Molecular mechanisms of muscle stem cells transitioning intoquiescence

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DESCRIPTION (provided by applicant): The mammalian musculature provides mechanical force for locomotion. Muscles display remarkable ability to regenerate from reservoirs of muscle stem cells. How muscle stem cells transition from an actively proliferating population during development into a reserved population in adulthood for muscle repair/regeneration throughout lifetime is not known. I have obtained evidence that, in the mouse model, this process depends on the function of the Pax7 gene, which encodes a transcription factor. I plan the following aims to define 1) the precise role of Pax7, 2) the transcriptome changes during the development-to-adult transition, and 3) other essential genes (acting in parallel or downstream of Pax7) that mediate this transition. Aim 1: Determining the cellular and molecular defects of Pax7 mutant satellite cells. When and how do Pax7 mutant cells become defective in the development-to-adult transition? I will combine a plethora of assays for determining several muscle stem cell properties, including muscle differentiation, cell proliferation, cell death, niche occupancy, and cell polarity. These assays will be conducted with time course studies from newborn to adult. Aim 2: Performing mRNA-seq of Pax7 control and Pax7 mutant satellite cells. What are the transcriptome changes during this transition and what are the specific processes governed by Pax7? A comparative time course study of transcriptomes between control and Pax7 mutant cells throughout the development-to-adult transitional period will be performed using mRNA-seq, followed by bioinformatic analysis. Aim 3: Screening candidate genes for their function in satellite cells. What are the genes (in addition to Pax7) critical for the development-to-adult muscle stem cell transition? From Aim 2, I will select up to 50 genes to test for their function in this aspect; they may either act independently or downstream of Pax7. I will develop a novel assay for rapid screening of these genes' functions by combining gene knock-down and intramuscular transplantation of muscle stem cells. PUBLIC HEALTH RELEVANCE: Muscle stem cells provide the replenishing source for muscle regeneration after injury. This proposal aims to determine the genes that control the genesis of muscle stem cells, using the mouse as a model. Results from this study will have clinical implications for stem cell based therapies of muscular dystrophies and of acute myotrauma from combat or accident.