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Apoptotic myonuclei appear during myogenesis and in diseased muscles. To investigate cell death regulation in skeletal muscle, we examined how members of the Bcl-2 family of apoptosis regulators are expressed and function in the C2C12 muscle cell line and in primary muscle cells at different stages of development. Both anti-apoptotic (Bcl-W, Bcl-X(L)) and pro-apoptotic (Bad, Bak, Bax) members of the Bcl-2 family were expressed in developing skeletal muscle in vivo. Each was also expressed in embryonic (E11-12), fetal (E15-16), and neonatal muscle stem cells, myoblasts, and myotubes in vitro. In contrast, Bcl-2 expression was limited to a small group of mononucleate, desmin-positive, myogenin-negative muscle cells that were seen in fetal and neonatal, but not embryonic, muscle cell cultures. The cell surface protein Sca-1, which is associated with muscle and blood stem cells, was found on approximately 1/2 of these Bcl-2-positive cells. Loss of Bcl-2 did not affect expression of other family members, because neonatal muscles of wild-type and Bcl-2-null mice had similar amounts of Bcl-X(L), Bcl-W, Bad, Bak, and Bax mRNAs. Loss of Bcl-2 did have functional consequences; however, because neonatal muscles of Bcl-2-null mice had only approximately 2/3 as many fast muscle fibers as muscles in wild-type mice. Thus, Bcl-2 function is required for particular stages of fetal and postnatal myogenesis., (Copyright 2001 Wiley-Liss, Inc.)

The four muscle regulatory factors (MRFs) of the MyoD family are expressed in distinct temporal and spatial patterns in developing somites. To examine MRF function and regulation in somites, we generated myogenin promoter-MRF4 transgenic mice in which MRF4 was expressed in rostral somites about a half day earlier than normal. We found that the transgene, which was expressed at about the same level as endogenous MRFs, did not noticeably alter developing or adult mice, whereas the rostral somites of transgenic embryos showed accelerated myocyte formation, as well as precocious expression of the endogenous MRF4 gene. In an individual transgenic somite, MRF4 was expressed in both presumptive myotomal (mesenchymal) and dermatomal (epithelial) cells. Transgenic dermatomal cells also contained myogenin, which is expressed early in myogenesis, but did not contain myosin, which is expressed late in myogenesis. In transgenic myotomal cells, in contrast, precocious expression of MRF4 accelerated late events in myogenesis, including myosin expression and striated myofibril formation. MRF function, therefore, appears to be differentially regulated in dermatomal and myotomal cells.

Skeletal muscle formation is studied in vitro with myogenic cell lines and primary muscle cell cultures, and in vivo with embryos of several species. We review several of the notable advances obtained from studies of cultured cells, including the recognition of myoblast diversity, isolation of the MyoD family of muscle regulatory factors, and identification of promoter elements required for muscle-specific gene expression. These studies have led to the ideas that myoblast diversity underlies the formation of the multiple types of fast and slow muscle fibers, and that myogenesis is controlled by a combination of ubiquitous and muscle-specific transcriptional regulators that may be different for each gene. We further review some unexpected results that have been obtained when ideas from work in culture have been tested in developing animals. The studies in vivo point to additional molecular and cellular mechanisms that regulate muscle formation in the animal.