PROJECT SUMMARY Persistent muscle weakness and physical disability decrease quality of life for millions of Americans. These physical limitations are linked with pathological accumulation of inter/intramuscular adipose tissue (IMAT) across numerous populations and morbidities. This link combined with the resistance of IMAT accumulation to standard diet and exercise remodeling cues points to IMAT as being a persistent and unaddressed clinical problem, likely impeding rehabilitative efforts to maintain or increase muscle strength. The long-term objective of this research is to leverage our understanding of fat-muscle cross-talk to therapeutically modulate IMAT paracrine signaling to promote muscle growth and regeneration. Recently, IMAT has been identified as a specialized sub-type of fat called beige fat, that can be stimulated to adopt features of brown fat, potentially including the secretion of pro-regenerative factors. It is the primary objective of this proposal to determine the mechanisms of pro-regenerative action of brown fat on muscle and whether this action can be mimicked by IMAT stimulation. Our central hypothesis is that brown fat promotes muscle regeneration by enhancing myogenic differentiation via paracrine secreted follistatin. To address our hypothesis, our approach will use our novel system of mouse intermuscular fat transplant to model paracrine fat-muscle cross-talk. We will use this system to control the timing and composition of brown fat signaling for mechanistic exploration of the cellular response during muscle regeneration (aim 1) and the role of follistatin in mediating that response (aim 2). We will also explore the therapeutic potential of mimicking brown fat paracrine signaling via stimulation in mouse and human IMAT (aim 3). Altogether, these studies will broaden our understanding of the basic biology of brown fat signaling, the nature of IMAT-muscle cross-talk and the therapeutic potential for stimulating IMAT and other depots of beige fat to improve tissue physiology.