned almost 45% of Yellowstone National Park in 1988. The goal of this dissertation research was to learn how fire would affect grazing by ungulates, plant production and nutrient cycling mainly in the sagebrush grasslands of Yellowstone National Park. Winter, transitional and summer range for ungulates provided the study locations for this project. Findings from this study showed that burning can increase aboveground production in the Yellowstone grasslands. Increases may last up to three years after fire but only on winter range. There were no discernable effects of fire on productivity in summer and transitional ranges. Burned soils on winter range typically held more water and had greater concentrations of phosphorus and ammonium, all of which may contribute to increases in aboveground production. Ungulates consumed more forage in burned areas, but only on winter range in the spring of 1991. Immediately following fire, nutrients become concentrated in ungulate forage--up to 3$\times$ higher than forage on unburned areas. Fires, however, may diminish this nutritional benefit to ungulates because burning also increases the aboveground biomass of unpalatable forbs like lupine (Lupinus sericeus).
Effects of the fire were also compared between a lodgepole pine forest and adjacent grassland. Burning greatly increased the production of grasses growing in the understorey of a lodgepole pine forest even five years after fire. Elk, however, consumed little of this productive forage suggesting that forage quality rather than quantity may influence grazing behavior in some areas of Yellowstone. Data from this dissertation suggest that ungulate activities may influence the productivity of Yellowstone grasslands more than the general physical environment. Overall, the Yellowstone grasslands appear very resilient to burning. Fire effects may require further evaluation during dryer periods to achieve a better understanding of fire's role in the Yellowstone ecosystem.
https://surface.syr.edu/bio_etd/54
oai:surface.syr.edu:bio_etd-1053
2010-11-04T11:33:25Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Identification and molecular characterization of a type-A histone acetyltransferase from Tetrahymena thermophila
Brownell, James Edward
1996-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
C. David Allis
chromatin
posttranslational acetylation
Cellular biology
Molecular biology
Cell Biology
Gene expression in eukaryotes requires the coordination of at least three events; the binding of activators and co-activators to DNA enhancer elements, the recruitment of the basal transcription apparatus to DNA promoter elements, and the relief of repressive chromatin structure. Indirect evidence has long suggested that posttranslational acetylation of the four core histones, the primary protein components of chromatin, alters chromatin structure and potentiates transcription. In this dissertation, I describe the isolation and cloning of a catalytically active subunit (p55) of type-A histone acetyltransferase (HAT) from Tetrahymena, an enzyme that catalyzes transcription-related acetylation.
The sequence of Tetrahymena p55 reveals striking homology to the highly conserved yeast co-activator GCNS, which, like p55, possesses intrinsic histone acetyltransferase activity. Amino acid sequence analyses of Tetrahymena p55 and its counterparts in yeast, Drosophila, and humans reveals several conserved domains, including single copies of the bromodomain. The bromodomain is a highly conserved motif found in an extremely limited number of polypeptides with related functions in activated gene expression. Thus the bromodomain may provide a mechanism for targeting histone acetyltransferase and related activities to individual genes within specific chromatin domains.
Analyses of Tetrahymena macronuclear extracts suggest that p55 is a component of a multimeric enzyme complex with a molecular mass of approximately 220 kDa. This value agrees closely with the theoretical mass of a predicted complex in yeast containing the gene products of GCN5, ADA1, ADA2, ADA3, and ADA5, suggesting that both the Tetrahymena and yeast HAT A activities are composed of similar heteromeric complexes. Together with independent genetic and biochemical evidence implicating Gcn5p, Ada1p, Ada2p, Ada3p, and Ada5p as components of a putative transcriptional adaptor complex necessary for the full activity of several genes in yeast, these findings establish a direct link between histone acetyltransferase activity and factors required for activated gene expression. Moreover, defining a biochemical function for yGcn5p and its homologs as HATs implies that other transcriptional co-activators may similarly affect gene expression by directly engaging and modifying chromatin components.
https://surface.syr.edu/bio_etd/53
oai:surface.syr.edu:bio_etd-1054
2010-11-04T18:27:28Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Genetic and molecular characterization of genes involved in Caenorhabditis elegans germ line development
Qiao, Li
1997-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Eleanor Maine
signal transduction
@ego
Cell Biology
Genetics
Cell-cell interactions control many cell fate choices during the development of multicellular organisms. In Caenorhabditis elegans, the distal tip cell (DTC) regulates the mitosis/meiosis choices of the germ cells. This cell signaling requires a putative signal from the DTC, encoded by the lag-2 gene, a putative receptor in the germline, encoded by the glp-1 gene, and at least one downstream effector encoded by the lag-1 gene. To identify additional genes involved in the glp-1 signaling pathway, genetic suppressors and enhancers of glp-1 mutations have been isolated; they are named sog (suppressor of glp-1) and ego (enhancer of glp-1) mutations.
My dissertation studies involve the genetic characterization of a sog-10 mutation and a set of ego mutations, and the molecular cloning of the ego-3 gene. The sog-10 mutation suppresses glp-1 (lf) germline defects and feminizes XX germline, indicating its role in both germline proliferation and sex determination. lag-1 and glp-4 are known genes whose mutations can enhance glp-1(lf). lag-1 mutations affect both germline proliferation and embryogenesis, while glp-4 mutations affect both germline mitosis and oogenesis. Finally, ego-3 mutations have multiple phenotypes indicating its role in multiple aspects of germline development as well as in somatic tissues. The predicted ego-3 gene encodes a novel protein. Its weak similarity with several enzymes suggests EGO-3 may be involved in protein-protein interactions. These studies suggest that these suppressors and enhancers of glp-1 may have functions in multiple aspects of development, as does glp-1 itself.
https://surface.syr.edu/bio_etd/57
oai:surface.syr.edu:bio_etd-1055
2010-11-05T15:25:12Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Identification and characterization of Hhp1p, a heterochromatin-associated protein in Tetrahymena thermophila
Huang, Hui
1997-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
C. David Allis
Hhp1p
heterochromatin-associated protein in Tetrahymena
Cell Biology
Molecular Biology
The identification and characterization of heterochromatin-associated proteins has greatly improved our understanding of heterochromatin assembly and dynamics in eukaryotes. In this dissertation, I identified and cloned a 28 KDa polypeptide, Hhp1p, from Tetrahymena macronuclei that possesses many features of the heterochromatin-associated protein HP1 from Drosophila. Like other HP1-like proteins, Hhp1p contains both a chromo domain and a chromo shadow domain. However, a short domain between the chromo domain and its shadow domain in Hhp1p displays some features of linker histone H1. Thus, this protein is being referred to as Hhp1p (for H1/HP1-like protein). In Tetrahymena, Hhp1p specifically localizes in transcriptionally active macronuclei where Hhp1p displays a punctate staining pattern.
To further investigate Hhp1p function, I have disrupted all of the expressed copies of the HHP1 gene in somatic macronuclei. HHP1 knock-out ($\Delta$HHP1) cells grow at normal rates, demonstrating that Hhp1p is not essential in Tetrahymena. However, the survival rate of $\Delta$HHP1 cells is markedly reduced compared to that of wildtype cells during prolonged starvation. Upon starvation, $\Delta$HHP1 cells display a roughly 1.5-fold increase in total macronuclear size along with a reduction in the size of the electron-dense chromatin bodies. In contrast, in wildtype cells, Hhp1p becomes hyperphosphorylated during prolonged starvation concomitant with an increase in chromatin body size. In addition, the activation of two starvation-induced genes is reduced in $\Delta$HHP1 cells, while transcription rates of growth-related genes are comparable to wildtype cells. These results suggest that Hhp1p functions in heterochromatin assembly and/or maintenance, and it is required for the appropriate expression of certain genes in physiological states such as starvation.
To investigate the potential relationship between Hhp1p and linker histone H1, a double knockout strain was created and studied. It appears that knocking out H1 in $\Delta$HHP1 cells partially rescues some wildtype phenotypes, suggesting that H1 may function as a suppressor of HHP1 in certain physiological states. These results, together with preliminary biochemical evidence that Hhp1:H1 products are detected in cross-linked macronuclei, suggest that linker histone H1 may interact with Hhp1p in vivo.
https://surface.syr.edu/bio_etd/56
oai:surface.syr.edu:bio_etd-1056
2010-11-05T18:49:25Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Integrating prior experiences into behavioral decisions: The effect of prior fighting experiences on the contest behavior of Rivulus marmoratus, a hermaphroditic fish
Hsu, Yuying
1997-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Larry L. Wolf
Rivulus marmoratus
hermaphroditic fish
behavioral decisions
fighting experiences
Behavior and Ethology
This research is concerned with whether animals integrate multiple prior fighting experiences into behavioral decisions in subsequent contests. The effect of prior fighting experience on behavior of contestants at different stages of a contest was examined. Further, a probability model of integrating fighting experiences into current aggressive behavior was proposed and tested.
