Involvement of protein phosphatase-1-mediated MARCKS translocation in myogenic differentiation of embryonic muscle cells.

Myristoylated alanine-rich C kinase substrate (MARCKS) translocates from the cytosol to the plasma membrane while mononucleated myoblasts fuse to form multinucleated myotubes. Here, we show that protein phosphatase-1-mediated dephosphorylation of MARCKS largely influences its subcellular localization and the fusion process. Treatment with okadaic acid or tautomycin, which are potent inhibitors of protein phosphatases and cell fusion, was found to reversibly block the MARCKS translocation. Moreover, the dephosphorylating activity against MARCKS markedly increased during myogenesis, and this increase was closely correlated with the membrane fusion of the cells. In addition, protein phosphatase-1 was identified as a major enzyme that is responsible for dephosphorylation of MARCKS. Furthermore, a mutation preventing MARCKS phosphorylation and thus facilitating MARCKS translocation resulted in promotion of the cell fusion. In contrast, overexpression of MARCKS carrying a mutation that blocks myristoylation and thus prevents the MARCKS translocation impaired the myoblast fusion. Together with the fact that MARCKS regulates the cytoskeleton dynamics by crosslinking the actin filaments in the plasma membrane and that myoblast fusion accompanies massive cytoskeleton reorganization, these results suggest that protein phosphatase-1-mediated MARCKS localization at the membrane is required for the fusion of embryonic muscle cells.