Acetyl-CoA carboxylase beta is expressed primarily in heart and skeletal muscle, where it may play a role in the control of mitochondrial fatty acid uptake and oxidation by controlling carnitine-palmitoyl-CoA transferase 1, To investigate the relationship between ACC-beta expression and myoblast differentiation, we specifically blocked ACC-beta production by expressing an antisense gene to the specific N-terminal region of ACC-beta in H9c2 cells. The expression of the antisense ACC-beta mRNA retarded cell fusion, myosin formation, and creatine kinase induction; all of these parameters are hallmarks of muscle cell differentiation. On the other hand, the rate of fatty acid oxidation was not affected by the presence or absence of ACC-beta in the cells, (C) 1999 Academic Press.