Rivulus marmoratus, an internally self-fertilization hermaphroditic fish, has been repeatedly observed to display aggressive behavior in both field and laboratory, and thus was selected for this research. However, their contest behavior has never been described before. This project started with observing and comparing their behavior in both dyadic contests and mirror tests. Different clones did not behave differently in either test, but the two tests yielded different conclusions regarding the aggressiveness of different clones.
The fish were given different combinations of two prior fighting experiences to investigate how different prior fighting experiences were evaluated. The major conclusions are: (1) the effect of the most recent experience was more pronounced than the effect of the penultimate fighting experience; (2) the primary effect of the two experiences did not come from a potential damage asymmetry due to different prior fighting experiences; and (3) the changes in contest behaviors were consistent with the hypothesis that prior fighting experiences influence how an individual perceives its relative fighting ability but do not influence its actual relative fighting ability in subsequent contests. These results were used to develop a model that combines the effect of past experiences of two individuals to predict the probability of winning a dyadic contest. The model parameters were estimated with experimental data and correctly predicted the outcome of contests. These results indicate that the effects from past fighting experiences are accumulated systematically and that the relative past experiences of two contestants determine the outcome of their interaction when other factors are controlled. They also suggest that the fish offers a model system for examining how experiences are integrated into current decisions. Although the model was developed for animal contests, its general approach to integrating prior experiences for different individuals might be useful for understanding other interactions that are affected by learning.
https://surface.syr.edu/bio_etd/55
oai:surface.syr.edu:bio_etd-1057
2010-11-08T15:27:51Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Mapping sites of deposition-related acetylation in newly synthesized H3 and H4 during chromatin assembly and maturation in vitro and in vivo
Sobel, Richard E.
1997-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
C. David Allis
Drosophila melanogaster
Tetrahymena
Acetylation
Chromatin
Biochemistry
Biology
The amino termini of core histones H2A, H2B, H3 and H4 contain conserved lysine residues which are transiently modified by the addition and removal of acetyl groups. Non-random acetylation of these lysines in histone H4 is associated with histone deposition (Chicoine, et al. 1986), dosage compensation in Drosophila (Turner, et al. 1992) and mating efficiency in Saccharomyces (Park and Szostak 1990, Megee, et al. 1990). In most eukaryotes, H4 is "blocked" at its amino terminus preventing direct analysis of acetylation sites by microsequencing. Application of a new deblocking procedure (Wellner, et al. 1990) allowed me to examine the deposition-related sites of acetylation of new H3 and H4 during chromatin assembly and maturation in a wide range of eukaryotes.
A crude cytosolic histone acetyltransferase (HAT-type B) from Drosophila embryos was used to in vitro label H4 purified from either Tetrahymena or Drosophila. Deblocking and microsequencing showed that acetylation occurred almost exclusively on lysine 11/12 of both substrates. Similar results have been obtained with homogeneous HATB from yeast (Parthun, et al. 1996). In agreement with results previously obtained in Tetrahymena H4 (Chicoine, et al. 1986), newly synthesized and deposited H4 from Drosophila and HeLa cells was diacetylated exclusively at lysines 5 and 12, demonstrating conservation of this modification. A novel, simple method to pulse-label, extract and purify newly synthesized yeast histones in vivo showed that in contrast to the organisms above, new yeast H4 was deposited primarily as a monoacetylated form.
New H3 from all four organisms failed to show conservation of deposition-related acetylation sites. Tetrahymena H3 was deposited as mono- and diacetylated forms acetylated predominantly at lysine 9 and lysines 9 and 14. New Drosophila H3 was mono- and diacetylated with lysine 23 and lysines 23 and 14 being the preferred sites, respectively. New yeast H3 was also deposited as acetylated forms with lysines 9 and 27 being the major sites of acetylation. However, in HeLa no acetylation of deposition-related H3 is observed.
Deacetylation of H4 during chromatin maturation was also examined.
https://surface.syr.edu/bio_etd/65
oai:surface.syr.edu:bio_etd-1059
2010-11-08T19:32:01Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The association of space use and aggressive behavior in nesting male dwarf gourami (Colisa lalia Hamilton-Buchanan)
Tlusty, Michael Urbancic
1996-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Larry L. Wolf
game theory
territorial
nest defense
intersexual
Zoology
Ecology
Zoology
Animals that defend a focal point generally decrease their aggressive frequency to intruders at increasing distances from the nest. Aggressive frequency is comprised of two sequential behaviors, the use of space and aggressive tendency. The resident has to first approach the intruder (space use) before it can attack (aggressive tendency). This study examined how manipulations of the net benefit of a contest influenced a nesting male dwarf gourami's (Colisa lalia) spatial and aggressive response to an intruder. This study was conducted on single males in a laboratory. This reduced the complexity of this problem by eliminating male-male interactions, and limited a male's spatial behavior to a linear response. The net benefit of a contest was decreased by limiting the visual distance, by increasing the number of females in the local environment, and by adding eggs to the nest. Each of these manipulations increased the cost of leaving the nest. The net benefit of a contest was increased via an increase in benefits by presenting an intruder at decreasing distances from the nest, presenting different quality intruders (males vs. females, small vs. large females).
Each of these manipulations influenced the spatial behavior of the fish. Males tended to spend more time near the nest when the cost of the encounter increased or the value of chasing an intruder decreased. This decrease in space use resulted in a decrease in the frequency of aggressive behavior. Aggressive tendency also decreased with increasing cost or decreasing benefit, although not as consistently as the changes in spatial behavior. Escalated aggressive responses rarely changed with experimental manipulation, as the test intruders could not respond appropriately to the resident's attacks. The use of space is a critical factor in determining the aggressive response of a resident to an intruder, and future studies need to be more rigorous in the study of space use and aggression as a two component sequential behavioral process.
https://surface.syr.edu/bio_etd/63
oai:surface.syr.edu:bio_etd-1058
2010-11-08T18:48:31Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Study of the S. cerevisiae mitochondrial chaperonin, Hsp60, and the identification and characterization of SCS1/RTS1, a high-copy suppressor of Hsp60(ts)
Shu, Youmin
1996-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Richard L. Hallberg
SCS1/RTS1
molecular chaperone
Saccharomyces cerevisiae
Molecular biology
Cellular biology
Genetics
Molecular Genetics
In order to study the functions of the S. cerevisiae mitochondrial chaperonin, Hsp60, I have generated 25 temperature-sensitive lethal mutant alleles of the HSP60 gene. Mutant strains expressing these alleles manifested a wide variety of abnormal phenotypes, both at permissive and non-permissive temperatures.
To identify proteins functionally related to Hsp60, I identified genes which, when over-expressed, suppressed the temperature sensitive phenotype of cells expressing a HSP60 mutant allele. One particular gene, RTS1, was characterized in detail. RTS1 is not an essential gene, but RTS1-null strains are temperature sensitive. RTS1 encodes an 86 kDa cytoplasmic protein which is heavily modified in vivo. Over-expression of RTS1 had significant effects on the cellular levels of mRNAs encoding the heat-inducible (heat shock) proteins Cpn10 and Mge1p, two other mitochondrial protein co-chaperones, but not on mRNAs encoding a number of other mitochondrial or cytosolic heat shock proteins analyzed. It was this property, presumably, that enabled overexpression of RTS1 to suppress mutant alleles of HSP60.
Besides defining the role of RTS1 in regulating the expression of mitochondrial chaperones, I also identified its role in cell cycle control. First, CLB2, a gene encoding a regulatory subunit of protein kinase, was identified as a high-copy suppressor of the RTS1-null strain. The activity of Clb2 is required for controlling cell progression from G2 to M phase. Second, RTS1-null cells were observed to have a typical cell division cycle (cdc) mutant phenotype at high temperatures. Finally the RTS1 protein was found modified in a cell cycle dependent manner, implying that the specific activity of RTS1 is cell cycle dependent.
