A Chemically Induced Colitis Screen Reveals the Necessity for Membrane Traffic in Intestinal Homeostasis




McAlpine, William Elliott

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Inflammatory bowel disease is most commonly a complex disorder caused by the interaction of environmental and genetic aberrations. Under normal conditions, a genetic program actively prevents inflammatory bowel disease, preventing invasion of microbes without permitting severe inflammation of the gut. To identify genes that maintain this balance, we performed a sensitized screen of 49,420 third generation (G3) germline mutant mice derived from N-ethyl-N-nitrosourea-mutagenized grandsires, bearing 104,658 coding/splicing mutations. We induced mild mucosal damage in these mice by orally administering dextran sodium sulfate (DSS) and found mutations that led to diarrhea and weight loss under these conditions. Causative mutations were mapped concurrently with screening using an automated mapping procedure. Among 114 DSS phenotypes identified and mapped, 36 have been validated by CRISPR/Cas9 targeting. Three vesicle trafficking genes, Myo1d, Smcr8, and Tvp23b, were selected for mechanistic evaluation. MYO1D is a class I myosin that binds both actin and lipid. MYO1D localizes to the basolateral membrane of enterocytes and functions in the intestinal epithelium to protect against colitis. SMCR8, along with C9ORF72 and WDR41, is a member of a tripartite complex that functions as a guanine exchange factor. SMCR8 localizes to the lysosome, and its absence results in perturbations to endocytic and phagocytic pathways. Hyperactivation of endosomal Toll-like receptors in Smcr8-/- mice causes spontaneous inflammation, and hyperactivation of multiple pathways contributes to DSS susceptibility. TVP23B is a trans-Golgi protein that binds YIPF6. Both TVP23B and YIPF6 are necessary for the formation of secretory granules in goblet and Paneth cells of the intestinal epithelium. These studies reveal non-redundant molecules required for the return of normal physiologic balance within the intestine after DSS insult.

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