The fat-6 gene may play a role in a recently-discovered mitochondrial stress response pathway

AJAS · 2022 Animal Science

Eden Anne Bauer

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Overview

"Interactions Between Fatty Acid Metabolism and the ESRE Pathway in C. elegans" This study investigates the mechanisms and expression of a novel stress pathway found in Caenorhabditis elegans (C. elegans) that contains the conserved nucleotide motif TCTGCGTCTCT, termed ESRE (Ethanol and Stress Response Element). The ESRE pathway is a highly conserved mitochondrial stress mediator pathway, acting separately from other well-studied mitochondrial stress pathways. We hypothesized that fatty acid metabolism plays a role in the expression and/or regulation of the ESRE pathway. RNA interference was used in C. elegans to knock down the fatty acid metabolism genes fat-6, elo-2, cbp-1, cbp-3, acc-1, and fat-2. RNAi knockdown in the nematodes was accomplished by feeding C. elegans with E. coli carrying dsRNA plasmids. ESRE activity with and without mitochondrial stress was measured using a 3xESRE::GFP reporter with fluorescence measured across multiple timepoints. Mitochondrial stress was induced through exposure to two mitochondrial-targeting drugs: complex I inhibitor rotenone and the iron chelator phenanthroline. Dimethyl sulfoxide (DMSO) was used as the vehicle control. Preliminary data from two repetitions of the experiment showed that knocking down the gene fat-6 significantly reduced ESRE expression (p<0.0078) by ~25% (Rep 1) and 31% (Rep 2) in controls without drug treatment as well as with nematode exposure to rotenone (p<0.0036, ~63%, 44%) and phenanthroline (p<0.00016, ~19%, 27%), and that knocking down elo-2 significantly reduced ESRE expression (p<0.0059) with exposure to phenanthroline by ~20% (Rep 1) and 28% (Rep 2). Knocking down the genes cbp-1 and cbp-3 yielded inconsistent results. Nematodes with the cbp-1 RNAi knockdown also appeared smaller than the other worms, which could cause them to die faster than worms with other genes knocked down, increasing auto-fluorescence unrelated to ESRE expression. This study indicates that fat-6 or related processes in fatty acid metabolism may play a role in expression of the ESRE pathway.

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From the student

I am very excited and grateful to attend the annual AJAS conference for the fifth time this year. I first attended AJAS in eighth grade, and presented a project on the effects of antihistamines on memory in C. elegans. I continued that project the next year. The following year, I researched the effects of chemical sunscreens on chicken embryos. In mid-2020, I took on a computer-based project that I could complete at home, due to the pandemic, and used protein modeling to look into if llama nanobodies could be used to treat SARS-CoV-2, the virus behind the COVID-19 illness. Around this time, my family moved from New Hampshire to Houston, TX. Throughout the chaos of 2020 and 2021, science kept me grounded since it let me explore how our world worked as we all became more isolated and cut-off from it, and allowed me to search for truth in a time of so much uncertainty.

One of my favorite parts of my research journey has been getting to share my work while learning about others' projects, hypotheses, and areas of study.

From the student

I would like to thank the many kind and generous individuals who supported me throughout this research: my bench mentor, Dr. Elissa Tjahono, the Kirienko Lab Principal Investigator, Dr. Natasha Kirienko, all Kirienko Lab members, and my family. I am incredibly grateful for the opportunity to conduct this work in the BioSciences Department at Rice University.

At the New Hampshire Academy of Science, I would like to thank two longtime mentors, Dr. Kelly Salmon and Dr. Peter Faletra. NHAS is supported by the Science Education Partnership Award (SEPA) program, from the National Institute of General Medical Science (NIGMS) of the National Institutes of Health, as well as The Couch Family Foundation.

From the student

The Kirienko Lab is full of complex technology and many years' worth of research experience, knowledge, and technique to learn from, but one piece of equipment I am particularly in awe of is this worm-sorting machine.

If you have C. elegans suspended in a liquid—in this picture, they're in the black 96-well plate near the left bottom corner of the picture—the machine will carefully suck up the liquid and use tiny bursts of air to push individual worms past the a sensor that registers the number of worms you have as well as the size of each worm. This is very useful for ensuring your sample sizes are precise and accurate. C. elegans are otherwise hard to count in liquid since they're microscopic (with adults being ~1 mm in length) and because the worms "swim" (wiggle around) when immersed in a liquid.