Analyses of diverse stresses, including hypoxia, cyanide, and hypergravity in Caenorhabditis elegans

Organisms often encounter various forms of stress during their lifespan. The response to stress involves the regulation of cellular processes by stress response modulators that function to ultimately enable resistance and survival. In this dissertation I used Caenorhabditis elegans as a model to study the effects of a variety of stresses including the response to cyanide, hypoxia, and hypergravity. Chapter 2 of this thesis focuses on understanding the mechanisms of cyanide resistance in C. elegans. We employed a novel microfluidic device to describe the resistance phenotypes with greater spatio-temporal resolution. The results shed light on the underlying genetic bases that contribute to cyanide resistance, including the role of the hypoxia-inducible factor HIF-1. They also reveal new findings about the cyanide resistance phenotype, and help establish the applicability of microfluidic devices in studying the effects of aqueous toxicants in real-time. Chapter 3 is a study investigating the crosstalk between the stress response modulators HIF-1, DAF-16, and HLH29 in C. elegans. We found significant over-representation of DAF-16 target genes, as well as HLH29 target genes in our lists of genes up- or down- regulated by hypoxia or in animals with over-active HIF-1. Genes identified in this study are known to play important roles in the response and resistance to diverse forms of stress. The findings from this study illustrate the complex mechanisms employed by cells to regulate the expression of subsets of genes in the response to specific forms of stress. In chapter 4 we studied the effects of hypergravity exposure on C. elegans mobility, behavior, reproduction, and lifespan. We found that the animals rapidly recoverered mobility after short, intense bouts of hypergravity exposure, but their reproductive capabilities and lifespans were altered after longer durations of treatment. The results suggest that long term exposure to stress in the form of hypergravity may be detrimental to the animal\u27s health and physiology. Collectively, the results from my thesis help elucidate the important roles played by stress response mediators in contributing to an organism\u27s survival. They also illustrate the rich and complex ways in which organisms modulate their responses to various forms of stress

Introductory biology undergraduate students\u27 mixed ideas about genetic information flow

The core concept of genetic information flow was identified in recent calls to improve undergraduate biology education. Previous work shows that students have difficulty differentiating between the three processes of the Central Dogma (CD; replication, transcription, and translation). We built upon this work by developing and applying an analytic coding rubric to 1050 student written responses to a three‐question item about the CD. Each response was previously coded only for correctness using a holistic rubric. Our rubric captures subtleties of student conceptual understanding of each process that previous work has not yet captured at a large scale. Regardless of holistic correctness scores, student responses included five or six distinct ideas. By analyzing common co‐occurring rubric categories in student responses, we found a common pair representing two normative ideas about the molecules produced by each CD process. By applying analytic coding to student responses preinstruction and postinstruction, we found student thinking about the processes involved was most prone to change. The combined strengths of analytic and holistic rubrics allow us to reveal mixed ideas about the CD processes and provide a detailed picture of which conceptual ideas students draw upon when explaining each CD process

C. elegans SWAN-1 Binds to EGL-9 and Regulates HIF-1-Mediated Resistance to the Bacterial Pathogen Pseudomonas aeruginosa PAO1

Pseudomonas aeruginosa is a nearly ubiquitous human pathogen, and infections can be lethal to patients with impaired respiratory and immune systems. Prior studies have established that strong loss-of-function mutations in the egl-9 gene protect the nematode C. elegans from P. aeruginosa PAO1 fast killing. EGL-9 inhibits the HIF-1 transcription factor via two pathways. First, EGL-9 is the enzyme that targets HIF-1 for oxygen-dependent degradation via the VHL-1 E3 ligase. Second, EGL-9 inhibits HIF-1-mediated gene expression through a VHL-1-independent mechanism. Here, we show that a loss-of-function mutation in hif-1 suppresses P. aeruginosa PAO1 resistance in egl-9 mutants. Importantly, we find stabilization of HIF-1 protein is not sufficient to protect C. elegans from P. aeruginosa PAO1 fast killing. However, mutations that inhibit both EGL-9 pathways result in higher levels of HIF-1 activity and confer resistance to the pathogen. Using forward genetic screens, we identify additional mutations that confer resistance to P. aeruginosa. In genetic backgrounds that stabilize C. elegans HIF-1 protein, loss-of-function mutations in swan-1 increase the expression of hypoxia response genes and protect C. elegans from P. aeruginosa fast killing. SWAN-1 is an evolutionarily conserved WD-repeat protein belonging to the AN11 family. Yeast two-hybrid and co-immunoprecipitation assays show that EGL-9 forms a complex with SWAN-1. Additionally, we present genetic evidence that the DYRK kinase MBK-1 acts downstream of SWAN-1 to promote HIF-1-mediated transcription and to increase resistance to P. aeruginosa. These data support a model in which SWAN-1, MBK-1 and EGL-9 regulate HIF-1 transcriptional activity and modulate resistance to P. aeruginosa PAO1 fast killing