Human and rabbit homologs of RTS1 were identified by others and shown to encode regulatory (B) subunits of protein phosphatase 2A. As another S. cerevisiae B regulatory subunit gene, CDC55, had been previously described and as PP2A is essential for cell viability, I created a strain null for both genes. This strain was viable, suggesting there are more B subunit genes remaining to be identified. Analyses of this strain also showed that RTS1 and CDC55 play distinctly different roles in vivo.
https://surface.syr.edu/bio_etd/64
oai:surface.syr.edu:bio_etd-1060
2010-11-09T16:55:57Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Structure-Function Relationship of Glucose 6-Phosphate Dehydrogenase from Leuconostoc Mesenteroides
Haghighi, Bahram
1982-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
H. Richard Levy
Lysine
ligand binding
conformational transitions
fluorescence reporter group
extrinsic probe
Biochemistry
The previous procedure for the isolation of glucose 6-phosphate dehydrogenase (G6PD) from Leuconostoc mesenteroides was modified to allow the preparation of large quantities of homogeneous enzyme suitable for structural studies. The enzyme was shown to consist of two identical subunits containing N-terminal valine and C-terminal glycine.
Three aspects were examined that probe the relationship between structure and function of L. mesenteroides G6PD. First, the enzyme was covalently labelled with a phosphopyridoxyl group at a unique lysine residue. Previous work had suggested that this lysine is located at the active site and that it participates in binding G6P {Milhausen and Levy, Eur., J. Biochem. 50 (1975), 453-461}. Two pyridoxyl peptides, DIIA and DIIB, were isolated from the tryptic digest of the modified enzyme. The amino acid sequence of each peptide revealed that the sequence of the pyridoxyl binding site was Phe-Leu-Leu-Lys(Pxy)-Ser-Pro-Ser-Tyr-(Asp, Val)-Lys. This is the first report of sequence information from the active site of any G6PD.
Support for the role of the modified lysine is provided in the second part of this dissertation, in which three fluorescent probes were exploited to provide information on ligand binding and conformational transitions that the enzyme undergoes.
First, the intrinsic protein fluorescence was quenched by binding various ligands to the enzyme. These measurements permitted the monitoring of overall conformational changes of the enzyme upon ligand binding and calculation of binding constants.
Second, the covalently bound phosphopyridoxyl group in the modified enzyme served as a fluorescence reporter group to monitor the conformational changes produced by NAD('+) or NADP('+) and to calculate their binding constants. The results of these experiments showed that pyridoxylation promotes a conformational change similar to that produced by NAD('+) or G6P (see below). Binding of NAD('+) to pyridoxyl enzyme is accompanied by a smaller conformational change than that seen when it binds to the native enzyme. Conversely, NADP('+) binding leads to a larger conformational change in the pyridoxylated than in the native enzyme. This was supported by the fact that the dissociation constant for NAD('+) is lower, and for NADP('+) is higher, for pyridoxyl enzyme than for native enzyme.
Third, the fluorescence of S-NADPH was used as an extrinsic probe of the coenzyme binding region of the enzyme. The fluorescence properties of S-NADPH are reported for the first time. ...
https://surface.syr.edu/bio_etd/62
oai:surface.syr.edu:bio_etd-1061
2010-11-10T17:26:51Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Conformational Properties of Polyribonucleic Acids: Studies with Yeast Transfer-RNA, Synthetic Homopolyribonucleotides and Rabbit Globin Messenger-RNA
Vamvakopoulos, Nikos Constantinos
1980-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
John Vournakis
Biopolymers
Ethidium bromide
Conformation
Magnesium
Environmental factors
Biophysics
The effect of various physical parameters on the conformation of three classes of biopolymers in solution, was investigated under equilibrium conditions. Using ethidium bromide as a probe, a description of both the conformational properties and conformational transitions occurring in transfer RNA became possible. A novel class of interactions between ethidium bromide and tRNA was revealed. This interaction occurred when the probe reacted with a solution of tRNA containing 5mM magnesium ions. Such an interaction can be considered as a useful index for the occurrence of tertiary interactions in tRNA in solution.
The effect of magnesium and manganese ions on the conformation of the hydrogen bonded hybrids polyribocytidylate(.)oligodeoxyguanylate and poly2'-O-methylcytidylate(.)oligodeoxyguanylate was studied. The results were compared with the ability of the hybrids to serve as efficient substrates for avian myeloblastosis virus DNA polymerase under the same solution conditions. It was found by this comparison that the conformation of these substrates as affected by the aforementioned ionic species constitutes an important parameter controlling enzymatic activity.
The effect of environmental factors (such as heat, pH, ionic composition, etc.) as potential determinants of the conformation of rabbit globin messenger RNA (mixture of two messenger RNA species) was investigated. Findings provided evidence for the occurrence of major conformational changes in mRNA. From a macroscopic viewpoint both acidic pH and magnesium ions strongly enhanced the conformation and stability of the message.
The conformation of this particular class of biological macromolecules was studied in further detail, at the nucleotide level, for each mRNA species. The most important finding from this study was the strong susceptibility, to the single strand specific enzymatic probe used in these experiments, of the initiator region in (beta) globin mRNA as opposed to the very limited susceptibility of the analogous region of (alpha) globin mRNA by the enzyme. This finding can provide a molecular interpretation to the more efficient initiation of protein synthesis by rabbit (beta) globin mRNA chain than by rabbit (alpha) chain.
A search for 18s ribosomal RNA binding sites in the 5'-non coding region of rabbit globin mRNA was undertaken in an attempt to reveal similarities in the mode of initiation of protein biosynthesis among eucaryotic and procaryotic systems. Results indicated an absence of interacting sequences with 18s rRNA in the 5'-non coding region of (beta) globin mRNA. This result is consistent with the lack of protection against enzymatic hydrolysis of eucaryotic mRNA sequences in a procaryotic ribosome binding system observed by other investigators, and reveals one more possible difference between procaryotic and eucaryotic mechanisms of initiation of protein synthesis.
https://surface.syr.edu/bio_etd/61
oai:surface.syr.edu:bio_etd-1062
2010-11-12T15:41:00Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Synthesis Of 1-Fluoro-3-Hydroxyacetone Phosphate And Its Effects On Glycerol-3-Phosphate Dehydrogenase
Silverman, Joan Lee Basloe
1973-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Thomas P. Fondy
Biochemistry
Biochemistry
Discusses methods, toxicity, antitumor studies, etc.
https://surface.syr.edu/bio_etd/60
oai:surface.syr.edu:bio_etd-1064
2010-11-12T17:29:15Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The role of nitrogen and leaf development in plant-insect interactions
Wait, D. Alexander
1997-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
James S. Coleman
herbivory
Biology
Botany
Increased nitrogen (N) availability to plants via fertilization, atmospheric deposition, or land use practices generally increases plant growth and tissue N concentration, and decreases tissue secondary chemical concentrations. Consequently, "N fertilized" plants should provide a greater quantity of higher quality leaf resources to insect herbivores. In turn, fertilized plants should experience greater levels of herbivory and proportionally more damage than unfertilized plants. Yet, we have had limited success in predicting both the effects of fertilization on plant tissue quality and quantity for herbivores, and the impact of herbivory on plant growth. The primary objective of this research was to increase our understanding of the effects of N fertilization on plant-insect interactions by determining the plant processes that are affected by N, and in turn, regulate herbivore responses.
The research addressed the following questions: (1) does fertilization affect the biochemical quality of leaves for herbivores through changes in leaf development? (2) does fertilization increase the availability of leaf resources for herbivores through changes in leaf development? and (3) how does fertilization influence the impact of herbivore damage on plant growth? My experimental system included: saplings of an economically and ecologically important tree species, Populus deltoides Bart.; varying N supply rates (concentration x time); two folivorous beetles, Chrysomela scripta F. and Plagiodera versicolora Laich. (Coleoptera: Chrysomelidae); and artificial defoliation.
The results demonstrated that: (1) foliage biochemical quality is mostly controlled by an interaction between fertilization and leaf development; (2) fertilization increases the availability of leaf resources that are suitable for feeding by insects to a greater extent than plant productivity; and (3) fertilization influences the effect of defoliation on plant growth by affecting the trajectory of growth through time. These results were independent of the degree to which fertilization affected plant growth rate and leaf N concentration. This research illustrates how N fertilization, through changes in leaf development and plant growth, regulates plant-insect interactions, and thus provides mechanistic predictions of the impact of fertilization on plants and herbivores.
https://surface.syr.edu/bio_etd/58
oai:surface.syr.edu:bio_etd-1063
2010-11-12T17:18:29Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
The role of metapopulation dynamics in the persistence of rare species
Latham, Brenda Price
1997-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
William T. Starmer
Dipodascus starmeri
Pichia cactophila
Stenocereus gummosus
cacti
yeast
Drosophila mojavensis
Ecology and Evolutionary Biology
The relative importance of factors determining the distribution of a common and a rarer cactophilic yeast inhabiting the stem necroses of Stenocereus gummosus were investigated. In nature, Candida ingens (now known as Dipodascus starmeri), occurs in a low but steady proportion of rots, while Pichia cactophila is both locally abundant and widespread. Metapopulation theory can be used to understand those types of rarity in which local populations are distributed across patches of habitat.
