M-Ras/R-Ras3, a transforming ras protein regulated by Sos1, GRF1, and p120 Ras GTPase-activating protein, interacts with the putative Ras effector AF6.
M-Ras is a Ras-related protein that shares approximately 55% identity with K-Ras and TC21. The M-Ras message was widely expressed but was most predominant in ovary and brain. Similarly to Ha-Ras, expression of mutationally activated M-Ras in NIH 3T3 mouse fibroblasts or C2 myoblasts resulted in cellular transformation or inhibition of differentiation, respectively. M-Ras only weakly activated extracellular signal-regulated kinase 2 (ERK2), but it cooperated with Raf, Rac, and Rho to induce transforming foci in NIH 3T3 cells, suggesting that M-Ras signaled via alternate pathways to these effectors. Although the mitogen-activated protein kinase/ERK kinase inhibitor, PD98059, blocked M-Ras-induced transformation, M-Ras was more effective than an activated mitogen-activated protein kinase/ERK kinase mutant at inducing focus formation. These data indicate that multiple pathways must contribute to M-Ras-induced transformation. M-Ras interacted poorly in a yeast two-hybrid assay with multiple Ras effectors, including c-Raf-1, A-Raf, B-Raf, phosphoinositol-3 kinase delta, RalGDS, and Rin1. Although M-Ras coimmunoprecipitated with AF6, a putative regulator of cell junction formation, overexpression of AF6 did not contribute to fibroblast transformation, suggesting the possibility of novel effector proteins. The M-Ras GTP/GDP cycle was sensitive to the Ras GEFs, Sos1, and GRF1 and to p120 Ras GAP. Together, these findings suggest that while M-Ras is regulated by similar upstream stimuli to Ha-Ras, novel targets may be responsible for its effects on cellular transformation and differentiation.
Quilliam
L A
LA
Department of Biochemistry and Molecular Biology and Walther Oncology Center, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA. lquillia@iupui.edu
Castro
A F
AF
Rogers-Graham
K S
KS
Martin
C B
CB
Der
C J
CJ
Bi
C
C
D016475:3T3 Cells; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000502:physiology*; D002454:Cell Differentiation; D002471:Cell Transformation, Neoplastic; D004789:Enzyme Activation; D005419:Flavonoids / Q000494:pharmacology; D005656:Fungal Proteins / Q000502:physiology*; D020558:GTP Phosphohydrolases / Q000032:analysis / Q000502:physiology*; D020690:GTPase-Activating Proteins; D016547:Kinesins / Q000502:physiology*; D048728:MAP Kinase Kinase Kinase 1; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1; D020559:Monomeric GTP-Binding Proteins; D009218:Myosins / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D011506:Proteins / Q000502:physiology*; D012097:Repressor Proteins / Q000502:physiology*; D020837:SOS1 Protein; D020703:ras GTPase-Activating Proteins; D018631:ras Proteins; D020666:ras-GRF1
C490657:Afdn protein, mouse; D018797:Cell Cycle Proteins; D005419:Flavonoids; D005656:Fungal Proteins; D020690:GTPase-Activating Proteins; D011506:Proteins; D012097:Repressor Proteins; D020837:SOS1 Protein; D020703:ras GTPase-Activating Proteins; D020666:ras-GRF1; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048728:MAP Kinase Kinase Kinase 1; C482576:MAP3K1 protein, human; C482578:Map3k1 protein, mouse; D020558:GTP Phosphohydrolases; C109418:Mras protein, mouse; D009218:Myosins; D016547:Kinesins; D020559:Monomeric GTP-Binding Proteins; D018631:ras Proteins; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one