Heterokaryons derived from polyethylene glycol-mediated fusion of myoblasts at different stages of development were used to investigate the transition of cells in the skeletal muscle lineage from the determined to the differentiated state. Heterokaryons were analyzed by immunofluorescence, using rabbit antibodies against the skeletal muscle isoforms of chicken creatine kinase and myosin, and a mouse monoclonal antibody that cross-reacts with chicken and rat skeletal muscle myosin. When cytochalasin B-treated rat L8(E63) myocytes (Konieczny S.F., J. McKay, and J. R. Coleman, 1982, Dev. Biol., 91:11-26) served as the differentiated parental component and chicken limb myoblasts from stage 23-26 or 10-12-d embryos were used as the determined, undifferentiated parental cell, heterokaryons exhibited a progressive extinction of rat skeletal muscle myosin during a 4-6-d culture period, and no precocious expression of chicken differentiated gene products was detected. In the reciprocal experiment, 85-97% of rat myoblast X chicken myocyte heterokaryons ceased expression of chicken skeletal muscle myosin and the M subunit of chicken creatine kinase within 7 d of culture. Extinction was not observed in heterokaryons produced by fusion of differentiated chicken and differentiated rat myocytes and thus is not due to species incompatibility or to the polyethylene glycol treatment itself. The results suggest that, when confronted in a common cytoplasm, the regulatory factors that maintain myoblasts in a proliferating, undifferentiated state are dominant over those that govern expression of differentiated gene products.

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