Variability in species occurrence in nature was found primarily between plants in a locality. Yeast community composition and physiological abilities were persistent across time. Region, seasonality, and negative effects of other yeast species are not important determinants of the presence of C. ingens in S. gummosus necroses. Candida ingens was more frequent in rots of neutral pH.
Relative attractiveness, vector acquisition and deposition, and mortality of yeasts during transport by D. mojavensis were investigated. Flies are attracted by the necrosis-initiating bacterium and P. cactophila, but C. ingens was less attractive. Mortality did not differ between yeast species. Acquisition and deposition were generally proportionate to species presence in the substrate, however P. cactophila was deposited more variably than C. ingens.
Potential sources of differences in colonization success between a rarer and a common yeast species, including the presence of other species, cactus chemistry, and bacterial species presence were investigated. Colonization by C. ingens is unaffected by P. cactophila. Candida ingens' carrying capacity is highest when it arrives early and in large numbers. Variation in stem chemical composition influences the carrying capacity of P. cactophila but not C. ingens; initial growth rates of both yeasts are unaffected. The rate of increase and the carrying capacity of both yeasts are significantly affected by the bacterial species initiating the rot.
A simulation model detected small effects of differential attractiveness and cactus chemistry on the distribution of C. ingens. The proportion of patches inhabited by C. ingens was more sensitive to growth conditions, especially carrying capacity, under different bacterial regimes. Bacterial succession, along with deposition variability, are likely to contribute to the metapopulation persistence of a rarer yeast species.
https://surface.syr.edu/bio_etd/59
oai:surface.syr.edu:bio_etd-1065
2010-11-16T21:54:30Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Biomass Turnover, Energy Balance, and Interpopulation Variation in the Stream Limpet, Ferrissia Rivularis (Say), With Special Reference to Respiration, Growth, and Fecundity
Burky, Albert John
1969-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
W. D. Russell-Hunter
New York State
Life-cycle
Reverse respiratory acclimation
Freshwater environments
Zoology
This investigation extends from 1965 through 1968 and involves 42,926 individual measurements on 19,479 limpets from populations of F. rivularis in the Canandaigua Outlet at Alloway, New York (AL)* and in Black Creek at Cleveland, New York (BC)*, with some comparative results from a population in Chittenango Creek at Cazenovia, New York (CC)*. These localities are located in the Seneca-Clyde-Oneida drainage system which flows by way of the Oswego River into Lake Ontario. For populations at AL (eutrophic environment) and BC (mesotrophic environment) variations in life-cycle, accompanied by marked differences in fecundity and biomass turnover rates (growth and egg reproduction), are reported. A quantification of fecundity is reported for groups of limpets maintained in cages in the field. Estimates of biomass are based on total organic carbon (equivalent to calorific measures), and the problems involved in making such estimates are discussed. Also, individual and seasonal variation in carbon, in nitrogen content, and in oxygen consumption are reported. These are related to the little known phenomenon of reverse respiratory acclimation. The population differences have significance in relation to peculiarities of evolution in the freshwater environments.
https://surface.syr.edu/bio_etd/67
oai:surface.syr.edu:bio_etd-1066
2010-11-17T14:43:49Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Studies in the Genus, Typha: I. Metaxenia, Xenia, and Heterosis. II. Interspecific Hybridization and the Origin of Typha Glauca. III. Autecology, with Special Reference to the Role of Aerenchyma
Marsh, Leland C.
1962-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Norman Gillette
Hybridization
Seed characteristics
Fecundity
Seed yield
Botany
The present study is intended to cast some light on the problem of T. glauca, but in so doing a slightly different experimental approach has been developed and applied. In the first part, intra-and interspecific hybridization of the three species, T. latifolia, T. angustifolia, and T. glauca, produced significant alterations in certain seed characteristics. There were color, dimensions of the external cellular layer, endosperm width, embryo width, and embryo length. ... In the second part, the relative fecundity of the species as measured by seed yield, together with external seed dimensions, seed weight and the above seed characteristics, were related to the issue of the origin of T. glauca. ...
https://surface.syr.edu/bio_etd/66
oai:surface.syr.edu:bio_etd-1067
2010-12-02T21:23:05Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Arbuscular Mycorrhizal Fungi in Grasslands of Yellowstone National Park: A Role for Plant-Fungal Mutualism in Grassland Sustainability
Murray, Tanya R.
2010-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Douglas Frank
Mycorrhizal fungi
Grasslands
Yellowstone National Park
Plant-fungal mutualism
Arbuscular mycorrhizae
Herbivory
Biology
Arbuscular mycorrhizal fungi are ubiquitous soil organisms that form symbioses with many families of terrestrial plants. Although mycorrhizae have been extensively researched, the relationships between arbuscular mycorrhizal fungal (AMF) community composition, AMF morphological structural investment, and the rate of nutrient exchange between symbionts is still unclear, especially in a field setting that includes multiple trophic levels and spatial scales. The research presented in this manuscript examined the influence of herbivory on plant-fungal dynamics across microbial, plant, and landscape level scales through mathematical modeling and field experiments. The first series of field studies demonstrated that some mycorrhizal species were more successful than others when host plants were grazed and/or when soil resource limitation varied. Grazing and soil nutrient availability induced shifts in the relative abundance of AMF spore species across a naturally occurring resource gradient, that resulted in maintained or increased nutrient delivery to the plant, and differentially influenced sporulation and hyphal production of AMF species. In the second field study soil nitrogen (N) fertilization by simulated bovine urine (SBU) amendment induced shifts in AMF species composition that resulted in higher benefit for the fungus than the host plant, demonstrated by increased sporulation of AMF, especially large-spored AMF species, and reduced abundance of plant supply structures. These results contribute to the understanding of how landscape variability and consumers influence the degree of benefit that plants and AMF species receive from their symbiotic relationship and contribute to community composition.
https://surface.syr.edu/bio_etd/70
oai:surface.syr.edu:bio_etd-1068
2010-12-03T13:28:04Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Fire and the reasons for its influence on mammalian herbivore distributions in an African savanna ecosystem
Eby, Stephanie L.
2010-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Mark E. Ritchie
Fire
African savanna
Herbivores
Predator-prey interactions
Biology
Fire is an abiotic factor which has long played a role in savanna and grassland ecosystems. Fire causes reductions in plant vegetation height and biomass and increases in plant nutrient content. Mammalian herbivores are attracted to post-fire burned areas and this attraction has largely been attributed to increases in plant nutrient content. However, because of the reduction in vegetation height and subsequent increase in sighting distance that fire causes, burned areas might also be safer habitats from predators. This dissertation investigates how fire influences the distribution of herbivores and carnivores in the post fire landscape. The results show that generally smaller sized herbivores prefer burned areas while larger sized herbivores do not. Both vegetation nutrient quality and vegetation height play a role in explaining this preference for burned areas. The role of predator avoidance as one possible explanation for herbivore preference of burned areas is further supported by data showing that lions ( Panthera leo ) do not prefer burned areas and do not kill more in burned areas despite increases in prey availability in these areas. To further investigate the potential impact of predators on herbivore use of burned areas, field studies were conducted on the vigilance behavior of Thomson's gazelles ( Gazella thomsonii ) in burned and unburned areas before and after exposure to a model cheetah ( Acinonyx jubatus ). Prior to cheetah presentation there was no difference in vigilance between the two habitats. However, right after cheetah presentation and removal vigilance levels were lower in burned areas than in unburned areas, indicating that Thomson's gazelles perceive burned areas to be safer habitats. Lastly, this dissertation explores three alternative hypotheses for herbivore preference of burned areas; (1) to avoid disease carrying and behavior changing invertebrates, (2) because burned areas are warmer microclimates, or (3) to obtain minerals from the ash. Of these three hypotheses only the ingestion of ash to obtain minerals is supported as a reason for herbivore preference of burned areas, but this would only be for time periods shortly after burning. The results from this dissertation contribute to our understanding of how and why fire influences herbivore distributions.
