Clinical trials utilizing the transfer of myoblasts for the treatment of debilitating myopathies have yielded disappointing results. Failure of these therapies reflects an inadequate knowledge of the fundamental regulatory mechanisms of myogenic stem cells. We have recently described a novel marker for the myogenic stem cell population and have named it myocyte nuclear factor (MNF). Using a gene knockout strategy, we have observed that mice without MNF display a growth deficit, impaired muscle regeneration, dysregulation of p21CIP and perturbed cell cycle progression in the myogenic stem cell. Furthermore, we have determined that an upstream fragment of the MNF gene confers expression in the myogenic stem cell population of adult skeletal muscle. These data strongly suggest that MNF is an important transcription factor in the regulation of myogenic stem cells. The principle hypothesis of this proposal is that MNF regulates the cell cycle of the myogenic stem cell and modulates muscle regeneration. In the proposed studies, we will evaluate the regulation of MNF gene expression in the myogenic stem cell population and analyze the cell cycle progression of this cell population to enhance our understanding of the regulation of muscle specific stem cells and muscle regeneration. To accomplish our goals we will pursue the following three specific aims: l) To define the molecular basis for selective expression of MNF in the myogenic stem cell, 2) To define the profile of gene expression and cell cycle progression in wild-type and MNF mutant myogenic stem cells; 3) To define the regulatory interaction between MNF and the cell cycle dependent kinase inhibitor, p21CIP. The experimental plans have been designed to take maximum advantage of emerging technologies and powerful model systems. The experiments we propose are hypothesis-driven and focused. The results of this analysis, using unique transgenic mouse models, will enhance our understanding of muscle stem cell biology and have therapeutic applications in the treatment of debilitating myopathies.
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