P2Y6 regulates cytoskeleton reorganization and cell migration of C2C12 myoblasts via ROCK pathway

Migration of skeletal muscle precursor cells is required for limb muscle development and skeletal muscle repair. This study aimed to examine the role of P2Y6 receptor in C2C12 myoblasts migration. C2C12 myoblasts were treated with P2Y6 agonist UDP, P2Y6 antagonist MRS2578, Ca2+ channel blocker BTP2,...

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Published in:Journal of cellular biochemistry 2018-02, Vol.119 (2), p.1889-1898
Main Authors: Wang, Wei, Chen, Mengjie, Gao, Yingna, Song, Xianmin, Zheng, Hongliang, Zhang, Kaiyong, Zhang, Bimeng, Chen, Donghui
Format: Article
Language:English
Subjects:
Actin
Calcium (intracellular)
Calcium influx
Calcium signalling
Cell adhesion & migration
Cell migration
Cofilin
Cytoskeleton
cytoskeleton reorganization
Elongation
Filaments
migration
Muscles
Musculoskeletal system
Myoblasts
P2Y6
Rho-associated kinase
RhoA protein
RhoA/ROCK
Signal transduction
Skeletal muscle
Tubulin
UDP
Wound healing
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Summary:Migration of skeletal muscle precursor cells is required for limb muscle development and skeletal muscle repair. This study aimed to examine the role of P2Y6 receptor in C2C12 myoblasts migration. C2C12 myoblasts were treated with P2Y6 agonist UDP, P2Y6 antagonist MRS2578, Ca2+ channel blocker BTP2, or ROCK inhibitor GSK269962 or Y27632, and the migration ability of C2C12 cells was assessed by wound healing assay. The cellular Ca2+ content was analyzed with fluo‐4 probe and the activation of ROCK (phosphorlyation of LIMK and cofilin) was assayed by western blot. The cytoskeleton was labeled with Actin‐Tracker Green and Tubulin‐Tracker‐Red. Silencing P2Y6 expression in C2C12 myoblasts reduced intracellular Ca2+ content and cell motility. Whereas UDP increased cellular Ca2+ content, actin filaments, and cell migration, MRS2578 had the opposite effects. The effects of UDP were abrogated by BTP2 and GSK269962 (and Y27632). Disruption of P2Y6 signaling pathway caused C2C12 myoblasts to have an elongated morphology. These results demonstrated that P2Y6 signaled through Ca2+ influx and RhoA/ROCK to reorganize cytoskeleton and promote migration in myoblasts. P2Y6 promotes cytoskeleton reorganization and migration while inhibits differentiation of C2C12 myoblasts through a CA2+ mediated activation of RhoA/ROCK pathway.
ISSN:0730-2312
1097-4644
DOI:10.1002/jcb.26350