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Cultured chick embryo skeletal muscle cells normally synthesize only the embryonic isoform of mysoin. We have found that aneural muscle cultures that become or are provoked into an extremely contractile state will begin to synthesize a pattern of myosin light chains typical of maturing muscle. Immunoblots with neonatal and adult specific monoclonal antibodies did not reveal a corresponding isozyme transition in myosin heavy chain. These results demonstrate a correlation between contractility and the regulation of myosin light chain maturation, and also suggest that the transitions of heavy and light chain synthesis during development do not appear to be under close coordinate regulation.

The synthesis and accumulation of myosin was studied in subcultures of fusion-blocked, postmitotic embryonic chicken myogenic cells. Electron micrographs and fluorescent microscopy with antimyosin revealed that most, if not all, of these cells contain myosin. It was also found that these cells are capable of accumulating myosin at rates comparable to fused cells. Incipient T-tubule formation was also present in some of the blocked cells. It is concluded that cell fusion is not a prerequisite for myosin synthesis and accumulation or T-tubule formation during myogenesis in vitro .

Publisher Summary In spite of the diversity of non-muscle cells, which have been shown to synthesize myosin or myosin-like peptides, it remains appropriate and indeed extremely useful to focus on myosin polypeptides as unique markers for terminal myogenesis. The basis for continuing to use myosin as a marker is the immunological specificity of myosin peptides and the lack of cross-reactivity of antibodies prepared for skeletal muscle myosin with myosins from other cell types. Terminal myogenesis involves (1) withdrawal of myoblasts from the cell cycle, (2) fusion of these myoblasts to form the embryonic myotube or precursor to the cross-striated muscle fiber, and (3) the initiation of the synthesis of unique peptides of myofibril. In some cases, the synthesis of unique peptides may occur prior to cell fusion, but fusion remains one of the major morphological markers for terminal skeletal muscle cell differentiation in most experimental systems. Myosin is the major structural protein of skeletal muscle and measurement of its synthesis and accumulation has been used in vivo and in vitro as a biochemical marker for the transition of the myoblast or myotube to the fully differentiated state. Although antimyosin has been used to quantitate myosin synthesis during normal myogenesis in vitro and in vivo, one of the more valuable uses of this antibody has been in the determination of myosin synthesis in unfused myogenic cells.

Messenger RNA which stimulates the synthesis of myosin heavy chain in a reticulocyte lysate has been isolated from single myogenic cell cultures. Specific myosin polypeptides have been identified by immunoprecipitation with an antibody made to purified adult chicken skeletal muscle myosin. This mRNA binds to oligo(dT)-cellulose, and an active fraction from sucrose gradients migrates as 26S on formamide-polyacrylamide gels. The relative amount of this RNA increases dramatically at the time of terminal differentiation.