title: Regulation of extracellular signal-regulated kinase activity by p120 RasGAP does not involve its pleckstrin homology or calcium-dependent lipid binding domains but does require these domains to regulate cell proliferation.
abstract: The gene encoding for p120 RasGAP, has been disrupted in mice (M. Henkemeyer et al., Nature (Lond.), 377: 695-701, 1995). In this study, using fibroblasts derived from these mouse embryos (Gap-/-; P. van der Geer et al., Mol. Cell Biol., 17: 1840-1847, 1997), we demonstrate that mitogen-activated protein kinase (MAPK) activation is prolonged after epidermal growth factor (EGF), but not lysophosphatidic acid, stimulation as compared with wild-type cells. Furthermore, these cells exhibited a moderate increase in their proliferative rate and saturation density, as well as a limited ability to form colonies in soft agar. Stable cell lines expressing full-length p120GAP not only restored the ability to down-regulate MAPK after EGF stimulation but also lowered their saturation densities. Similarly, expression of p120GAP, missing either its pleckstrin homology (PH) or its calcium-dependent lipid binding (CaLB)/C2 domain, restored MAPK down-regulation and retained the ability to associate with p190 RhoGAP and to be phosphorylated by v-src but exhibited higher saturation densities similar to Gap-/- cells. Our results, therefore, suggest that p120GAP functions not only by down-regulating the Ras/MAPK pathway after growth factor stimulation but is also important in regulating cell proliferation that involves its PH and CaLB domains.
authors.affiliation: Banting and Best Department of Medical Research, Medical Genetics, University of Toronto, Toronto, Ontario M9W 7H4, Canada.
authors.forename: J A
authors.initials: JA
authors.lastname: Koehler
authors.forename: M F
authors.initials: MF
authors.lastname: Moran
mesh_terms: D000818:Animals; D001798:Blood Proteins / Q000737:chemistry*; D002118:Calcium / Q000378:metabolism*; D002452:Cell Count; D002455:Cell Division; D002460:Cell Line; D003260:Contact Inhibition; D004268:DNA-Binding Proteins; D015536:Down-Regulation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology; D020690:GTPase-Activating Proteins; D017353:Gene Deletion; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D050356:Lipid Metabolism; D020935:MAP Kinase Signaling System; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D015688:Oncogene Protein pp60(v-src) / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism; D012097:Repressor Proteins; D020729:p120 GTPase Activating Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020666:ras-GRF1
chemical_list: C074560:ARHGAP35 protein, human; C513043:ARHGAP5 protein, human; C513041:Arhgap35 protein, mouse; C513042:Arhgap5 protein, mouse; D001798:Blood Proteins; D004268:DNA-Binding Proteins; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; D010750:Phosphoproteins; C513039:RASGRF1 protein, human; C513037:Rasgrf1 protein, mouse; D011956:Receptors, Cell Surface; D012097:Repressor Proteins; D020729:p120 GTPase Activating Protein; C039077:platelet protein P47; D020666:ras-GRF1; D004815:Epidermal Growth Factor; D020794:Receptor Protein-Tyrosine Kinases; D015688:Oncogene Protein pp60(v-src); D019908:Proto-Oncogene Proteins c-raf; D020928:Mitogen-Activated Protein Kinases; D020962:Heterotrimeric GTP-Binding Proteins; D002118:Calcium