The effector loop and prenylation site of R-Ras are involved in the regulation of integrin function.
The closely related small GTP-binding proteins H-Ras and R-Ras have opposing effects on the regulation of integrin cell adhesion receptors. To gain insight into the properties of R-Ras with respect to the regulation of integrin function and interactions with downstream effectors we performed an analysis of R-Ras variants containing mutations in the effector binding domain and C-terminal prenylation site. We found that the activation of the downstream effector PI 3-kinase was sensitive to mutations in the effector binding domain, as was the binding to the effectors, Ral-GDS, Raf-1 and the novel effector Nore1. Furthermore, specific mutations in the effector binding loop and C-terminal prenylation motif impaired the ability of R-Ras to regulate integrin function in CHO cells. However, the ability of the R-Ras effector loop mutants to bind, and activate known effectors did not correlate with their ability to regulate integrin function. Thus, the known R-Ras effectors are not critical for regulating integrin activation, at least in CHO cells. Consequently, these studies provide insight into the structural basis of the interactions between R-Ras and its candidate effectors and suggest the existence of novel mechanisms through which this GTPase could regulate cell adhesion.
Oertli
B
B
Department of Vascular Biology, The Scripps Research Institute, La Jolla, California, CA 92037, USA.
Han
J
J
Marte
B M
BM
Sethi
T
T
Downward
J
J
Ginsberg
M
M
Hughes
P E
PE
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D016466:CHO Cells / Q000378:metabolism; D020134:Catalytic Domain; D002462:Cell Membrane / Q000378:metabolism; D017124:Conserved Sequence; D006224:Cricetinae; D004789:Enzyme Activation; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016023:Integrins / Q000502:physiology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011485:Protein Binding; D017368:Protein Prenylation; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D020726:ral Guanine Nucleotide Exchange Factor / Q000378:metabolism; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
D016023:Integrins; D020726:ral Guanine Nucleotide Exchange Factor; D019869:Phosphatidylinositol 3-Kinases; D019908:Proto-Oncogene Proteins c-raf; D020558:GTP Phosphohydrolases; D018631:ras Proteins