H-ras and raf-1 cooperate in transformation of NIH3T3 fibroblasts.
We examined the effect of overexpression of growth factor-regulated second messenger enzymes, alone and in combination, on transformation of NIH3T3 cells. Signal transducers included phospholipase C-gamma (PLC-gamma), protein kinase C-gamma (PKC-gamma), and two proto-oncogenes, c-H-ras and c-raf-1. Three of these proteins, PLC-gamma, PKC-gamma and Raf-1, did not transform NIH3T3 cells alone or in combination. c-H-ras, which under its own promoter control has low transforming activity, also did not cooperate with PLC-gamma or PKC-gamma. In contrast, the combination of normal or oncogenic p21 H-Ras with the Raf-1 kinase dramatically increased transformation efficiency. The level of Ras protein required for transformation was reduced in Raf-1 co-transfectants, implying that, at low levels of p21 Ras, p74 Raf-1 is rate limiting. As transformation by Ras depends on jun-mediated transcriptional events, we also examined H-ras and c-raf-1 cooperation in transcriptional transactivation of TPA-responsive element (TRE)-dependent reporters. Like the H-ras/c-raf-1 cooperation in transformation, we observed this synergistic stimulation of TRE-dependent transcription. This pathway for transformation and transcriptional activation by increased levels of normal Ras and Raf may be important in tumors that show overexpression but lack mutationally activated forms of these two proto-oncogenes.
Cuadrado
A
A
Laboratory of Cellular and Molecular Biology, National Cancer Institute, Bethesda, Maryland 20892.
Bruder
J T
JT
Heidaran
M A
MA
App
H
H
Rapp
U R
UR
Aaronson
S A
SA
D016475:3T3 Cells; D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D015870:Gene Expression; D011905:Genes, ras; D006133:Growth Substances / Q000502:physiology; D066298:In Vitro Techniques; D051379:Mice; D011493:Protein Kinase C / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D019908:Proto-Oncogene Proteins c-raf; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D015290:Second Messenger Systems; D014158:Transcription, Genetic; D014162:Transfection; D010738:Type C Phospholipases / Q000502:physiology
D006133:Growth Substances; D011518:Proto-Oncogene Proteins; D011505:Protein-Tyrosine Kinases; D019908:Proto-Oncogene Proteins c-raf; D011493:Protein Kinase C; D010738:Type C Phospholipases; D016283:Proto-Oncogene Proteins p21(ras)