https://surface.syr.edu/bio_etd/69
oai:surface.syr.edu:bio_etd-1069
2010-12-03T13:51:46Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Metabolic and genetic engineering of Escherichia coli for the production of nonulosonic acid sugars
Lundgren, Benjamin Robert
2010-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Robert P. Doyle
Amino sugars
Metabolic
Genetic engineering
Nonulosonic acid
Sialic acid
Biochemistry, Biophysics, and Structural Biology
Microbial fermentation has proven to be a valuable and essential method for producing both high-value low-volume products such as pharmaceuticals and low-value high-volume goods, e.g., fuels and nutraceuticals. Compared to other industrial processes, the scalability, low-cost and green chemistry associated with Escherichia coli -based fermentation has driven it into the forefront of bioprocess engineering. Indeed, this bacterium is a vital recombinant microorganism for accessing not only common bioproducts such as amino acids and sugars, but is a source of more structurally complex, pharmaceutical-precursors, including 6-deoxyerythronolide B and artemisinic acid. Through metabolic and genetic engineering approaches presented herein, we were able to construct E. coli strains that could biosynthesize and produce the non-native, nonulosonic acid sugars, sialic acid and pseudaminic acid. These nine-carbon keto acids perform unique, key cellular functions, making them highly sought after in the medical and biotechnological industries. In addition, with the limited access to the nonulosonates via enzymatic synthesis, advancements in the basic research on the biochemistry and physiology of these sugars remains hindered. To increase the availability of these compounds, fully functional, biosynthetic pathways for sialic and pseudaminic acid were constructed and successfully expressed in E. coli. However, optimal production of either sugar required inimitable modifications to E. coli 's amino sugar metabolism for maximizing carbon flow into the desired nonulosonate and obtaining industrial-relevant titers. Known as an industrial workhorse, the E. coli processes developed herein will serve as valuable platforms for accessing nonulosonic sugars and their derivatives.
https://surface.syr.edu/bio_etd/68
oai:surface.syr.edu:bio_etd-1070
2010-12-03T14:13:56Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Sophisticated regulation of histone H3 lysine 9 dimethylation accumulation during meiosis in Caenorhabditis elegans
She, Xingyu
2010-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Eleanor M. Maine
Unpaired chromatin
Meiotic silencing
Caenorhabditis elegans
Histone H3 lysine 9 dimethylation
Meiosis
Biology
Meiotic silencing of unpaired chromatin (MSUC) is a process that has been proposed to play an essential role in defending against genetic parasites and maintaining gamete quality during sexual reproduction. Our lab uses the model organism, Caenorhabditis elegans, to study this phenomenon. In C. elegans, MSUC is thought to limit transcription of unpaired chromatin by labeling it with a silencing epigenetic marker, histone H3 lysine 9 dimethylation (H3K9me2), and inducing heterochromatin formation. A germline specific RNA-directed RNA polymerase (RdRP), EGO-1, has been shown to be required for the accumulation of H3K9me2. This study, along with the research of several other labs, has also revealed additional regulators that affect the H3K9me2 pattern during meiosis. Two of them function in small RNA pathways: RNA Helicase A (RHA-1), and another RdRP (RRF-3). These results strongly suggest a connection between the small RNAs and MSUC.
In this study, I discovered five genes that are involved in regulating the H3K9me2 accumulation pattern during MSUC. Specifically, removal of CSR-1, DRH-3 and EKL-1 function causes an abnormally high level of H3K9me2 on paired chromatin; removal of SIN-3 function leads to absence of H3K9me2 on unpaired chromatin regions that may be highly histone-acetylated; removal of ERI-1 function delays removal of H3K9me2 from developing sperm cells, suggesting that ERI-1, like RRF-3, is a negative regulator of MSUC. The majority of my dissertation focuses on studying the functions of CSR-1, DRH-3 and EKL-1. My results suggest that CSR-1, DRH-3, EKL-1, and EGO-1 may represent a functional pathway in MSUC. I also provide additional information about the functions of SIN-3, HIM-17, RHA-1 and ERI-1. Based on these results, alternative models of the H3K9me2 modification pathway in C. elegans are described.
https://surface.syr.edu/bio_etd/72
oai:surface.syr.edu:bio_etd-1072
2011-09-15T19:38:05Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
publication:oa_etd
The Maintenance of Male Color Polymorphism in Poecilia parae
Hurtado-Gonzales, Jorge L.
2011-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
J. Albert C. Uy
Competition
Female mating preferences
Genetic color polymorphism
Poeciliidae
Sexual selection
Visual environment
Biology
<p>Genetic color polymorphisms are common in nature, and a major challenge for evolutionary biologists has been to understand how they are maintained despite the effects of directional selection. Recent studies suggest that frequency-dependent selection may explain the persistence of intraspecific variation in color. However, it remains to be determined whether frequency-dependent selection is the most predominant (or perhaps the only) mechanism to maintain such genetically-based polymorphisms. Using a South American fish, Poecilia parae, my dissertation aims to elucidate the relative roles of natural and sexual selection in the maintenance of polymorphisms in natural populations. Poecilia parae males exhibit five distinct, Y-linked and co-occurring color morphs: (i) `immaculata', the smallest and drab-colored males that resemble juvenile females; (ii) `parae', the largest males that exhibit a striped tail and black vertical body bars that intensify during social interactions; and (iii) the blue, red, and yellow morphs that are of intermediate body size and display colorful body flanks. Field surveys indicate that the frequency of each morph remains relatively stable and consistent over multiple years. Using a combination of observational and experimental studies, accompanied by techniques that aimed to characterize the visual ecology (e.g., water light transmission, visual sensitivity) of Poecilia parae, I found that this striking color polymorphism is maintained by a complex balance between different components of natural and sexual selection. First, males of the five color morphs employ different behavioral mating strategies (hereafter alternative mating strategies, AMSs) to maximize their reproductive success. These AMSs are also accompanied by differences in morphological traits, such as testes investment and sperm morphometry that complement the specific tactic. For instance, immaculata males are often categorized as an `unattractive' male by females, which is correlated to its phenotypical appearance (smaller and drab). However, this morph is relatively abundant. I found that immaculata males specialize in `sneak' copulations and have adaptations (i.e., larger testes and unique sperm morphology) that possibly confer a fertilization advantage during postcopulatory events, when competing with the sperm of more attractive males. My studies also indicate that females have strong mating preferences for red and yellow males, with visual predators also favoring those males as prey. These findings suggest that the antagonistic interaction between pre-mating sexual selection favoring and predation acting against the red and yellow morphs may prevent them from eliminating other color morphs from the population. In fact, the red and yellow males were consistently found to be the rarest morphs across populations. Further, despite overall preference for red or yellow males, my analysis also detected female preferences for blue males. These results suggest that the interaction between female mating preferences and predation accompanied by variation in male reproductive strategies may allow for the maintenance of complex color polymorphism in natural populations. Males of the different morphs further vary in their levels of aggressive behaviors. In a series of controlled lab experiments, I found that parae males gain successful matings by preventing other males from accessing females and/or modifying female mating preferences after test females witness successful agonistic interactions. Finally, I found that the signaling environment of Poecilia parae in nature is variable and that this contributes to temporal and spatial variation in how males are perceived by females. Although red males were typically the most conspicuous morph, blue males were sometimes more conspicuous than red males at several sampling sites. The results also revealed that an aquatic model predator is able to discriminate the same male color morphs that resulted also conspicuous for females. However, it is possible that males and females of Poecilia parae are using parts of the background spectral that are least sensitive for predators. In conclusion, my dissertation suggests that balancing selection defined by the interaction between various episodes of selection allows for the invasion of AMSs and thus the maintenance of the male color polymorphism in Poecilia parae.</p>
https://surface.syr.edu/bio_etd/73
oai:surface.syr.edu:bio_etd-1074
2011-09-15T19:37:34Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
publication:oa_etd
The Consequence of Anthropogenic Disturbance on Communication and the Operation of Sexual Selection in the Eastern Bluebird (Sialia Sialis)
Wisner, Ellen Michale
2011-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
J. Albert C. Uy
anthropogenic disturbance
communication
Eastern bluebird
urban noise
Biology
<p>Signals that are efficiently transmitted and easily detected in their signaling environment are favored by natural selection. Anthropogenic disturbance can rapidly alter the signaling environment, and recent studies have shown that acoustic and visual signals change in response to these altered habitats. Although these studies provide important insight into the effects of urbanization on animal signals and have served as experiments testing the role of the environment in shaping signal design, several key aspects of how signal design can be influenced by environmental changes remain unclear.</p>
