The goal of the proposed research is to understand the differentiative events which occur during the transition of muscle progenitor cells, myoblasts, into functional muscle cells and the manner in which this transition is coupled to the cell division cycle of the progenitor cells. Using synchronized myoblasts that block in G1 and differentiate in mitogen-free media but resume proliferation if refed with growth-promoting medium, studies have been initiated to determine: 1) Does withdrawal from the cell cycle require higher levels of Ca ions than are employed to block fusion? 2) Do myocytes eventually lose the ability to re-enter the cycle? and 3) Is myosin synthesis turned off when myocytes re-enter the cycle? As the synthesis of muscle-specific proteins serves as a major marker of differentiation in these studies, immunochemical probes for the major contractile proteins (and their ubiquitous homologues) are being developed using the monoclonal antibody technology. Hybridomas against several muscle proteins have been isolated and are currently undergoing characterization. Temperature sensitive mutants of a murine myoblast line have been isolated. These cell lines are being screened to determine the position of the TS block in the cell cycle. Mutants which block in G1 will be used to determine whether or not any specific portion of G1 is required for differentiation. The plasticity of gene expression in differentiated muscle cells is also being examined in an in vivo coupled transcription-translation system. Experiments in which Xenopus oocytes are injected with quail myoblast nuclei indicate that the synthesis of a discrete set of oocyte proteins may be repressed by the donor nuclei. These experiments will be extended to compare the protein synthetic patterns of control oocytes with those containing myoblast or differentiated myocyte nuclei.
