SUMMARY The over-arching goal of this proposal is to delineate the signaling networks mediating the instructive role of the myofiber on MuSC function. MuSC, also known as satellite cells, are the main source of skeletal muscle growth and regeneration. In healthy adult tissues MuSC exist in a quiescent state, and upon stress or injury they are activated to proliferate and generate large numbers of progenitors to efficiently repair damaged muscles. While this process is efficient in healthy conditions, in several diseased conditions and during aging the numbers and function of MuSC are impaired. Thus, there is a major need to understand the molecular networks regulating their function, in order to identify novel tools or targets to enhance their tissue repair potential that can be utilized in regenerative medicine approaches. Significant efforts in the recent years have defined a critical role of the microenvironment in regulating MuSC function, by uncovering the integrated coordination of several tissue- resident cells, such as inflammatory cells, fibroadipogenic progenitors and endothelial cells in regulating MuSC tissue repair function. However, the role of the myofiber within the tissue microenvironment and in the MuSC niche is still poorly defined. Our preliminary findings provide evidence that the E3 ubiquitin ligase Fbxw7 in myofibers regulates MuSC pool size in a cell non-autonomous manner, suggesting that the myofiber plays a major role in instructing stem cell function. We further identify PGC1alpha as a direct Fbxw7 target and the upregulation of the PGC1alpha target irisin, a myokine that promotes MuSC differentiation. The identification of signaling pathways that mediate the instructive role of the myofiber on MuSC pool size could identify strategies to expand MuSC, thus allowing their use in regenerative medicine approaches. The objective of this application is to take advantage of genetic models, unbiased proteomics and ubiquitinome profiling, gene expression profiling and cellular biology techniques in order to delineate the signaling networks mediating the instructive role of the myofiber on MuSC function. The central hypothesis to be tested is that the Fbxw7 in myofibers regulates MuSC numbers in a cell non-autonomous manner by altering the MuSC niche. This integrated approach will improve our understanding of myofiber/MuSC interactions, define the signaling networks mediating this process and evaluate Fbxw7/PGC1?/Irisin as a critical axis to regulate MuSC pool size.