Transforming growth factor beta1 is up-regulated by activated Raf in skeletal myoblasts but does not contribute to the differentiation-defective phenotype.
The Raf/MEK/MAPK signaling module elicits a strong negative impact on skeletal myogenesis that is reflected by a complete loss of muscle gene transcription and differentiation in multinucleated myocytes. Recent evidence indicates that Raf signaling also may contribute to myoblast cell cycle exit and cytoprotection. To further define the mechanisms by which Raf participates in cellular responses, a stable line of myoblasts expressing an estrogen receptor-Raf chimeric protein was created. The cells (23A2RafER(DD)) demonstrate a strict concentration-dependent increase in chimeric Raf protein synthesis and downstream phosphoMAPK activation. Initiation of low-level Raf activity in these cells augments contractile protein expression and myocyte fusion. By contrast, induction of high level Raf activity in 23A2RafER(DD) myoblasts inhibits the formation of myocytes and muscle reporter gene expression. Interestingly, treatment of myoblasts with conditioned medium isolated from Raf-repressive cells inhibits all of the aspects of myogenesis. Closer examination indicates that the transforming growth factor-beta(1) (TGF-beta(1)) gene is up-regulated in Raf-repressive myoblasts. The cells also direct elevated levels of Smad transcriptional activity, suggesting the existence of a TGF-beta(1) autocrine loop. However, extinguishing the biological activity of TGF-beta(1) does not restore the myogenic program. Our results provide evidence for the involvement of Raf signal transmission during myocyte formation as well as during inhibition of myogenesis.
Wang
Xu
X
Department of Poultry Science, The Pennsylvania State University, University Park, Pennsylvania 16802, USA.
Thomson
Season R
SR
Starkey
Jessica D
JD
Page
Jeanine L
JL
Ealy
Alan D
AD
Johnson
Sally E
SE
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D004789:Enzyme Activation; D051379:Mice; D018482:Muscle, Skeletal / Q000166:cytology / Q000378:metabolism; D032448:Myoblasts, Skeletal / Q000166:cytology / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D015398:Signal Transduction; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D053773:Transforming Growth Factor beta1; D015854:Up-Regulation
C508961:Tgfb1 protein, mouse; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D019908:Proto-Oncogene Proteins c-raf