Myristoylated alanine-rich C kinase substrate (MARCKS) is involved in myoblast fusion through its regulation by protein kinase Cα and calpain proteolytic cleavage

Abstract

MARCKS (myristoylated alanine-rich C kinase substrate) is a major cytoskeletal protein substrate of PKC (protein kinase C) whose cellular functions are still unclear. However numerous studies have implicated MARCKS in the stabilization of cytoskeletal structures during cell differentiation. The present study was performed to investigate the potential role of Ca(2+)-dependent proteinases (calpains) during myogenesis via proteolysis of MARCKS. It was first demonstrated that MARCKS is a calpain substrate in vitro. Then, the subcellular expression of MARCKS was examined during the myogenesis process. Under such conditions, there was a significant decrease in MARCKS expression associated with the appearance of a 55 kDa proteolytic fragment at the time of intense fusion. The addition of calpastatin peptide, a specific calpain inhibitor, induced a significant decrease in the appearance of this fragment. Interestingly, MARCKS proteolysis was dependent of its phosphorylation by the conventional PKCalpha. Finally, ectopic expression of MARCKS significantly decreased the myoblast fusion process, while reduced expression of the protein with antisense oligonucleotides increased the fusion. Altogether, these data demonstrate that MARCKS proteolysis is necessary for the fusion of myoblasts and that cleavage of the protein by calpains is involved in this regulation.

Transcriptional profile of GTP-mediated differentiation of C2C12 skeletal muscle cells

“…The myristoylated alanine rich protein kinase c substrate (Macs) gene, which expresses a major cytoskeletal protein substrate of protein kinase C, was upregulated at 24 h. MACS proteolysis is necessary for fusion of myoblasts and its cleavage is mediated by calpain [31]. The sarcoglycan, beta (dystrophin-associated glycoprotein) gene was up-regulated at 24 h; its protein product forms a complex with three further subunits on the skeletal muscle cell surface membrane.…”

TRPC1 regulates skeletal myoblast migration and differentiation

“…However, comparing the proteolytic pattern of these proteins in normal myoblasts and in myoblasts overexpressing calpastatin, Dedieu and collaborators found accumulation of MARCKS only in cells showing a reduced calpain activity, suggesting the involvement of this actin-binding protein in cell migration (Dedieu et al, 2003;Dedieu et al, 2004). Dulong and colleagues demonstrated the direct cleavage of MARCKS by calpain and its involvement in myoblast migration (Dulong et al, 2004). Calpain-dependent MARCKS proteolysis was also shown to activate its actin binding activity and therefore its ability to modulate actin dynamics and cell migration (Tapp et al, 2005).…”

“…Myristoylated alanine-rich C kinase substrate (MARCKS) is an actin-binding protein that is reported to be involved in myoblast fusion through its regulation by calpain proteolytic cleavage (Dulong et al, 2004). It accumulates in cells showing a reduced calpain activity, such as in calpastatin-overexpressing cells (Dedieu et al., 2004).…”

Calcium's Role in Mechanotransduction during Muscle Development

“…This process involves the activation of calpains, which are cysteine proteases activated by calcium, and the subsequent proteolysis of myristoylated alanine-rich C-kinase substrate (MARCKS), which is an actin-binding protein [104]. Indeed, the expression of calpastatin, a specific inhibitor of calpains, is decreased in fusing myoblasts [105], and when calpain activity is blocked, so is muscle differentiation [106].…”