SWAP-70 is a guanine-nucleotide-exchange factor that mediates signalling of membrane ruffling

Shirakawa, Masahiro; Terada, Yoh; Iwamatsu, Akihiro; Shinohara, Azusa; Mochizuki, Naoki; Higuchi, Masakazu; Gotoh, Yukiko; Ihara, Sayoko; Nagata, Satoshi; Itoh, Hiroshi; Fukui, Yoshihiro; Jessberger, Rolf

doi:10.1038/416759a

Cited by 207 publications

(258 citation statements)

References 31 publications

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“…As phosphatidylinositol 3-kinase has been reported to regulate Rac1 activity through the several guanine nucleotide exchange factors, such as SWAP70 and P-REX2 downstream of Gab1 (53)(54)(55), and Vav and Tiam-1 are also known to mediate Rac1 activity involved in transformation (56, 57), we initially speculated that the implication of Crk in HGF-mediated Rac1 activation may be partial. However, the significant suppression of HGF-dependent activation of Rac1, cell motility, and scattering was shown in Crk knockdown cells, which may suggest the crucial role for Crk in HGF-evoked activation of Rac1 and following cell motility in human synovial sarcoma cells.…”

Section: Discussionmentioning

confidence: 99%

Adaptor Molecule Crk Is Required for Sustained Phosphorylation of Grb2-Associated Binder 1 and Hepatocyte Growth Factor–Induced Cell Motility of Human Synovial Sarcoma Cell Lines

Watanabe

Tsuda

Makino

et al. 2006

Molecular Cancer Research

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Activation of the c-Met receptor tyrosine kinase through its ligand, hepatocyte growth factor (HGF), promotes mitogenic, motogenic, and morphogenic cellular responses. Aberrant HGF/c-Met signaling has been strongly implicated in tumor cell invasion and metastasis. Both HGF and its receptor c-Met have been shown to be overexpressed in human synovial sarcoma, which often metastasizes to the lung; however, little is known about HGF-mediated biological effects in this sarcoma. Here, we provide evidence that Crk adaptor protein is required for the sustained phosphorylation of c-Met-docking protein Grb2-associated binder 1 (Gab1) in response to HGF, leading to the enhanced cell motility of human synovial sarcoma cell lines SYO-1, HS-SY-II, and Fuji. HGF stimulation induced the sustained phosphorylation on Y307 of Gab1 where Crk was recruited. Crk knockdown by RNA interference disturbed this HGF-induced tyrosine phosphorylation of Gab1. By mutational analysis, we identified that Src homology 2 domain of Crk is indispensable for the induction of the phosphorylation on multiple Tyr-X-X-Pro motifs containing Y307 in Gab1. HGF remarkably stimulated cell motility and scattering of synovial sarcoma cell lines, consistent with the prominent activation of Rac1, extreme filopodia formation, and membrane ruffling.

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Section: Discussionmentioning

confidence: 99%

Adaptor Molecule Crk Is Required for Sustained Phosphorylation of Grb2-Associated Binder 1 and Hepatocyte Growth Factor–Induced Cell Motility of Human Synovial Sarcoma Cell Lines

Watanabe

Tsuda

Makino

et al. 2006

Molecular Cancer Research

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“…It has been shown that several GEFs activate Rac1 in response to EGF and PDGF signaling [13,[23][24][25]. To determine which exchange factors activate Rac1 downstream of VEGF, we first investigated the endogenous expression of candidate Rac1 GEFs in endothelial cells.…”

Section: Exchange Factor Vav2 Couples Vegf Signaling To Rac1mentioning

confidence: 99%

VEGF-induced Rac1 activation in endothelial cells is regulated by the guanine nucleotide exchange factor Vav2

Garrett

Buul

Burridge

2007

Experimental Cell Research

152

132

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Vascular endothelial growth factor (VEGF) signaling is critical for both normal and diseaseassociated vascular development. Dysregulated VEGF signaling has been implicated in ischemic stroke, tumor angiogenesis, and many other vascular diseases. VEGF signals through several effectors, including the Rho family of small GTPases. As a member of this family, Rac1 promotes VEGF-induced endothelial cell migration by stimulating the formation of lamellipodia and membrane ruffles. To form these membrane protrusions, Rac1 is activated by guanine nucleotide exchange factors (GEFs) that catalyze the exchange of GDP for GTP. The goal of this study was to identify the GEF responsible for activating Rac1 in response to VEGF stimulation. We have found that VEGF stimulates biphasic activation of Rac1 and for these studies we focused on the peak of activation that occurs at 30 min. Inhibition of VEGFR-2 signaling blocks VEGF-induced Rac1 activation. Using a Rac1 nucleotide-free mutant (G15ARac1), which has a high affinity for binding activated GEFs, we show that the Rac GEF Vav2 associates with G15ARac1 after VEGF stimulation. Additionally, we show that depleting endothelial cells of endogenous Vav2 with siRNA prevents VEGF-induced Rac1 activation. Moreover, Vav2 is tyrosine phosphorylated upon VEGF treatment, which temporally correlates with Rac1 activation and requires VEGFR-2 signaling and Src kinase activity. Finally, we show that depressing Vav2 expression by siRNA impairs VEGF-induced endothelial cell migration. Taken together, our results provide evidence that Vav2 acts downstream of VEGF to activate Rac1.

