Summary Immunological control of mucosal barrier development and maintenance is a critical process, as imbalances in immune sentinel activity can lead to skin, lung and gut inflammatory diseases. Central in the mucosal tissue homeostasis are type 3 cytokine producing lymphocytes. Cytokines, cytokines receptors and master transcriptions involved in tissue development and barrier fortification have been identified based on analyses of loss of function models in human and mice. However, non-inflammatory factors that orchestrate the type 3 responses in tissues to assure proper immune reactions and prevent aberrant inflammation have not been well characterized. We discovered that oxysterols, a major class of cholesterol metabolites with established immunomodulatory activities, are the primary positional cues for the stereotypical type 3 lymphocytes in the skin called Tγδ17 cells. Ebi2 (encoded by Gpr183) is the G protein coupled receptor (GPCR) that senses specific types of oxysterols, and is expressed on all type 3 cytokine producing lymphocytes. Increased dietary cholesterol enhances Tgd17 activity at the skin interface via augmented oxysterol production and sensing. However, how tissue resident cells contribute to the anatomical framework that generate the final positional cues for dermal IL-17 secreting innate T lymphocytes is not known. Moreover, whether oxysterols are involved in the development of skin-tropic Tgd17 and other type 3 cytokine producing lymphocytes is not known. In this project we will test the hypothesis that Tgd17 cells maintain barrier tissue homeostasis by sensing cues that reflect nutritional states and inputs from the peripheral nerve system. Metabolic changes in the epithelial- neuronal niche in the skin control optimal microanatomical Tgd17 positioning and function via GPR183. This integrated sensory mechanism for cholesterol metabolism and neuronal inputs is imprinted early in Tγδ17 thymocyte maturation through interactions with a novel subset of thymic epithelial cells programmed to produce oxysterols. The conceptual basis of this project is innovative as there are no precedents for the oxysterol- regulated mucosal tissue hub integrating epithelial-neuronal-immune communications critical for barrier tissue fitness.

Relevance Excessive type 3 cytokine response is responsible for psoriasis and eczema, a skin disorder involving impaired skin barrier integrity. Obesity, often driven by intake of high fat western diet, is one of the non-genetic factors associated with morbidity and severity of these two chronic inflammatory skin diseases. Despite major progress in understanding the immunological effect of dietary lipids, little is known about the molecular mechanism linking high fat diet to tissue inflammatory diseases. We have discovered that dietary cholesterol and its metabolites oxysterols shapes innate T lymphocytes resident in the skin that are equipped with sensors of altered oxysterol homeostasis. We will test the hypothesis that the fine specificity of skin positioning of these dermal T cells guided by oxysterols produced in a specialized skin niche is obligatory for proper immune reactions. Skin-homing innate T cells are imprinted to sense cholesterol byproducts during their development in the thymus and we will seek to decipher the molecular features controlling T cell-thymic niche crosstalk. This project will establish the functional importance of tissue cholesterol metabolism to achieve mucosal tissue immune homeostasis and provide new intervention strategies to treat inflammatory disorders in the skin, lung and gut.