Regionalized expression of myosin isoforms in heterotypic myotubes formed from embryonic and fetal rat myoblasts in vitro.

The development of mammalian limb muscles involves the appearance and fusion of at least two separate populations of muscle precursor cells. These two populations, termed embryonic and fetal myoblasts, are first detected within the limb bud at different stages of development. We have previously demonstrated that, in the rat, each myoblast population expresses a unique pattern of myosin heavy chains (MyHCs) during differentiation in vitro (Pin and Merrifield [1993] Dev. Genet. 14:356-368). Embryonic myoblasts accumulate embryonic and slow MyHCs, whereas fetal myoblasts accumulate embryonic, neonatal, and adult fast MyHCs but not slow MyHC. To determine if the two populations can fuse with each other and whether the pattern of MyHC expression is altered in the resulting heterokaryons, embryonic and fetal myoblasts were labelled with the lipophilic dye PKH26, [3H]-thymidine, or 5-bromodeoxyuridine (BRDU) and cocultured for 24-48 hr. Our results demonstrate that fusion occurs between embryonic and fetal myoblasts in vitro. Moreover, analysis of the resulting heterokaryons revealed regionalized accumulations of MyHC around individual nuclei. Interestingly, these accumulations were typical of the default pattern of expression that individual nuclei would have normally expressed in single culture. Nuclei contributed by embryonic myoblasts were surrounded by localized accumulations of slow MyHC, whereas nuclei from fetal myoblasts were surrounded by neonatal/fast MyHC. The occurrence of such nuclear domains indicates that the myoblast-specific expression of MyHC isoforms is dictated by cis-acting factors established prior to fusion.