Little is known about the factors which regulate vascular smooth muscle (vsm) actin gene expression during skeletal myogenesis in culture. We have therefore looked for differences in the levels of accumulation of vsm actin mRNA among six mouse cell lines differing in apparent myogenic potential or in the complement of myogenesis determination genes which they express: NIH 3T3 and 10T1/2 non-myogenic fibroblasts and four myogenic lines--3T3-MyoD1 and 10EMc11s, MyoD/myogenin expressing sublines of the fibroblast lines, derived by transfer into the parent lines of a MyoD cDNA expression construct; C2C12, which expresses all four known myogenesis determination genes; and BC3H1, which expresses myf-5, myogenin, little herculin, and no MyoD. In differentiated cells of all four myogenic lines, vsm actin mRNA was expressed at levels dramatically higher than in growth-arrested NIH 3T3 cells, consistent with expression of vsm actin mRNA as an intrinsic part of the skeletal myogenic program somehow directed by myogenesis determination gene products. Interestingly, however, the level of vsm actin mRNA in growth arrested C3H10T1/2 fibroblasts was also dramatically higher than that in NIH 3T3. In view of these findings, and of the relative ease with which 10T1/2 as opposed to NIH 3T3 cells can be converted to myogenic lines, we hypothesize that factors which can act to regulate vsm actin gene expression in the absence of myogenesis determination gene expression may also influence the skeletal myogenic potential of the cells in which they are found. Among the myogenic lines, the ratio of vsm to skm actin mRNA was highest in BC3H1 cells, raising the possibility that were these cells forced to express MyoD and/or more herculin, as do the other myogenic lines, the ratio would decrease. Thus both fibroblast and myogenic lines will be useful for investigating the mechanisms controlling skeletal myogenesis and vsm and skm actin gene expression during myogenesis.