Both Myoblast Lineage and Innervation Determine Fiber Type and Are Required for Expression of the Slow Myosin Heavy Chain 2 Gene

Abstract

Skeletal muscle fibers express members of the myosin heavy chain (MyHC) gene family in a fiber-type-specific manner. In avian skeletal muscle it is the expression of the slow MyHC isoforms that most clearly distinguishes slow- from fast-contracting fiber types. Two hypotheses have been proposed to explain fiber-type-specific expression of distinct MyHC genes during development-an intrinsic mechanism based on the formation of different myogenic lineage(s) and an extrinsic, innervation-dependent mechanism. We developed a cell culture model system in which both mechanisms were evaluated during fetal muscle development. Myoblasts isolated from prospective fast (pectoralis major) or slow (medial adductor) fetal chick muscles formed muscle fibers in cell culture, none of which expressed slow MyHC genes. By contrast, when muscle fibers formed from myoblasts derived from the slow muscle were cocultured with neural tube, the muscle fibers expressed a slow MyHC gene, while muscle fibers formed from myoblasts of fast muscle origin continued to express only fast MyHC. Motor endplates formed on the fibers derived from myoblasts of both fast and slow muscle origin in cocultures, and slow MyHC gene expression did not occur when neuromuscular transmission or depolarization was blocked. We have cloned the slow MyHC gene that is expressed in response to innervation and identified it as the slow MyHC 2 gene, the predominant adult slow isoform. cDNAs encoding portions of the three slow myosin heavy chain genes (MyHC1, slow MyHC 2, and slow MyHC 3) were isolated. Only slow MyHC 2 mRNA was demonstrated to be abundant in the cocultures of neural tube and muscle fibers derived from myoblasts of slow muscle origin. Thus, expression of the slow MyHC 2 gene in this in vitro system indicates that formation of slow muscle fiber types is dependent on both myoblast lineage (intrinsic mechanisms) and innervation (extrinsic mechanisms), and suggests neither mechanism alone is sufficient to explain formation of muscle fibers of different types during fetal development.

Proximal Sequences of the Aldolase A Fast Muscle-specific Promoter Direct Nerve- and Activity-dependent Expression in Transgenic Mice

“…This has been previously suggested by the studies of the expression patterns of muscle sarcomeric genes during regeneration in the presence or the absence of nerve (45) or after muscle stimulation (43,44). Recently, it has been shown that in chicken myotubes, the expression of the myosin heavy chain 2 slow gene depends on both intrinsic (type of myoblasts) and extrinsic (nerve-induced contraction) factors (46). The present study shows that the precise regulation of pM results from the combination of different transcriptional regulatory elements associated with these different programs whose relative contribution may evolve during muscle maturation.…”

Sox6 is required for normal fiber type differentiation of fetal skeletal muscle in mice

“…Because differentiation of fiber types in fetal skeletal muscle appears to be regulated by both the intrinsic genetic programs within myoblasts and the extrinsic local environment (DiMario and Stockdale, 1997;Robson and Hughes, 1999;Pin et al, 2002), we want to determine whether Sox6 is involved in intrinsic myogenic commitment or/and in regulating the extrinsic signals. To begin to answer this question, we have obtained Sox6 conditional knockout mice (Dumitriu et al, 2006a) to inactivate the Sox6 gene in a myogenic cell-specific manner.…”

Regulation of skeletal muscle fiber type and slow myosin heavy chain 2 gene expression by inositol trisphosphate receptor 1

“…This was most evident in the curare-treated fetal MA in which secondary muscle fibers did not contain detectable levels of slow MyHC2 (DiMario and Funk, 1999). Normal innervation-dependent regulation of slow MyHC2 gene expression was observed in vitro by innervation of MA muscle fibers and subsequent slow MyHC2 gene expression (DiMario and Stockdale, 1997). Fast PM muscle fibers in vitro were resistant to innervation-induced slow MyHC2 gene expression, suggesting a muscle fiber typespecific mechanism of slow MyHC2 gene repression in these fibers.…”

“…Spinal cord explants containing motor neurons from E5 chick embryos were added to some of the cultures on day 3 of incubation as previously described (DiMario and Stockdale, 1997). On day 4 of incubation, medium for some of the cultures was supplemented with 100 µM 2-aminoethoxydiphenylborate (2-APB) (Sigma) or 10 µM xestospongin D (Calbiochem).…”