title: RA-GEF-1, a guanine nucleotide exchange factor for Rap1, is activated by translocation induced by association with Rap1*GTP and enhances Rap1-dependent B-Raf activation.
abstract: We previously identified RA-GEF-1, a novel guanine nucleotide exchange factor (GEF) for Rap1 with the ability to associate with Rap1.GTP at its Ras/Rap1-associating (RA) domain. Because it possesses a PSD-95/DlgA/ZO-1 (PDZ) domain, it was also named PDZ-GEF. In this report, we have examined the role of the RA domain of this protein in Rap1-mediated cellular responses. A mutant of RA-GEF-1 (RA-GEF-1DeltaRA) carrying a 21-residue deletion at its RA domain fully retains the in vitro GEF activity toward Rap1 but completely loses the Rap1 binding activity. In contrast, RA-GEF-1DeltaRA, expressed in COS-7 cells, exhibits a 3-fold reduction in its in vivo GEF activity toward Rap1 compared with wild-type RA-GEF-1 as examined by the Rap1 pull-down assay. Correspondingly, when coexpressed with wild-type Rap1, RA-GEF-1DeltaRA is unable to further activate B-Raf, whereas RA-GEF-1 stimulates B-Raf as efficiently as activated Rap1. Consistent with these observations, coexpression of activated Rap1 induces translocation of RA-GEF-1, which is otherwise located in the cytoplasm, to the perinuclear compartment, where Rap1 is also predominantly localized. This localization almost coincides with that of the Golgi apparatus, which was detected by anti-trans-Golgi-network 38 antibody. RA-GEF-1DeltaRA fails to show the translocation. These results indicate that RA-GEF-1 defines a novel category of GEF that is translocated to a particular subcellular compartment by association with the GTP-bound form of a small GTPase and catalyzes activation of the GDP-bound form present in the compartment, thereby causing an amplification of cellular responses induced by the small GTPase.
authors.affiliation: Division of Molecular Biology, Department of Molecular and Cellular Biology, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650-0017, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Liao
authors.forename: T
authors.initials: T
authors.lastname: Satoh
authors.forename: X
authors.initials: X
authors.lastname: Gao
authors.forename: T G
authors.initials: TG
authors.lastname: Jin
authors.forename: C D
authors.initials: CD
authors.lastname: Hu
authors.forename: T
authors.initials: T
authors.lastname: Kataoka
mesh_terms: D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D004789:Enzyme Activation; D017353:Gene Deletion; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics* / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009419:Nerve Tissue Proteins; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D051381:Rats; D013997:Time Factors; D014162:Transfection; D020827:rap1 GTP-Binding Proteins / Q000378:metabolism*; D020663:ras Guanine Nucleotide Exchange Factors / Q000502:physiology*
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D009419:Nerve Tissue Proteins; C403525:RAPGEF2 protein, human; D020663:ras Guanine Nucleotide Exchange Factors; D000242:Cyclic AMP; D019908:Proto-Oncogene Proteins c-raf; D020827:rap1 GTP-Binding Proteins; D006152:Cyclic GMP