Thesis (M.Sc.)--York University, 2008. Graduate Programme in Biology. / Typescript. Includes bibliographical references (leaves 83-89). Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:MR38831

Thesis (M.S.)--Rutgers University, 2009. / "Graduate Program in Cell and Developmental Biology." Includes bibliographical references (p. 58-65).

Thesis (M.S.)--Rutgers University, 2009. / "Graduate Program in Cell and Developmental Biology." Includes bibliographical references (p. 44-48).

Thesis (Ph. D.)--University of Oregon, 2003. / Typescript. Includes vita and abstract. Includes bibliographical references (leaves 159-180). Also available for download via the World Wide Web; free to University of Oregon users.

Insulin signaling is highly conserved across animals, and is known for its ubiquitous function in all aspects of animal physiology. Despite its relatively well-studied role in metabolism and energy expenditure, how it is involved in learning and memory remains a mystery, due to the complex nature of the nervous system. In this thesis, I have used C. elegans, a tractable model organism with a sophisticated behavioral repertoire, to investigate molecular and cellular mechanisms of insulin signaling in learning.

No description available.

Radiation induced bystander effects have changed our understanding of the biological effects of ionizing radiations. The original assumption was that biological effects require direct damage to DNA. The bystander effect eliminated that requirement and has become one main stream in radiation research ever since first reported over 20 years ago. Most bystander studies to date have been carried out by using conventional single cell in vitro systems , 2D cell array and 3D tissue samples, which are useful tools to characterize basic cellular and molecular responses. But to reveal the complexity of radiation responses and cellular communication, live animal models have many advantages. In recent years, models such as C. elegans and Zebrafish have been utilized in bystander effects research. In the Loma Linda/TAMU experiment, a L1 larva C. elegans model was devloped to study the radiation bystander effects by irradiating single intestine cell nuclei with a microbeam of protons. Due to the stochastic nature of particle interactions with matter and changing stopping power when protons slow down, precise dosimetry in the target nucleus is a difficult problem. This research was undertaken to provide a detailed description of the energy deposition in the targeted and surrounding non-targeted cell nuclei, and to evaluate the probabilities of the non-targeted cell nucleus being irradiated. A low probability is required to exclude the possibility of radiation biological an effect in non-targeted cells is caused by scattered particles. Mathematical models of the microbeam system and the worm body were constructed in this research. Performing Monte Carlo simulations with computer code, Geant 4, this research provided dosimetry data in cell nuclei in different positions and Geant 4, this research provided dosimetry data in cell nuclei in different positions and probabilities of scattering to non-targeted cell nuclei in various microbeam collima- tor configurations. The data provided will be useful for future collimated microbeam design.

Small molecules are powerful reagents for biological investigation. They provide an alternative to genetic perturbation and may offer more control over a target’s activity. C. elegans has recently gained prominence as a platform to discover new chemical tools. Through large-scale screens for compounds that induce phenotypes consistent with the disruption of conserved pathways, we identified two previously uncharacterized molecules of interest that we named nemadipine and migrazole. Here, I describe my efforts to understand their mechanism of action. Nemadipine is structurally analogous to 1,4-dihydropyridines (DHPs), which target the Cav1 calcium channel and are used clinically to lower blood pressure. Phenotypic and genetic evidence suggest that nemadipine targets the worm Cav1 channel, EGL-19. To identify the target of nemadipine in an unbiased manner, I performed a forward genetic screen for mutants resistant to its effects. The majority of the mutants from my screen had polymorphisms in EGL-19, providing additional evidence that it is the target of nemadipine. I also found that nemadipine is the only DHP that robustly elicits phenotypes in the worm. Therefore, I used this unique chemical to investigate the in vivo interactions between DHPs and the Cav1 channel. I identified residues in EGL-19 important for DHP-sensitivity in worms and showed that some of these residues are also important for mammalian DHP-interaction. Other labs have since exploited nemadipine’s in vivo properties to demonstrate new biological insights for EGL-19. Chemical genetic analyses indicated that migrazole disrupts multiple signal transduction pathways. This, together with experiments that I performed in yeast, suggests that migrazole may affect multiple pathways by perturbation of protein transport. To identify migrazole’s target, I performed a forward genetic screen for mutants resistant to migrazole’s effects. However, I was unable to identify the target of migrazole through analysis of the mutants I isolated. This result illustrates that while forward genetic screens can be very successful for target identification, their effectiveness is likely dependent on the nature of the compound-target interaction. My work shows that all aspects of developing a small molecule into a tool for biological analysis, from its discovery to its characterization, can be accomplished using C. elegans.

