Project Summary The nematode Caenorhabditis elegans is one of the most important model organisms for biomedical research, because of its biological tractability and because much of its physiology and behavior relies on signaling pathways that are conserved in humans. The goal of this project is to complement the highly developed genomics and genetics of C. elegans with a comprehensive structural and functional characterization of its metabolome. In recent work we have shown that C. elegans utilizes small- molecule architectures of unanticipated diversity and complexity in endocrine and exocrine signaling that control almost every aspect of its life history, including development, aging, stress resistance, and a wide range of behaviors. One major focus of our investigations is the elucidation of the biosynthesis and perception mechanisms of newly identified small molecule signals, which will reveal how metabolism and conserved signaling pathways interact to control phenotypes. Of particular interest are (i) the roles of recently identified endogenous agonists of the nuclear receptor NHR-49, a key regulator of lipid metabolism and functional ortholog of vertebrate PPARα, (ii) male germline- dependent metabolites that accelerate development and aging via conserved pathways, including homologs of mammalian steroid receptors, and (iii) our recent discovery of a modular glucosides that appear to function as a “second layer” of neurotransmitter signaling. In addition, we will investigate the biological functions of a new family of nucleoside derivatives we discovered that may interact with purinergic signaling. Another focus area will be the influence of dietary bacteria on C. elegans small molecule signaling, motivated by our discovery of unexpected roles for bacterial metabolites, e.g. in modulating fat metabolism via NHR-49 or in serotonin signaling. Central to the proposed research is the use of synthetic samples and derivatives of identified signaling molecules for bioassays, mutant screens, and transcriptomic studies, as well as the in-house developed metabolomics platform (Metaboseek), which greatly accelerates compound identification and functional annotation. Successful conclusion of this project will provide a structural and functional annotation small molecule signaling networks in C. elegans, substantially increasing our understanding of conserved pathways that control development, aging and metabolism, as well as corresponding disease-relevant pathways in mammals. The small-molecule knowledge generated will not only enable future efforts aimed at more varied chemical genetic screens exploring additional aspects of the biology and ecology of C. elegans, but also of nematode species relevant in agriculture or medicine. Furthermore, methodology developed for characterizing C. elegans signaling molecules will facilitate similar studies toward structural and functional characterization of small molecule metabolites from other model organisms.