Abstract
The functionally undefined Stac3 gene, predicted to encode a SH3 domain- and C1 domain-containing protein, was recently found to be specifically expressed in skeletal muscle and essential to normal skeletal muscle development and contraction. In this study we determined the potential role of Stac3 in myoblast proliferation and differentiation, two important steps of muscle development. Neither siRNA-mediated Stac3 knockdown nor plasmid-mediated Stac3 overexpression affected the proliferation of C2C12 myoblasts. Stac3 knockdown promoted the differentiation of C2C12 myoblasts into myotubes as evidenced by increased fusion index, increased number of nuclei per myotube, and increased mRNA and protein expression of myogenic markers including myogenin and myosin heavy chain. In contrast, Stac3 overexpression inhibited the differentiation of C2C12 myoblasts into myotubes as evidenced by decreased fusion index, decreased number of nuclei per myotube, and decreased mRNA and protein expression of myogenic markers. Compared to wild-type myoblasts, myoblasts from Stac3 knockout mouse embryos showed accelerated differentiation into myotubes in culture as evidenced by increased fusion index, increased number of nuclei per myotube, and increased mRNA expression of myogenic markers. Collectively, these data suggest an inhibitory role of endogenous Stac3 in myoblast differentiation. Myogenesis is a tightly controlled program; myofibers formed from prematurely differentiated myoblasts are dysfunctional. Thus, Stac3 may play a role in preventing precocious myoblast differentiation during skeletal muscle development.
Highlights
Skeletal muscle is composed of multinucleated cells called myofibers, which are formed from the fusion of myoblasts
We investigated the potential role of Stac3 in myoblast proliferation by siRNA-mediated knockdown of Stac3 in C2C12 myoblasts
To determine the potential role of Stac3 in myoblast differentiation, C2C12 myoblasts transfected with Stac3 siRNAs or scrambled siRNA were induced to differentiate, and their differentiation status was assessed by measuring the fusion index and average number of nuclei per myotube
Summary
Skeletal muscle is composed of multinucleated cells called myofibers, which are formed from the fusion of myoblasts. While Myf and MyoD determine the myogenic lineage of muscle progenitor cells [1,2,3], MyoG and Mrf drive the terminal differentiation and fusion of myoblasts into myotubes, the developing myofibers [4,5,6,7]. The functionally undefined Stac gene appears to be a new factor regulating skeletal muscle development and function. An essential role of Stac in myoblast differentiation does not appear to be supported by the facts that Stac knockdown or mutation did not prevent the formation of myofibers in zebrafish [12,14] and that Stac knockout did not block the formation of myofibers in mice [11,13]. Our results suggest an inhibitory role of Stac in myoblast differentiation and myotube formation
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