Extrinsic Regulation of Muscle Stem Cell Quiescence

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Stem cells maintaining and repairing adult tissues have a strong dependency on signals provided by the surrounding microenvironment, often referred to as the niche. In skeletal muscle this niche regulation of stem cells is particularly complex and depends on a multitude of different cell types secreting growth factors and instructive extracellular matrix (ECM) in a tightly regulated spatio-temporal manner. Fundamental properties of muscle stem cells (MuSCs), such as quiescence, self-renewal and the ability to differentiate, are determined by the composition of the niche. However, in spite of its important role for the function of MuSCs, the architecture and the instructive signals the niche generates remain incompletely understood. Notably, it remains particularly enigmatic which cellular and extracellular cues in the niche control MuSC quiescence, the important dormant maintenance state that is essential to the life-long ability of mammals to maintain their muscles and recover from injuries. A major focus of our proposed NSERC research program will be to fill this gap in our biological understanding of the niche by studying the microenvironmental regulation of quiescent MuSCs.******Our recent progress and preliminary experiments have revealed that quiescent MuSCs employ a receptor called dystroglycan (DAG) to adhere to the niche ECM. We observed that loss of DAG in a tamoxifen inducible MuSC specific DAG knockout mouse model (iDAG-KO) leads to a break quiescence and to premature differentiation. Importantly, we also discovered that mice deficient for the DAG ligand Laminin alpha 2 (LAMA2) have hyperactivated excessively differentiating MuSCs. Here we propose a number of objectives to investigate the molecular mechanisms through which DAG and its ECM ligand LAMA2 maintain MuSC quiescence. ******Specific objectives are:******(I) To study the role of DAG as an ECM receptor for MuSCs and provide insights into the pathways it induces to maintain the quiescent state. This objective will be based on the analysis of iDAG-KO mice and will rely on novel hydrogel based assays that allow to study MuSCs quiescence in culture.******(II) To determine the role of the abundant DAG ligand LAMA2 in the quiescent MuSC niche. This objective involves the in-vivo and in-vitro interrogation of the hyperactivated phenotype MuSC in a LAMA2 depleted environment. ******(III) To study the deposition dynamics and cellular sources of LAMA2 in muscle tissue. This objective is based on the generation and analysis of an innovative reporter mouse model for LAMA2.******Taken together, the experiments outlined in this proposal will provide fundamental insights into the niche regulation of quiescent MuSCs that may well be extrapolated from skeletal muscle to other tissues. In addition, our work will contribute to a better understanding of the biology of Laminins with all their important implications for development, organ structure and function. **