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“…The inositol-polyphosphate 5-phosphatases (referred to as 5-phosphatases) are a large family of signal-modifying enzymes that hydrolyze the 5-phosphate from the inositol ring of both inositol phosphates, such as Ins(1,4,5)P 3 and Ins(1,3,4,5)P 4 , and/or PtdIns-derived messenger molecules, including PtdIns(4,5)P 2 , PtdIns(3,4,5)P 3 , and PtdIns(3,5)P 2 (9). Ten mammalian and four yeast homologs have been identified and characterized.…”

Section: Phosphatidylinositolmentioning

confidence: 99%

Identification of a Novel Domain in Two Mammalian Inositol-polyphosphate 5-Phosphatases That Mediates Membrane Ruffle Localization

Gurung

Tan

Ooms

et al. 2003

Journal of Biological Chemistry

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SKIP (skeletal muscle and kidney enriched inositol phosphatase) is a recently identified phosphatidylinositol 3,4,5-trisphosphate-and phosphatidylinositol 4,5-bisphosphate-specific 5-phosphatase. In this study, we investigated the intracellular localization of SKIP. Indirect immunofluorescence and subcellular fractionation showed that, in serum-starved cells, both endogenous and recombinant SKIP colocalized with markers of the endoplasmic reticulum (ER). Following epidermal growth factor (EGF) stimulation, SKIP transiently translocated to plasma membrane ruffles and colocalized with submembranous actin. Data base searching demonstrated a novel 128-amino acid domain in the C terminus of SKIP, designated SKICH for SKIP carboxyl homology, which is also found in the 107-kDa 5-phosphatase PIPP and in members of the TRAF6-binding protein family. Recombinant SKIP lacking the SKICH domain localized to the ER, but did not translocate to membrane ruffles following EGF stimulation. The SKIP SKICH domain showed perinuclear localization and mediated EGF-stimulated plasma membrane ruffle localization. The SKICH domain of the 5-phosphatase PIPP also mediated plasma membrane ruffle localization. Mutational analysis identified the core sequence within the SKICH domain that mediated constitutive membrane association and C-terminal sequences unique to SKIP that contributed to ER localization. Collectively, these studies demonstrate a novel membrane-targeting domain that serves to recruit SKIP and PIPP to membrane ruffles.Phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P 2 ) 1 serves as a precursor to two major signaling pathways. This central phosphoinositide is phosphorylated by phosphoinositide 3-kinase, forming PtdIns(3,4,5)P 3 , which transiently accumulates at the plasma membrane of growth factor-activated cells and is subsequently rapidly metabolized by PtdIns(3,4,5)P 3 5-or 3-phosphate-specific lipid phosphatases. PtdIns(3,4,5)P 3 recruits various effector proteins that contain PH domains, such as the serine/threonine kinases Akt and PDK1 (1), and Rho, Rac, and ADP-ribosylation factor guanine nucleotide exchange factors, including P-Rex1 (2) and SWAP-70 (3), which in turn regulate many signaling pathways, including inhibition of cell death, cell cycle progression, actin polymerization, membrane ruffling, cell migration, and secretion (4 -7). In addition, PtdIns(4,5)P 2 is hydrolyzed by phospholipase C to produce inositol 1,4,5-trisphosphate (Ins(1,4,5)P 3 ) and diacylglycerol, which mobilize intracellular calcium and activate protein kinase C, respectively. PtdIns(4,5)P 2 itself regulates the activity of actin-binding proteins by suppressing the function of vinculin, cofilin, gelsolin, and profilin and by activating the actin-gelatinizing activity of ␣-actinin (8).The inositol-polyphosphate 5-phosphatases (referred to as 5-phosphatases) are a large family of signal-modifying enzymes that hydrolyze the 5-phosphate from the inositol ring of both inositol phosphates, such as Ins(1,4,5)P 3 and Ins(1,3,4,5)P 4 , and/or PtdIns-...

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SWAP-70 is a guanine-nucleotide-exchange factor that mediates signalling of membrane ruffling

Cited by 207 publications

References 31 publications

Adaptor Molecule Crk Is Required for Sustained Phosphorylation of Grb2-Associated Binder 1 and Hepatocyte Growth Factor–Induced Cell Motility of Human Synovial Sarcoma Cell Lines

Adaptor Molecule Crk Is Required for Sustained Phosphorylation of Grb2-Associated Binder 1 and Hepatocyte Growth Factor–Induced Cell Motility of Human Synovial Sarcoma Cell Lines

VEGF-induced Rac1 activation in endothelial cells is regulated by the guanine nucleotide exchange factor Vav2

Identification of a Novel Domain in Two Mammalian Inositol-polyphosphate 5-Phosphatases That Mediates Membrane Ruffle Localization

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