R-Ras C-terminal sequences are sufficient to confer R-Ras specificity to H-Ras.
Activated versions of the similar GTPases, H-Ras and R-Ras, have differing effects on biological phenotypes: Activated H-Ras strongly transforms many fibroblast cell lines causing dramatic changes in cell shape and cytoskeletal organization. In contrast, R-Ras transforms fewer cell lines and the transformed cells display only some of the morphological changes associated with H-Ras transformation. H-Ras cells can survive in the absence of serum whereas R-Ras cells seem to die by an apoptotic-like mechanism in response to removal of serum. H-Ras can suppress integrin activation and R-Ras specifically antagonizes this effect. To map sequences responsible for these differences we have generated and investigated a panel of H-Ras and R-Ras chimeras. We found that the C-terminal 53 amino acids of R-Ras were necessary and sufficient to specify the contrasting biological properties of R-Ras with respect to focus morphology, reactive oxygen species (ROS) production and reversal of H-Ras-induced integrin suppression. Surprisingly, we found chimeras in which the focus formation and integrin-mediated phenotypes were separated, suggesting that different effectors could be involved in mediating these responses. An integrin profile of H-Ras and R-Ras cell pools showed no significant differences; both activated H-Ras and R-Ras expressing cells were found to have reduced beta(1) activity, suggesting that the activity state of the beta(1) subunit is not sufficient to direct an H-Ras transformed cell morphology.
Hansen
Malene
M
Department of Molecular Cell Biology, Institute of Molecular Biology, University of Copenhagen, Øster Farimagsgade 2A, Denmark.
Rusyn
Elena V
EV
Hughes
Paul E
PE
Ginsberg
Mark H
MH
Cox
Adrienne D
AD
Willumsen
Berthe M
BM
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005353:Fibronectins / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000502:physiology*; D017353:Gene Deletion; D011905:Genes, ras / Q000502:physiology*; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008969:Molecular Sequence Data; D010957:Plasmids; D017382:Reactive Oxygen Species; D011993:Recombinant Fusion Proteins / Q000235:genetics; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014162:Transfection; D018631:ras Proteins / Q000502:physiology*
D005353:Fibronectins; D017382:Reactive Oxygen Species; D011993:Recombinant Fusion Proteins; D008156:Luciferases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D020558:GTP Phosphohydrolases; C513254:Rras protein, mouse; D018631:ras Proteins