Honors Capstone Project
Date of Submission
Arts and Science
Capstone Prize Winner
Won Capstone Funding
Sciences and Engineering
Biochemistry | Biochemistry, Biophysics, and Structural Biology
One of the most critical structures in cellular biology is the plasma membrane, due to its ability to respond to environmental stresses. Saccharomyces cerevisiae is a model, single-celled eukaryote that has been used to investigate many aspects of cell biology. A recent genetic screen in yeast for plasma membrane homeostatic proteins identified three related proteins of unknown molecular function that participate in these processes. These proteins, termed PDR19, PDR20, and PDR21 for Pleiotropic Drug Resistance, are each approximately one hundred amino acids in size and share a small conserved domain, namely the core sequence KITRYDL. In the case of PDR21, the core sequence is VITRHDL. The coding sequences for this set of proteins are found in the ORFs YGR035c, YLR346c and YPR145w-a, respectively. A triple mutation of these genes led to an observed decrease in membrane homeostasis, when cells were treated with the membrane- disrupting compound digitonin, natural products that disturb membranes and in the presence of the clinical antifungal drug amphotericin B. The observed phenotype suggests this set of novel proteins functionally regulate membranes in response to membrane-altering conditions, as an observed fifty-fold increase in membrane sensitivity of the triple mutant was observed. In order to help determine the molecular function(s) of the PDR proteins, a GAL4 two-hybrid system is being used to screen for proteins that may associate with the PDR 19/20/21 family proteins and help mediate their cellular functions. That this system can be used has been confirmed through negative autoactivation tests involving a Gal4DBD-PDR fusion construct and done in a modified Y187/Y190 mating strain carrying the pACT II activation domain plasmid containing the ADGal4. Plasmid sequencing of the Gal4DBD-PDR fusion proteins is in process to help confirm proper cloning of the bait proteins in addition to library screening. In addition to this work, bioinformatic characterization of genes involved in TTG cellular responses was conducted. In a previous screen in the Erdman lab, 4,851 deletion strains were screened, of which 991 strains demonstrated a degree of sensitivity or resistance to TTGs. In an attempt to further understand and classify these results, a bioinformatics tool
was used to reveal underlying modes of genetic control governing the range of observed phenotypic sensitivities.
White, Casey, "Identifying Proteins that Interact with the Novel Saccharomyces cerevisiae Proteins PDR19 and PDR20 and Bioinformatic Characterization of Genes Involved in TTG Cellular Responses" (2009). Syracuse University Honors Program Capstone Projects. 446.
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