TY - JOUR

T1 - The differentiation of skeletal muscle cells involves a protein-tyrosine phosphatase-alpha-mediated C-Src signaling pathway.

AU - Lu, Huogen

AU - Shah, Poonam

AU - Ennis, David

AU - Shinder, Gail

AU - Sap, Jan

AU - Le-Tien, Hoang

AU - Fantus, I George

N1 - Keywords: Animals; Blotting, Western; Cell Differentiation; Cell Division; Cell Line; Fluorescent Antibody Technique; Muscle, Skeletal; Protein Tyrosine Phosphatases; Proto-Oncogene Proteins pp60(c-src); Rats; Signal Transduction

PY - 2002

Y1 - 2002

N2 - Protein-tyrosine phosphatase-alpha (PTPalpha) plays an important role in various cellular signaling events, including proliferation and differentiation. In this study, we established L6 cell lines either underexpressing or overexpressing PTPalpha by stable transfection of cells with antisense PTPalpha or with full-length wild-type human or mouse or double catalytic site Cys --> Ala mutant (DM8) PTPalpha cDNA. Expression of PTPalpha in these cell lines was determined by immunoblotting and immunofluorescence. Cells harboring antisense PTPalpha exhibited a significantly reduced growth rate and thymidine incorporation when compared with the wild-type L6 cells. In contrast, cells overexpressing PTPalpha showed more rapid (2-fold) proliferation. Myoblasts with diminished PTPalpha failed to undergo fusion and did not form myotubes in reduced serum whereas overexpression of PTPalpha promoted myogenesis 2 days earlier than wild-type L6 cells. Overexpression of phosphatase-inactive mutant PTPalpha recapitulated the phenotype of the antisense cells. The different myogenic activities of these cell lines were correlated with the expression of myogenin and creatine kinase activity. Consistent with previous reports, PTPalpha positively regulated the activity of the protein-tyrosine kinase Src. Treatment of L6 cells with PP2 or SU6656, specific inhibitors of Src family kinases, and transient transfection of dominant-inhibitory Src inhibited the formation of myotubes and expression of myogenin. Moreover, enhanced expression of PTPalpha and activation of Src was detected during myogenesis. Together, these data indicate that PTPalpha is involved in the regulation of L6 myoblast growth and skeletal muscle cell differentiation via an Src-mediated signaling pathway.

AB - Protein-tyrosine phosphatase-alpha (PTPalpha) plays an important role in various cellular signaling events, including proliferation and differentiation. In this study, we established L6 cell lines either underexpressing or overexpressing PTPalpha by stable transfection of cells with antisense PTPalpha or with full-length wild-type human or mouse or double catalytic site Cys --> Ala mutant (DM8) PTPalpha cDNA. Expression of PTPalpha in these cell lines was determined by immunoblotting and immunofluorescence. Cells harboring antisense PTPalpha exhibited a significantly reduced growth rate and thymidine incorporation when compared with the wild-type L6 cells. In contrast, cells overexpressing PTPalpha showed more rapid (2-fold) proliferation. Myoblasts with diminished PTPalpha failed to undergo fusion and did not form myotubes in reduced serum whereas overexpression of PTPalpha promoted myogenesis 2 days earlier than wild-type L6 cells. Overexpression of phosphatase-inactive mutant PTPalpha recapitulated the phenotype of the antisense cells. The different myogenic activities of these cell lines were correlated with the expression of myogenin and creatine kinase activity. Consistent with previous reports, PTPalpha positively regulated the activity of the protein-tyrosine kinase Src. Treatment of L6 cells with PP2 or SU6656, specific inhibitors of Src family kinases, and transient transfection of dominant-inhibitory Src inhibited the formation of myotubes and expression of myogenin. Moreover, enhanced expression of PTPalpha and activation of Src was detected during myogenesis. Together, these data indicate that PTPalpha is involved in the regulation of L6 myoblast growth and skeletal muscle cell differentiation via an Src-mediated signaling pathway.

U2 - 10.1074/jbc.M209643200

DO - 10.1074/jbc.M209643200

M3 - Journal article

C2 - 12351660

VL - 277

SP - 46687

EP - 46695

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 48

ER -