Collaborative research: Signaling mechanisms in the crustacean molting gland

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Crabs, lobsters, and shrimp are ecologically and economically important crustaceans in marine environments. The hard, calcified exoskeleton provides protection and support, but restricts growth. As a result, these animals must periodically shed the exoskeleton, a process called molting. Upon molting, animals stretch the new exoskeleton before it hardens, providing more space for tissue growth. The entire process is controlled by molting hormone produced by a pair of molting glands. The activity of the molting gland is controlled by environmental signals mediated by the nervous system. Much remains to be known about the signaling genes that control the molting gland, in particular the genes required for (1) committing the animal to molt and (2) repressing the molting gland after the animal molts. Commitment is a “point of no return” decision that is critical for survival and growth in all crustaceans. Repression prevents molt induction until synthesis and calcification of the exoskeleton is completed. This collaborative project involves a team of investigators from four universities who will use state-of-art DNA and peptide sequencing technologies to identify genes and proteins essential for the activation, commitment, and repression of the molting gland and its regulation by environmental signals. The data generated will be made available to researchers so that they can better understand how to manage fisheries, develop effective aquaculture practices, and mitigate the effects of pollutants and climate change. Three postdoctoral fellows, four graduate students, and 6-8 undergraduates will receive training in advanced molecular techniques and bioinformatics. An educational outreach program for teachers recruited from junior high and high schools in northeastern Colorado will be developed.In crustaceans, paired molting glands (Y-organs or YOs) produce ecdysteroids necessary for systemic molt cycle control. Molt-inhibiting hormone (MIH), produced in the eyestalk ganglia, inhibits the synthesis of ecdysteroids by the YO via a putative G protein-coupled receptor (GPCR) and cyclic nucleotide second messengers. The project will use transcriptomic and proteomic tools to determine the signaling mechanisms that drive YO phenotypic changes over the molt cycle; these are designated basal in intermolt, activated in early premolt, committed in mid and late premolt, and repressed in postmolt. The specific aims are to: (1) determine how MIH signaling inhibits Mechanistic Target of Rapamycin (mTOR) activity; (2) determine the role of Transforming Growth Factor beta/Activin signaling in YO commitment; (3) determine the role of ecdysteroids on YO entry into, and exit from, the repressed state; and (4) characterize the MIH receptor. The first three aims will quantify the phenotypic effects of experimental signaling manipulations on YO state transitions. Network analysis of RNA-sequence and liquid chromatography-tandem mass spectrometry data will determine the effects of experimental treatments on gene interactions and downstream targets of signaling pathways. Aim #4 uses transcriptomic tools and a luciferase ligand/receptor binding assay with recombinant neuropeptides to characterize candidate GPCRs for receptor function. A thorough understanding of the hormonal regulation of decapod molting and growth is essential to manage fisheries, develop effective aquaculture practices, and mitigate potential effects of pollutants and environmental factors on an economically and ecologically important group of marine organisms. Three postdocs, four graduate students, and 6-8 undergraduates will receive training in advanced molecular techniques and bioinformatics, and a grade 6-12 teacher will be involved in educational outreach on the research topic.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.