Treatment of human muscle myotube cultures with 2 microM-cytochalasin D (CD) for 6 h stimulated synthesis of both the (muscle-specific) alpha-actin and the (non-muscle) beta and gamma-actins usually expressed by these cells. In non-muscle (HEp-2) cell cultures, CD enhanced synthesis of beta and gamma-actin, but did not induce synthesis of alpha-actin, which is not normally present in these cells. Thus, synthesis of both muscle and non-muscle actins can be increased by CD, but enhancement of actin synthesis results from increases in the isoactins usually present, rather than induction of new isotypes. Comparison of CD-treated (fused) myotube cultures with (unfused) myoblast cultures indicated that beta and gamma-actin synthesis was similarly enhanced in both cultures, but that alpha-actin synthesis was stimulated to a greater extent in the myoblast cultures. Desmin synthesis was also stimulated in the myoblasts but not the myotubes, suggesting that the effect of CD on synthesis of these developmentally regulated cytoskeletal proteins (alpha-actin, desmin) might be modulated by fusion or the state of differentiation of the muscle cell.