Muscle injury stimulates normally quiescent muscle satellite cells to re-enter the cell cycle and execute the myogenic program, resulting in the restoration of muscle structure and function. While the essential function of satellite cells in muscle repair has been recognized for many years, fundamental questions concerning their embryological origin, developmental potential, and mechanism of renewal remain unresolved. The present proposal uses unique transgenic mouse lines and lineage tracing approaches based on Cre/loxP recombination to investigate these key aspects of satellite cell biology in vivo. To investigate the competing hypotheses that satellite cells are derived from fetal myoblasts and cells of the vasculature (e.g. endothelial precursors), relevant cell types will be specifically and permanently labeled with lacZ by intercrossing two mouse strains, transgenic mice that express the gene for Cre recombinase under the control of a cell type- specific promoter/enhancer (MyoD-cre for myoblasts, Tie2-cre and Flk1-cre for vascular-derived cells) and a reporter mouse strain in which lacZ expression is Cre-dependent. Whether these cells represent satellite cell progenitors will be determined by assessing p-galactosidase labeling of satellite cells using transmission electron microscopy, and by standard and confocal microscopy of single-fiber preparations. Similar studies in injured muscle will determine whether the satellite cell pool is maintained by self-renewal, or by a stem cell source within or outside of skeletal muscle. The developmental potential of satellite cells will be determined by tracing the fates of satellite cells following muscle injury and in single-fiber cultures. Experiments in wild- type mice will be compared to parallel experiments in mice that are mutant for either MyoD or Myf-5 to test the function of these muscle regulatory genes in satellite cell myogenic commitment. Analysis of MyoD and Myf-5 functions in the embryo will define the full repertoire of cell fate choices available to embryonic myogenic precursors in the absence of these regulatory genes and will provide new comparative insights into the regulation of embryonic and postnatal myogenesis. Collectively, these studies will significantly advance current knowledge of satellite cell biology and will provide a foundation for future investigations into the signaling and gene transcriptional pathways that regulate cell fate choices of satellite cells and their progenitors.
