Sphingosine 1-phosphate induces myoblast differentiation through Cx43 protein expression: a role for a gap junction-dependent and -independent function.
Although sphingosine 1-phosphate (S1P) has been considered a potent regulator of skeletal muscle biology, acting as a physiological anti-mitogenic and prodifferentiating agent, its downstream effectors are poorly known. In the present study, we provide experimental evidence for a novel mechanism by which S1P regulates skeletal muscle differentiation through the regulation of gap junctional protein connexin (Cx) 43. Indeed, the treatment with S1P greatly enhanced Cx43 expression and gap junctional intercellular communication during the early phases of myoblast differentiation, whereas the down-regulation of Cx43 by transfection with short interfering RNA blocked myogenesis elicited by S1P. Moreover, calcium and p38 MAPK-dependent pathways were required for S1P-induced increase in Cx43 expression. Interestingly, enforced expression of mutated Cx43(Delta130-136) reduced gap junction communication and totally inhibited S1P-induced expression of the myogenic markers, myogenin, myosin heavy chain, caveolin-3, and myotube formation. Notably, in S1P-stimulated myoblasts, endogenous or wild-type Cx43 protein, but not the mutated form, coimmunoprecipitated and colocalized with F-actin and cortactin in a p38 MAPK-dependent manner. These data, together with the known role of actin remodeling in cell differentiation, strongly support the important contribution of gap junctional communication, Cx43 expression and Cx43/cytoskeleton interaction in skeletal myogenesis elicited by S1P.
Squecco
R
R
Departments of Biochemical Sciences, University of Florence, Interuniversity Institute of Myology, Florence I-50134, Italy.
Sassoli
C
C
Nuti
F
F
Martinesi
M
M
Chellini
F
F
Nosi
D
D
Zecchi-Orlandini
S
S
Francini
F
F
Formigli
L
L
Meacci
E
E
D000818:Animals; D015415:Biomarkers; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D018031:Connexin 43 / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects; D004553:Electric Conductivity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017629:Gap Junctions / Q000187:drug effects*; D015870:Gene Expression / Q000187:drug effects; D007700:Kinetics; D008246:Lysophospholipids / Q000494:pharmacology*; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D050505:Mutant Proteins / Q000378:metabolism; D032448:Myoblasts, Skeletal / Q000166:cytology / Q000187:drug effects*; D018008:Myogenin / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors
D015415:Biomarkers; D018031:Connexin 43; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D008246:Lysophospholipids; D050505:Mutant Proteins; D018008:Myogenin; C060506:sphingosine 1-phosphate; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases; D013110:Sphingosine; D002118:Calcium