title: Src Family Tyrosine Kinase Signaling Regulates FilGAP through Association with RBM10.
abstract: FilGAP is a Rac-specific GTPase-activating protein (GAP) that suppresses lamellae formation. In this study, we have identified RBM10 (RNA Binding Motif domain protein 10) as a FilGAP-interacting protein. Although RBM10 is mostly localized in the nuclei in human melanoma A7 cells, forced expression of Src family tyrosine kinase Fyn induced translocation of RBM10 from nucleus into cell peripheries where RBM10 and FilGAP are co-localized. The translocation of RBM10 from nucleus appears to require catalytic activity of Fyn since kinase-negative Fyn mutant failed to induce translocation of RBM10 in A7 cells. When human breast carcinoma MDA-MB-231 cells are spreading on collagen-coated coverslips, endogenous FilGAP and RBM10 were localized at the cell periphery with tyrosine-phosphorylated proteins. RBM10 appears to be responsible for targeting FilGAP at the cell periphery because depletion of RBM10 by siRNA abrogated peripheral localization of FilGAP during cell spreading. Association of RBM10 with FilGAP may stimulate RacGAP activity of FilGAP. First, forced expression of RBM10 suppressed FilGAP-mediated cell spreading on collagen. Conversely, depletion of endogenous RBM10 by siRNA abolished FilGAP-mediated suppression of cell spreading on collagen. Second, FilGAP suppressed formation of membrane ruffles induced by Fyn and instead produced spiky cell protrusions at the cell periphery. This protrusive structure was also induced by depletion of Rac, suggesting that the formation of protrusions may be due to suppression of Rac by FilGAP. We found that depletion of RBM10 markedly reduced the formation of protrusions in cells transfected with Fyn and FilGAP. Finally, depletion of RBM10 blocked FilGAP-mediated suppression of ruffle formation induced by EGF. Taken together, these results suggest that Src family tyrosine kinase signaling may regulate FilGAP through association with RBM10.
authors.affiliation: Division of Cell Biology, Department of Biosciences, School of Science, Kitasato University, Kanagawa, 252-0373, Japan.
authors.forename: Hazuki
authors.initials: H
authors.lastname: Yamada
authors.affiliation: Division of Cell Biology, Department of Biosciences, School of Science, Kitasato University, Kanagawa, 252-0373, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Tsutsumi
authors.affiliation: Division of Cell Biology, Department of Biosciences, School of Science, Kitasato University, Kanagawa, 252-0373, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Nakazawa
authors.affiliation: Division of Biochemistry, School of Pharmaceutical Sciences, Kitasato University, Tokyo, 108-8641, Japan.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Shibagaki
authors.affiliation: Division of Biochemistry, School of Pharmaceutical Sciences, Kitasato University, Tokyo, 108-8641, Japan.
authors.forename: Seisuke
authors.initials: S
authors.lastname: Hattori
authors.affiliation: Division of Cell Biology, Department of Biosciences, School of Science, Kitasato University, Kanagawa, 252-0373, Japan.
authors.forename: Yasutaka
authors.initials: Y
authors.lastname: Ohta
mesh_terms: D001943:Breast Neoplasms / Q000378:metabolism*; D002384:Catalysis; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D003094:Collagen / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D005260:Female; D020690:GTPase-Activating Proteins / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D057809:HEK293 Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism*; D015398:Signal Transduction; D014443:Tyrosine / Q000737:chemistry; D019061:src-Family Kinases / Q000378:metabolism*
chemical_list: C497033:ARHGAP24 protein, human; D018076:DNA, Complementary; D020690:GTPase-Activating Proteins; C512260:RBM10 protein, human; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D014443:Tyrosine; D003094:Collagen; D019061:src-Family Kinases