Date of Award

5-15-2015

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Physics

Advisor(s)

Kenneth W. Foster

Keywords

Active Diffusion, Analytical Modeling of Motility, Chlamydomonas reinhardtii, Control of Ciliary Motility, I1/f-dynein, IC138

Subject Categories

Physical Sciences and Mathematics

Abstract

The green alga Chlamydomonas reinhardtii has evolutionarily developed a range of receptors to detect light, chemical, or mechanical stimuli for its survival. It employs its two cilia, identified as cis and trans with respect to its photoreceptor-containing eye, to achieve appropriate behavioral responses. The sensory signals are relayed to the dynein motors in the cilia through complex networks of signal transduction pathways that have yet to be fully characterized. The first part of this work is an experimental study of one such signal transduction pathways, the membrane electric field. In the study, an experimental method is developed to monitor the membrane electric field transients in response to an external stimulus. The method is non-invasive and allows monitoring the membrane electric field of cell population over extended periods of time by using a voltage-sensitive fluorescence probe, di-8-ANNEPS. The method is also insensitive to cell orientations and is suitable for studying the effect of any stimuli that may influence the behavior of cells by changing the membrane electric field. In this work two such types of stimuli, green light and sound, are used. In response to impulses of green light, the membrane electric field was found to change in the same way for both positively and negatively phototactic strains, and all the processing due to green light detection at the eye appeared to take place in the cilia. In response to sound stimuli, amplitude-modulated as 1-second-on-1-second-off or sine waves at 8.0 Hz, no change in the membrane electric field was observed.

The second part of this work is devoted to tracking experiments of swimming Chlamydomonas reinhardtii cells. The measured cell trajectories are quantified using a suitable implementation of the cell-motility model developed in the third part of this work. Through quantifying the cell trajectories using the motility model, the activity of IC138 component of cilium's inner I1/f-dynein arm is characterized. This unit has a regulatory role in motility. When it is phosphorylated (due to increased level of cAMP), the probability of it acting like a transient brake or an extra drag on the trans-cilium increases. This in turn causes a low-amplitude extra beat relative to the cis-cilium that maintains a steady beat. The extra low amplitude beat causes the cell to change direction more frequently, which makes the motion less ballistic and more diffusive. This ballistic-diffusive ratio affects the behavior associated with mating and searching for food and light in opposing manners. More frequent activation of the brake, for example, worsens search for food and light but increases chances of mating. In order to quantify this regulatory mechanism, which is a part of the braking signal transduction network, individual tracks of six Chlamydomonas reinhardtii strains were recorded and the data was fit to the above mentioned motility model at the population level. Among the obtained set of statistical parameters from fitting, the persistence time was found to be the most suitable one for characterizing the activity of IC138.

In addition to this, a special realization of the cell-motility model, suitable for studying the effect of an external periodic force on motility, is also developed in the third part of this work. This realization provides a quantitative mean to discern between the pure mechanical effect of an external periodic force, such as sound, and its sensory detection on the cell behavior.

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