Stress-activated Protein Kinase-2/p38 and a Rapamycin-sensitive Pathway Are Required for C2C12 Myogenesis

Ana Cuenda,Philip Cohen

doi:10.1074/jbc.274.7.4341

Abstract

The differentiation of C2C12 myoblasts to myotubes was found to be accompanied by a strong activation of p70 S6 kinase and the mitogen-activated protein kinase (MAPK) family member SAPK2/p38, without significant activation of p42 MAPK and only slight activation of SAPK1/JNK and protein kinase Balpha. Consistent with these findings, SB 203580 (a specific inhibitor of SAPK2/p38) or rapamycin (which blocks the activation of p70 S6 kinase) prevented the formation of multinucleated myotubes, as well as the expression of muscle-specific proteins that included SAPK3 (another MAPK family member). PD 098059 (which prevents the activation of p42 MAPK) had no effect on myotube formation. Surprisingly, the slow activation of p70 S6 kinase during differentiation was not only prevented by rapamycin but also by SB 203580, and the activation of MAPKAP kinase-2 (an in vivo substrate of SAPK2/p38) was not only prevented by SB 203580 but also by rapamycin. In contrast, the acute activation of p70 S6 kinase in C2C12 myoblasts induced by phorbol esters was unaffected by SB 203580 and the acute activation of MAPKAP kinase-2 induced by anisomycin was unaffected by rapamycin. These results show for the first time that SAPK2/p38 plays an essential role in C2C12 cell differentiation.

Highlights

The differentiation of skeletal muscle starts when myoblasts withdraw from the cell division cycle and is characterized morphologically by the alignment, elongation, and fusion of mononucleated myoblasts into multinucleated myotubes
We demonstrate that the SAPK2/p38 pathway is activated during C2C12 cell differentiation and that SB 203580 blocks myotube formation and the expresnase; MKP1, mitogen-activated protein kinase (MAPK) phosphatase-1; SAPK, stress-activated protein kinase; JNK, c-Jun N-terminal kinase; MBP, myelin basic protein, IGF-1, insulin-like growth factor-1; GST, glutathione S-transferase; PMA, phorbol 12-myristate 13-acetate; MAPKAP-K2, MAPK activated protein kinase-2; FCS, fetal calf serum; PKB, protein kinase B; PI 3-kinase, phosphatidylinositide 3-kinase
The activities were measured of five protein kinases that lie in distinct signal transduction pathways, namely MAPKAP-K2, p70 S6 kinase, p42 MAPK, SAPK1/JNK, and PKB␣

Summary

Introduction

The differentiation of skeletal muscle starts when myoblasts withdraw from the cell division cycle and is characterized morphologically by the alignment, elongation, and fusion of mononucleated myoblasts into multinucleated myotubes. Cell division is prevented during muscle differentiation by the induction of cyclin-dependent protein kinase inhibitors (3, 4).

Results

Conclusion