<p>Previous studies have focused on signals used between adults, such as those used in mate choice, yet other signals should be similarly affected by anthropogenic disturbance. Thus, one facet of my research examines how anthropogenic disturbance can influence parent-offspring communication. I tested whether nestling mouth coloration in Eastern bluebirds (Sialia sialis) was a signal of quality, and if a parents ability to discriminate among the mouth coloration of their nestlings was affected by level of human disturbance. I found that the perceived color contrast of nestling mouths against its nest was significantly correlated with nestling body condition, suggesting that it may be signal of nestling quality. Additionally, I found that the parent's ability to perceived a difference in color contrast of a nestling's mouth among nest-mates was lower in disturbed habitats, than in undisturbed habitats, showing less discriminability among nestlings in disturbed habitats. These results suggest that parent-offspring communication can be affected by anthropogenic disturbance which may reduce a parent's ability to preferentially invest in high quality young.</p>
<p>Past research on anthropogenic disturbance and signaling has focused on the response of single signals, yet most organisms communicate using signals from multiple sensory systems (i.e., multimodal signals). Thus, I examined how anthropogenic disturbance can simultaneously influence components of multimodal signals in Eastern bluebirds. I measured the visual and acoustic environment at different disturbance levels and related them to male plumage and song characteristics. I found that in areas with high levels of anthropogenic noise, males sing at a higher minimum frequency, presumably to avoid overlap with low frequency background noise. I also found that the visual background is altered in disturbed sites; however, plumage characteristics did not covary with the altered habitats. These results suggest that human disturbance is interfering with both visual and acoustic signals, yet only acoustic signals have responded to the changes.</p>
<p>Few studies on anthropogenic disturbance directly explore the explicit evolutionary mechanisms underlying the changes in signal design. Thus, in the final chapter of my dissertation, I explored how selection on traits varied across habitats with different levels of disturbance. To do this, I determined paternity of nestlings using microsatellite. Then, I determined the major factors influencing rates of extra-pair paternity (i.e., proportion of nestlings within a nest that were sired by other males), and tested whether these factors varied with disturbance levels. I found that the minimum frequency of song, and the brightness of the male's chestnut breast are important predictors of extra-pair paternity rate across all disturbance levels. Additionally, I found an interaction between disturbance level and the minimum frequency of song in relation to extra-pair paternity. This interaction effect was due to differences in selection pressure on the minimum frequency of song in relation to habitat disturbance. Males that sing in higher minimum frequencies have lower rates of extra-pair paternity, in disturbed areas, but higher rates of extra-pair paternity in undisturbed areas. These results suggest that selection on signals vary across disturbance levels and this could drive the observed changes in the design of signals. Potential consequences of these changes include the possibility of long-term differentiation between bluebird populations living in disturbed and undisturbed habitats, and a shift in important traits across the entire species.</p>
https://surface.syr.edu/bio_etd/75
oai:surface.syr.edu:bio_etd-1076
2012-05-24T12:55:54Z
publication:dbio
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Enhancer Binding Proteins Initiate and Propagate Multicellular Development in Myxococcus xanthus
Giglio, Krista Michelle
2011-12-01T08:00:00Z
Dissertation/Thesis
Doctor of Philosophy (PhD)
Biology
Anthony Garza
Biofilms
Enhancer binding proteins
Sigma 54 promoters
Two-component systems
Biology
<p><em>Myxococcus xanthus</em> responds to starvation by engaging a programmed series of morphological changes that yield multicellular fruiting bodies filled with dormant spores. Previous work suggests that <em>M. xanthus</em> uses a series of transcriptional activators called enhancer binding proteins (EBPs) to coordinate expression of many developmental genes. This work focuses on six EBPs (Nla4, Nla18, Nla6, Nla28, ActB and MXAN4899) that regulate early development. It is during this critical stage when information from both external and internal environments must be integrated and cells must decide to continue to grow and divide, or to arrest growth and commit to development. We find that the early developmental EBPs function sequentially during development. DNA microarray analysis and quantitative PCR revealed a transcriptional hierarchy among the early-acting EBP genes. EBPs work in conjunction with σ<sup>54</sup> promoters, and bioinformatics revealed that five of the EBP gene operons have putative σ<sup>54</sup> promoters, which suggested that the EBPs might form a cascade involving direct transcriptional regulation. Subsequent electrophoretic mobility shift assays (EMSAs) performed with the purified DNA binding domains of the six EBPs and fragments of the EBP gene promoters, indicated that an EBP functioning at one developmental stage directly binds to the promoter region of the EBP gene that functions at the next developmental stage. In addition to activation of downstream EBP genes within the cascade, it is probable that each EBP in the cascade activates many genes that are important for development. Using a combination of DNA microarray analysis and bioinformatics, we identified hundreds of operons that are activated in the early stages of development that are potential targets of the EBPs in the cascade. To further investigate the role of EBP-mediated transcription during development, we focused on Nla6, and identified developmental target operons for this EBP. Alignment of the promoter regions positive for Nla6 binding revealed a 10 bp tandem repeat as a putative Nla6 binding site. Using both <em>in vitro</em> and <em>in vivo</em> methods, we find that the 10bp consensus is important for Nla6 binding to the target promoters, and for expression of target operons. Using the confirmed consensus and the sequence of the <em>M. xanthus </em>genome, we identified 42 putative developmental promoter targets for Nla6. EMSAs were used to confirm that Nla6 binds to six of the putative target promoters. Additionally, we find that mutations in four of the targets tested caused developmental phenotypes similar to that of an nla6 mutant strain. The abundance of putative EBP targets and their functional importance to development, suggest that the cascade is part of a complex regulatory network that is essential to the initiation and propagation of multicellular development.</p>
https://surface.syr.edu/bio_etd/77
oai:surface.syr.edu:bio_etd-1077
2012-04-02T13:23:32Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Molecular Characterization of Genetic Components of Pathogen Defense in Arabidopsis
Dutta, Aditya
2011-01-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Ramesh Raina
Arabidopsis
Histone demethylation
Plant defense
Pseudomonas syringae
Reactive oxygen species
Salicylic acid
Molecular Biology
<p>Plant defense against pathogens involve complex interplay of several defense-associated factors. This work characterizes three Arabidopsis thaliana genes that play critical roles in regulating defense responses against the bacterial pathogen, Pseudomonas syringae. We have previously described phenotypic characterization of the hrl1 mutant of Arabidopsis; a lesion-mimic mutant that is small in size compared to the wild type, spontaneously develops necrotic lesions mimicking the hypersensitive response, constitutively expresses several defense-related genes and has enhanced resistance against a variety of pathogens. In this work I cloned the HRL1 gene by a map-based cloning approach. HRL1 codes for a 4-hydroxybenzoate polyprenyl diphosphate transferase, which catalyzes a rate limiting step in ubiquinone biosynthesis. My studies suggest that mitochondrial metabolic reactive oxygen species are key regulators of Arabidopsis basal resistance against P. syringae. I further identified and characterized a suppressor of the hrl1 mutant; JMJ27 (ARABIDOPSIS JUMONJI 27) gene of Arabidopsis. JMJ27 codes for a JmjC domain-containing H3K9 histone demethylase and is required for defense against P. syringae. Additionally, another novel component of plant defense, SMALL DEFENSE-ASSOCIATED PROTEIN 1 (SDA1) gene of Arabidopsis, was characterized. SDA1 was identified as a gene that is constitutively expressed to high levels in the hrl1 mutant. SDA1 defines a novel class of small pathogen defense proteins required for defense against P. syringae.</p>
https://surface.syr.edu/bio_etd/78
oai:surface.syr.edu:bio_etd-1078
2012-08-17T19:12:16Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
publication:oa_etd
Role of the Nla6S and Nla28S histidine kinases in fruiting body development of <i>Myxococcus xanthus</i>
Sarwar, Zaara
2012-06-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Anthony Garza
Biofilm formation
histidine kinase
Myxococcus xanthus fruiting body formation
two component signal transduction systems
Biochemistry, Biophysics, and Structural Biology
<p>The complex life cycle of Myxococcus xanthus makes it a model organism for studying multicellular developmental processes in bacteria. In response to adverse environmental conditions, M. xanthus aggregates and forms multicellular structures known as fruiting bodies. Two component signal transduction systems (TCS) are widely used by bacteria to detect and respond to environmental cues by regulating large-scale changes in gene expression. They contain a histidine kinase sensor that detects environmental cues and a response regulator that modulates cellular processes. Two key regulators of the early stages of fruiting body development are the Nla6S/Nla6 and Nla28S/Nla28 TCSs. The response regulators of these TCSs, Nla6 and Nla28, are important for the successful completion of fruiting body formation. However, the histidine kinase sensors that modulate the activity of these key response regulators were previously unknown. Here we report the identification and characterization of the Nla6S and Nla28S histidine kinases. Analysis of Nla6S reveals that it represents a new family of bacterial histidine kinases. Furthermore, it plays an important role in the M. xanthus life cycle. Analysis of Nla28S shows that this is an important sensor of early developmental events. Our data suggests that Nla28S is involved in sensing the two important events of early development, nutrient depletion and cell density. In response to these signals Nla28S regulates sporulation of M. xanthus. Characterization of Nla6S and Nla28S expands our knowledge of the signaling networks that regulate initiation of the multicellular developmental process of M. xanthus.</p>
https://surface.syr.edu/bio_etd/79
oai:surface.syr.edu:bio_etd-1079
