Myristoylated alanine-rich C kinase substrate (MARCKS) is involved in myoblast fusion through its regulation by protein kinase Cα and calpain proteolytic cleavage
Myristoylated alanine-rich C kinase substrate (MARCKS) is involved in myoblast fusion through its regulation by protein kinase Cα and calpain proteolytic cleavage
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DOI：
10.1042/bj20040347
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发表时间：
2004-09-15
影响因子：
4.1
通讯作者：
Cottin, P
中科院分区：
文献类型：
--
作者：
Dulong, S;Goudenege, S;Cottin, P
MARCKS（肉豆蔻酰基化富丙氨酸C激酶底物）是PKC（蛋白激酶C）的主要细胞骨架蛋白底物，其细胞功能尚不清楚。然而，许多研究表明，在细胞分化过程中，marks与细胞骨架结构的稳定有关。本研究旨在研究Ca2+依赖性蛋白酶（calpain）在肌肉形成过程中通过MARCKS蛋白水解的潜在作用。首次在体外证明MARCKS是钙蛋白酶底物。然后，在肌形成过程中检测MARCKS的亚细胞表达。在这种情况下，强烈融合时出现55 kDa的蛋白水解片段，导致MARCKS表达显著下降。加入钙蛋白酶肽（一种特定的钙蛋白酶抑制剂）可显著减少该片段的出现。有趣的是，marks蛋白水解依赖于传统的PKCalpha的磷酸化。最后，MARCKS的异位表达显著降低了成肌细胞融合过程，而反义寡核苷酸蛋白的表达降低了成肌细胞融合过程。总之，这些数据表明，marks蛋白水解对于成肌细胞的融合是必要的，calpain对该蛋白的裂解参与了这一调控。
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 Ca2+-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.