Abstract
In this paper, we investigated the isoform-specific roles of certain protein kinase C (PKC) isoforms in the regulation of skeletal muscle growth. Here, we provide the first intriguing functional evidence that nPKCδ (originally described as an inhibitor of proliferation in various cells types) is a key player in promoting both in vitro and in vivo skeletal muscle growth. Recombinant overexpression of a constitutively active nPKCδ in C2C12 myoblast increased proliferation and inhibited differentiation. Conversely, overexpression of kinase-negative mutant of nPKCδ (DN-nPKCδ) markedly inhibited cell growth. Moreover, overexpression of nPKCδ also stimulated in vivo tumour growth and induced malignant transformation in immunodeficient (SCID) mice whereas that of DN-nPKCδ suppressed tumour formation. The role of nPKCδ in the formation of rhabdomyosarcoma was also investigated where recombinant overexpression of nPKCδ in human rhabdomyosarcoma RD cells also increased cell proliferation and enhanced tumour formation in mouse xenografts. The other isoforms investigated (PKCα, β, ε) exerted only minor (mostly growth-inhibitory) effects in skeletal muscle cells. Collectively, our data introduce nPKCδ as a novel growth-promoting molecule in skeletal muscles and invite further trials to exploit its therapeutic potential in the treatment of skeletal muscle malignancies.
Highlights
Cell lysates of pooled cultures were subjected to Western blotting employing isoform-specific antibodies that corresponded to the overexpressed recombinant protein kinase C (PKC) isoforms, we found that the levels of the overexpressed PKCs (Fig. 1A) were several-fold higher than those of the respective endogenous ones
We investigated the effect of overexpression of the PKC isoforms on the proliferation of C2C12 myoblasts
We show that the overexpression of nPKCd enhanced the insulin-like growth factor-I (IGF-I)-induced ERK 1/2 phosphorylation (Fig. 6) compared to the cells overexpressing the dominant-negative mutant (DN-nPKCd) or the empty vector suggesting the involvement of nPKCd in mediating the growth-promoting effect of IGF-I
Summary
The above findings strongly suggested that in C2C12 myoblasts (i) the cPKCa and b isoforms inhibit proliferation and promote differentiation; (ii) the nPKCd, in contrast, markedly stimulates cell growth but inhibits differentiation; and (iii) the nPKCe plays an insignificant role in regulating the above processes.
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