2012-08-31T13:40:29Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
publication:oa_etd
Roles for Chd7 in Zebrafish Development with Implications for Human Disease
McDaniels, Nicole Lynn
2012-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Meilssa Pepling
R. Craig Albertson
CHARGE Syndrome
chd7
development
RNA-Seq
spinal deformity
zebrafish
Biology
<p>The Chromodomain Helicase DNA Binding Protein (CHD) family consists of a group of nine known proteins that function in controlling DNA dynamics and transcription. The CHD family member of interest in this work, <em>Chromodomain helicase DNA-binding protein 7 (chd7)</em>, has been implicated in human CHARGE (coloboma of the eye, heart defects, atresia of the choanae, retardation of growth and/or development, genital and/or urinary abnormalities, and ear abnormalities and deafness) Syndrome and Idiopathic Scoliosis, however little is known about the roles this gene plays during development.</p>
<p>Using zebrafish as a model system, morpholino antisense technology, whole mount <em>in situ</em> hybridization (WISH), and a relatively new protocol, RNA-Seq, we provide evidence of several developmental defects resulting from Chd7 knockdown as well as describe several genes that exhibit significant differential expression upon Chd7 knockdown. We provide evidence that zebrafish embryos highly express <em>chd7</em> in the retina, brain and somite boundaries. We demonstrate that a reduction in Chd7 synthesis results in laterality defects in the expression of somitogenesis genes, consistent with the hypothesis that this chromatin remodeler is necessary for the proper development of the long axis of the body and may be involved in the onset of human scoliosis. We show that the presence of Chd7 is crucial for proper neural, retinal and vertebral development in zebrafish and that loss of function of Chd7 resulted in several morphological defects similar to those observed in human patients with CHARGE syndrome.</p>
<p>Finally, this work is the first transcriptome-wide study analyzing genetic changes in response to knockdown of a member of the Chromodomain Helicase DNA Binding Protein family. Using RNA-Seq we were able to identify and quantify differentially expressed genes in <em>chd7</em> morphant zebrafish embryos compared to control morphant zebrafish embryos. These data were consistent with our knockdown experiments as well as with putative roles for Chd7 in human diseases.</p>
<p>In an effort to make a broader impact, we extended the use of zebrafish beyond scientific research and described several different methods of using zebrafish in the undergraduate laboratory classroom. First, we described the use of WISH in the classroom to help students form connections between molecular and organismal biology and then described the use of two model organisms, zebrafish and <em>C. elegans</em>, in the classroom to help students understand how genes and the environment interact to affect organismal development.</p>
<p>Cumulatively, this work is an example of interdisciplinary study both in the research laboratory as well as in the classroom. It involves the intensive study of a chromatin remodeler, Chd7, and makes important connections between Chd7 knockdown in zebrafish and human diseases. It also describes the pedagogical advantages of model organisms with respect to providing a rich and integrative experience for undergraduate biology majors.</p>
https://surface.syr.edu/bio_etd/80
oai:surface.syr.edu:bio_etd-1080
2012-09-18T15:35:52Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
publication:oa_etd
High Throughput Screening of Aptamers
Kupakuwana, Gillian Vimbai
2011-05-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Philip N. Borer
Aptamer
High throughput screening of aptamers
SELEX alternative
Thrombin binding aptamer
Biochemistry, Biophysics, and Structural Biology
<p>Aptamers are functional oligonucleotides that are discovered through directed evolution. With affinities for their targets similar to antibodies, aptamers are useful in biosensors, diagnostics and therapeutics. Three elements of natural selection--variation, selection, and replication afford aptamer discovery methods the ability to enrich and distinguish aptamers from randomized pools. Here I describe an aptamer discovery method we are calling High Throughput Screening of Aptamers (HTSA), which incorporates two additional elements that eliminate the cyclic evolution bottleneck in traditional aptamer discovery methods, and expedite the discovery process with no complex automation required. We designed a method that (1) affords multiple representation of every possible sequence (oversampling) or alternatively the multiple representation of each present sequence (reduced complexity) in a library pool and (2) employs massively parallel sequencing after the selection process. The first element facilitates the rapid expulsion of low affinity binding sequences while retaining the high affinity binders at frequencies far above the background probable frequencies. The second element then enhances the observation of the enrichment by providing a comprehensive picture of the enriched pool that cannot be achieved by conventional sequencing of a few clones. We essentially used high throughput sequencing as a sequence reader of enriched library members and a directional sensor of the selection process. We could predict the relative affinity of a sequence for its desired target by simply counting its frequency in the enriched pool.</p>
<p>The goal of the work reported in this volume was to develop a high throughput screening method for aptamers that is viable across a variety of potential targets in single-target and multiplexedtarget environments using whole organisms/cells (live <em>Cryptosporidium Parvum</em> oocysts), protein targets (<em>a</em> thrombin, as well as <em>Cryptosporidium Parvum</em>'s surface proteins, Cp23 and Cp15) and small molecule targets (<em>a</em> thrombin's glycan moiety and saccharides). We demonstrated the efficacy of our method to rapidly screen and distinguish minimal aptamers by co-isolating, in a single selection step, a previously known DNA aptamer with nanomolar affinity for human á thrombin that was originally discovered after 5 selection cycles and an unprecedented aptamer with apparent specificity for hexose sugars with affinity in the top third of known aptamers for small molecules, from an oversampled library. We also demonstrated HTSA's superior ability to enrich undersampled RNA libraries without any cycling, by successfully isolating high affinity binding sequences against <em>Cryptosporidium Parvum</em> oocysts and two of its surface wall proteins Cp23 and Cp15. Undersampled libraries theoretically have a single copy of each present sequence and traditionally require cyclic enrichment for the copy number to increase enough to result in an observable enrichment; which is the goal of every aptamer discovery method. We circumvented the cycling by translating the undersampled library into a reduced complexity library, which is essentially an undersampled library depleted in lowaffinity sequences by partitioning against the target, then amplified to create multiple copies of each present sequence; this basically functions like an oversampled library that effects an observable enrichment after a single selection step.</p>
<p>Additionally, as a result of the dramatic increase in throughput from Sanger to second generation sequencing, we also demonstrate HTSA's ability to exhaustively search the space of sequences within an enriched library which simplifies the characterization of the core binding "domain." Ultimately, HTSA simplifies and shortens the discovery process from weeks to months of standard SELEX to less than a week, exhaustively searches the space of sequences within a library, may eliminate the need to truncate long aptamer sequences to find the core binding domain, reduces the quantity of the target required and cycling artifacts since it eliminates cycling altogether, and allows multiplexing of targets and experiments.</p>
https://surface.syr.edu/bio_etd/81
oai:surface.syr.edu:bio_etd-1081
2014-08-15T17:58:30Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
Identification and Characterization of Pathogen Defense Genes of Arabidopsis thaliana
Caruana, Julie
2012-08-01T07:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Ramesh, Raina
Arabidopsis thaliana
chromatin remodeling
microRNA
pathogen defense
Pseudomonas syringae
Biology
<p>Plants are constantly under attack from microbes and therefore employ complex and multilayered defense mechanisms to prevent infection. This work reports characterization of defense regulatory genes of <em>Arabidopsis thaliana</em>, including a microRNA, a family of histone demethylases, and a gene of unknown function. The microRNA <em>miR167</em> was previously found to regulate flower development by controlling patterns of expression of the <em>AUXIN RESPONSE FACTOR</em> genes <em>ARF6</em> and <em>ARF8</em>. I found that <em>miR167</em> is differentially expressed in response to treatment with the bacterial pathogen <em>Pseudomonas syringae</em>, and that overexpression of <em>miR167</em> confers very high levels of resistance to the pathogen. The high resistance of <em>miR167</em> overexpression plants appears to be due to shifts in the balance of hormones to favor those that strengthen defense, and to their tendency to maintain stomata in a closed state, thus preventing entry of bacterial cells into the leaf interior where they can cause disease. This work highlights a new role for a microRNA previously known only for its role in development. I also began characterization of several members of the JumonjiC domain-containing family of histone demethylases. We have previously shown that one member of this family, JMJ27, is required for defense against <em>P. syringae</em>. I identified several other family members that are differentially expressed upon pathogen treatment and two that are critical for defense. Finally, I characterized the <em>TolB-RELATED DEFENSE PROTEIN 1</em> gene of <em>Arabidopsis</em>. This gene is strongly induced by a variety of pathogens and abiotic stresses and encodes a protein of unknown function. <em>tdp1</em> knockout mutants did not have altered resistance to <em>P. syringae</em> or oxidative stress, but the high induction of this gene under so many conditions suggests that it plays an important, if not critical, role in stress management.</p>
https://surface.syr.edu/bio_etd/82
oai:surface.syr.edu:bio_etd-1082
2013-02-21T18:56:56Z
publication:dbio
publication:etd
publication:coscde
publication:bio_etd
publication:cas
publication:bio
publication:oa_etd
Molecular Mechanisms Regulating Neonatal Oocyte Survival and Primordial Follicle Formation in the Mouse Ovary
Jones, Robin L.