Small molecules are powerful reagents for biological investigation. They provide an alternative to genetic perturbation and may offer more control over a target’s activity. C. elegans has recently gained prominence as a platform to discover new chemical tools. Through large-scale screens for compounds that induce phenotypes consistent with the disruption of conserved pathways, we identified two previously uncharacterized molecules of interest that we named nemadipine and migrazole. Here, I describe my efforts to understand their mechanism of action. Nemadipine is structurally analogous to 1,4-dihydropyridines (DHPs), which target the Cav1 calcium channel and are used clinically to lower blood pressure. Phenotypic and genetic evidence suggest that nemadipine targets the worm Cav1 channel, EGL-19. To identify the target of nemadipine in an unbiased manner, I performed a forward genetic screen for mutants resistant to its effects. The majority of the mutants from my screen had polymorphisms in EGL-19, providing additional evidence that it is the target of nemadipine. I also found that nemadipine is the only DHP that robustly elicits phenotypes in the worm. Therefore, I used this unique chemical to investigate the in vivo interactions between DHPs and the Cav1 channel. I identified residues in EGL-19 important for DHP-sensitivity in worms and showed that some of these residues are also important for mammalian DHP-interaction. Other labs have since exploited nemadipine’s in vivo properties to demonstrate new biological insights for EGL-19. Chemical genetic analyses indicated that migrazole disrupts multiple signal transduction pathways. This, together with experiments that I performed in yeast, suggests that migrazole may affect multiple pathways by perturbation of protein transport. To identify migrazole’s target, I performed a forward genetic screen for mutants resistant to migrazole’s effects. However, I was unable to identify the target of migrazole through analysis of the mutants I isolated. This result illustrates that while forward genetic screens can be very successful for target identification, their effectiveness is likely dependent on the nature of the compound-target interaction. My work shows that all aspects of developing a small molecule into a tool for biological analysis, from its discovery to its characterization, can be accomplished using C. elegans.

In vertebrate and invertebrate systems, the role of glia-neuron interactions during development and behavior is becoming apparent. Recent studies have been aimed at characterizing glial-expressed proteins that affect the modulation of activities traditionally thought to be regulated by the neuron itself. The soil nematode Caenorhabditis elegans has recently emerged as an important invertebrate model to study glial roles in nervous system function and development. My dissertation work focuses on the characterization of HLH-17, a C. elegans basic helix-loop-helix transcription factor that is strongly and constitutively expressed in the glial cells that associate with four of the cephalic (CEP) neurons in the head of the animal. The CEP neurons are four of eight dopaminergic neurons with well characterized roles in the modulation of a number of behavioral activities in the worm. Although HLH-17 is required for neither the specification nor the development of the CEPsh glia or the CEP neurons, it does have a defined role during dopamine responses. We show that HLH-17 functions upstream of the dopamine receptors DOP-1, DOP-3 and the dopamine transporter DAT-1 to affect DA-dependent behaviors. Also, our microarray analyses provide preliminary evidence that HLH-17 targets factors responsible for receiving and transducing signaling molecules that are involved in the modulation of synaptic events in the worm nervous system. Together these results point to a role for HLH-17 in glia-neuron interactions in C. elegans. My dissertation studies therefore provide further support for the role of glial-expressed proteins in the regulation of activities mediated by the nervous system.