Active Ras-induced effects on skeletal myoblast differentiation and apoptosis are independent of constitutive PI3-kinase activity.
23A2 myoblasts expressing GAP-resistant, constitutively active G12V:H-Ras (A2:G12V:H-Ras myoblasts) display a transformed morphology and do not undergo mitogen-deprivation-induced differentiation or the associated apoptosis. To determine the phenotype induced by F156L:H-Ras, a constitutively active mutant with enhanced nucleotide exchange activity rather than impaired GAP-stimulated GTPase activity, myoblast cell lines were established that stably express F156L:H-Ras at levels of H-Ras comparable to the A2:G12V:H-Ras myoblasts. These A2:F156L:H-Ras myoblast cell lines do not possess a transformed morphology, and while differentiation and apoptosis are impaired, these processes are not abrogated as in the A2:G12V:H-Ras myoblasts. Surprisingly, while expression of either G12V:H-Ras or F156L:H-Ras results in constitutive signaling through PI3-kinase, only cells expressing G12V:H-Ras additionally possess constitutive signaling through MAPK, and NFkappaB. Pharmacological abrogation of the Ras-induced constitutive PI3-kinase signal, however, is not responsible for the impaired differentiation or apoptosis in either A2:G12V:H-Ras myoblasts or A2:F156L:H-Ras myoblasts. Thus, our data suggest that a pathway distinct from those that signals through MAPK, NFkappaB or PI3-kinase is responsible for the impaired differentiation and apoptosis in 23A2 skeletal myoblasts expressing constitutively active Ras.
Karasarides
M
M
Department of Cell Biology, Learner Research Institute, Cleveland Clinic Foundation, OH 44195, USA.
Dee
K
K
Schulman
D
D
Wolfman
A
A
Weyman
C M
CM
D000818:Animals; D017209:Apoptosis; D002454:Cell Differentiation; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D015870:Gene Expression; D020930:MAP Kinase Kinase Kinases / Q000037:antagonists & inhibitors; D051379:Mice; D017570:MyoD Protein / Q000235:genetics; D032448:Myoblasts, Skeletal / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism*; D016328:NF-kappa B / Q000037:antagonists & inhibitors; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000037:antagonists & inhibitors; D015398:Signal Transduction; D018631:ras Proteins / Q000235:genetics* / Q000378:metabolism*
D020930:MAP Kinase Kinase Kinases; D017570:MyoD Protein; D016328:NF-kappa B; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D018631:ras Proteins