2012-12-01T08:00:00Z
Dissertation
Doctor of Philosophy (PhD)
Biology
Melissa Pepling
Follicle
Mouse
Oocyte
Ovary
Reproductive Biology
Cell Biology
Developmental Biology
Molecular Biology
<p>In mammals, formation of the primordial follicle is a complex process involving the breakdown of germ cell cysts, where oocytes must separate from each other and subsequently become surrounded by somatic cells. As cysts separate, a large number of germ cells are lost by apoptosis, however the mechanisms by which cyst breakdown and germ cell death occur are not well understood. We first hypothesized that two anti-apoptotic regulators from the BCL2 family of proteins, BCL2 and MCL1, may be responsible for regulating neonatal oocyte survival. To elucidate the effects of BCL2 in the neonatal ovary, we examined ovaries of both <em>Bcl2</em> overexpressing and knockout transgenic mice. When compared to wild-type mice, neither <em>Bcl2</em> overexpression nor abrogation significantly altered ovarian histology. Another BCL2 family protein, MCL1, is expressed in human oocytes during ovarian development, suggesting a role for MCL1 in oocyte survival. We found that MCL1 was localized to both oocytes and somatic cells during cyst breakdown. Subsequently, we used an <em>in vitro</em> organ culture system to identify a role for MCL1 in oocyte survival. We found that inhibition of MCL1 with an antibody to MCL1 in culture resulted altered germ cell numbers and oocyte cyst breakdown. Our data demonstrate that while BCL2 is not likely involved in perinatal oocyte survival, MCL1 may be an important regulator of the ovarian primordial follicle reserve. Next, we hypothesized that the KIT signaling pathway may be important for oocyte survival and cyst breakdown in the neonatal ovary. The tyrosine kinase receptor, KIT, and its ligand, KITL, have been implicated in oocyte survival and follicle development in both fetal and adult ovaries but have not been well studied at the perinatal time point. To elucidate the functional role of KIT signaling in the neonatal ovary, we began by using immunohistochemistry to test the expression of KIT and KITL. We found both proteins to be expressed in the developing ovary from 17.5 dpc to PND 3, suggesting an important role for this protein in cyst breakdown or oocyte survival. To test this hypothesis, ovaries from 17.5 dpc fetal mice were cultured for 5 days in control media, or in media with the KIT blocking antibody, ACK2, or recombinant KITL. Our data demonstrated a role for KIT signaling in cyst breakdown, as inhibition and activation of the pathway altered ovarian histology. Using cell proliferation and TUNEL assays at the conclusion of culture, we identified a reduction in somatic cell proliferation when KIT signaling was inhibited and likewise, a decrease in cell death. Finally, we investigated which pathway downstream of KIT affects cyst breakdown and the effect of KIT signaling on MCL1 protein expression. After 3 days in culture with KITL supplemented media, Western blotting was used to analyze the total and phosphorylated forms of proteins from the PI3K, MAPK and JAK-STAT pathways as well as the BCL2 family protein, MCL1. We found that there was an increase in the phosphorylated forms of p44/p42 in the MAPK pathway and a downregulation of MCL1 on KIT activation. Overall our data have shown that while BCL2 may not contribute to oocyte survival during cyst breakdown, both MCL1 and KIT play important roles in formation of the primordial follicle pool.</p>
https://surface.syr.edu/bio_etd/83
oai:surface.syr.edu:bio_thesis-1000
2013-05-21T13:19:20Z
publication:dbio
publication:coscde
publication:thesis
publication:bio_thesis
publication:cas
publication:bio
Structural and Functional Studies on the Role of the Retinoblastoma Binding Protein Five (RbBP5) in the Histone Methyltransferase Activity of the Mixed Lineage Leukemia (MLL1) Core Complex
Sanders, Melody
2013-05-01T07:00:00Z
Thesis
Master of Science (MS)
Biology
Ramesh Raina
Michael S. Cosgrove
Biochemistry, Biophysics, and Structural Biology
Biology
<p>Cells employ elaborate mechanisms to introduce structural and chemical variation into chromatin in the form of covalent post-translational modifications. Covalent modifications of histones contribute to the dynamic states of chromatin structure that govern nearly all of DNA-coupled processes such as transcription, replication, and repair. The mechanism by which covalent modifications of histones contribute to these activities remains a central question to understanding genome regulation and its dysfunction in human disease. Although there is extensive literature documenting the identification of many of the enzymes that place histone modifications, far less is known about how the enzymes are targeted and how their enzymatic activities are regulated. The long-term goal of the work in this thesis seeks to understand the mechanisms underlying the accessibility of genes in chromatin. In particular, this study focuses on identifying the underlying molecular mechanisms involved in the regulation of one such element of variation in chromatin, the methylation of lysine four on histone H3.</p>
<p>Histone H3 lysine 4 methylation (H3K4me) is an evolutionarily conserved epigenetic mark that is correlated with transcriptional activation in eukaryotes. H3K4 methylation has been shown to be important for a number of biological processes including gene expression and DNA replication. This mark is mainly catalyzed by a group of enzymes that contain an evolutionarily conserved SET domain. The evolutionarily conserved SET domain was originally named for its presence in three <em>Drosophila melanogaster</em> proteins: the position effect variegation modifier SU(VAR)3-9, the polycomb group protein E(z), and the trithorax group protein (TRX) . In vertebrates, the Mixed Lineage Leukemia protein-1 (MLL1) belongs to the SET1 family of histone H3K4 methyltransferases. The catalytic activity of MLL1 is regulated by a conserved group of proteins that include the Tryptophan-Aspartate-repeat protein-5 (WDR5), the retinoblastoma-binding protein-5 (RbBP5) and the absent small homeotic-2-like protein (Ash2L).</p>
<p>The focus of this investigation is the RbBP5 component of the MLL1 core complex. The RbBP5 subunit of the MLL1 core complex has been shown to be required for enzymatic activity and disruption of RbBP5 is frequently observed in patients with malignant primary brain tumors. To gain insight into the functional role of RbBP5 in the regulation of the enzymatic activity of the MLL1 core complex, mutations targeting individual residues in a highly conserved stretch of amino acid residues in RbBP5 were generated. Biochemical and biophysical analyses of the variant proteins were utilized to assess the functional role of each amino acid on the intrinsic properties of RbBP5 alone or when assembled within the context of the entire complex. These studies identify several conserved aromatic residues and one acidic residue in RbBP5 required for interaction with MLL1 and the overall dimethyltransferase activity of the complex. The residues identified constitute a previously uncharacterized MLL1 interaction motif located in RbBP5. Therefore, this study provides important insights into how the RbBP5 subunit of the MLL1 core complex contributes to its overall enzymatic activity. Understanding the role that RbBP5 plays in facilitating proper H3K4 methylation may provide insight into how misregulation of RbBP5 can lead to brain tumor genesis.</p>
https://surface.syr.edu/bio_thesis/1
1460061/qualified-dublin-core/100//