The Genomics of Stress-Induced Life Cycle Decisions in Nematodes

Organisms including bacteria, insects, and mammals make decisions to alter aspects of their development based on signals from the environment. The roundworm Caenorhabditis elegans can escape environmental collapse by halting reproductive growth and entering the stress-resistant dauer larval stage. Dauer larvae are spore-like and have specialized behaviors for finding and stowing onto carrier animals for dispersal. The decision to enter dauer is an anticipatory decision that is based on the inputs of food, pheromone, and temperature. Here, I show that touch is an overlooked input into the dauer entry decision. Using quantitative dauer entry assays on CRISPR knock-ins and existing mutants in mechanosensation, I demonstrate that gentle, harsh, and piezo touch promote dauer entry. By measuring pheromone sensation and signal tranmission in mechnanosensation-defective mutants, I show that mechanosensation likely inputs into the decision in parallel with pheromone. Further confirmation that touch promotes dauer entry is provided using direct mechanical stimulation of C. elegans, and I provide a plausible role for touch in sensing dauer-promoting weather and crowding conditions. Using RNA-seq, I also show that 8,042 genes are differentially expressed between dauer and reproductive development. Within this dataset, we observed the striking up-regulation of 64 neuropeptide genes (encoding 215 peptides) during dauer. By comparison, the entire human genome contains 97 neuropeptide genes (encoding 270 peptides). In particular, we observed coordinated up-regulation of the FMRFamide-like neuropeptides (FLPs). Using sbt-1 mutants to knock down neuropeptide processing, we demonstrate that peptidergic signaling promotes the dauer entry decision, promotes vigorous waving during the dauer-specific nictation behavior (carrier animal-hitchhiking), and is necessary for switching from repulsion to CO2 (a carrier animal cue) in non-dauers to CO2 attraction in dauers. By testing individual neuropeptides using CRISPR knockouts and existing strains, we show that 7 FLPs promote dauer entry while 4 FLPs inhibit. I therefore propose plausible roles for these FLPs in acting downstream of and/or modulating the sensation of food, pheromone, temperature, and touch inputs. We also demonstrate that FLP-10/FLP-17, which are expressed in the CO2-sensing BAG neuron, promote CO2 chemotaxis and nictation in dauers. These findings reveal that neuropeptides can alter decision-making and behavior during C. elegans dauer entry. Through a meta-analysis, we discovered similar up-regulation of FLPs in the dauer-like infective juveniles of diverse parasitic nematodes, suggesting that this may be an ancient mechanism for expanding the behavioral repertoire of nematodes. Further utilizing our RNA-seq dataset, I identified several markers for conveniently tracking and manipulating the dauer entry decision. These include col-183 (which tracks dauer fate in the hypodermis), ets-10 (neurons and intestine), nhr-246 (intestine and muscle), and led-1 (reproductive fate in hypodermis). Using condition shift experiments, we demonstrate that the dauer markers label animals during dauer-commitment. We show that these markers can be used to manipulate the entry decision by driving the reproduction-promoting gene daf-9/Cytochrome P450 under the control of the dauer-commitment markers. We further demonstrate that the markers can be used to track tissue coordination and its breakdown in partial dauer mutants, and propose strategies for using the markers to identify the intercellular signals that coordinate the dauer entry decision. I have discovered that the C. elegans dauer entry decision is more complex than previously realized, I have shown that C. elegans dauers obtain new behaviors through FLP signaling, and I have engineered tools for conveniently tracking and manipulating the dauer entry decision. My findings may illuminate how animals make robust decisions in uncertain environments, and have implications for how densely information and behaviors can be packed into a nervous system.