title: Suppression of chemotaxis by SSeCKS via scaffolding of phosphoinositol phosphates and the recruitment of the Cdc42 GEF, Frabin, to the leading edge.
abstract: Chemotaxis is controlled by interactions between receptors, Rho-family GTPases, phosphatidylinositol 3-kinases, and cytoskeleton remodeling proteins. We investigated how the metastasis suppressor, SSeCKS, attenuates chemotaxis. Chemotaxis activity inversely correlated with SSeCKS levels in mouse embryo fibroblasts (MEF), DU145 and MDA-MB-231 cancer cells. SSeCKS loss induced chemotactic velocity and linear directionality, correlating with replacement of leading edge lamellipodia with fascin-enriched filopodia-like extensions, the formation of thickened longitudinal F-actin stress fibers reaching to filopodial tips, relative enrichments at the leading edge of phosphatidylinositol (3,4,5)P3 (PIP3), Akt, PKC-ζ, Cdc42-GTP and active Src (SrcpoY416), and a loss of Rac1. Leading edge lamellipodia and chemotaxis inhibition in SSeCKS-null MEF could be restored by full-length SSeCKS or SSeCKS deleted of its Src-binding domain (ΔSrc), but not by SSeCKS deleted of its three MARCKS (myristylated alanine-rich C kinase substrate) polybasic domains (ΔPBD), which bind PIP2 and PIP3. The enrichment of activated Cdc42 in SSeCKS-null leading edge filopodia correlated with recruitment of the Cdc42-specific guanine nucleotide exchange factor, Frabin, likely recruited via multiple PIP2/3-binding domains. Frabin knockdown in SSeCKS-null MEF restores leading edge lamellipodia and chemotaxis inhibition. However, SSeCKS failed to co-immunoprecipitate with Rac1, Cdc42 or Frabin. Consistent with the notion that chemotaxis is controlled by SSeCKS-PIP (vs. -Src) scaffolding activity, constitutively-active phosphatidylinositol 3-kinase could override the ability of the Src inhibitor, SKI-606, to suppress chemotaxis and filopodial enrichment of Frabin in SSeCKS-null MEF. Our data suggest a role for SSeCKS in controlling Rac1 vs. Cdc42-induced cellular dynamics at the leading chemotactic edge through the scaffolding of phospholipids and signal mediators, and through the reorganization of the actin cytoskeleton controlling directional movement.
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Hyun-Kyung
authors.initials: HK
authors.lastname: Ko
authors.affiliation: Div. of Genetic & Reproductive Toxicology, National Center for Toxicological Research, Jefferson, Arkansas, United States of America.
authors.forename: Li-wu
authors.initials: LW
authors.lastname: Guo
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Bing
authors.initials: B
authors.lastname: Su
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Lingqiu
authors.initials: L
authors.lastname: Gao
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Irwin H
authors.initials: IH
authors.lastname: Gelman
mesh_terms: D054758:A Kinase Anchor Proteins / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism*; D045744:Cell Line, Tumor; D002633:Chemotaxis; D004622:Embryo, Mammalian / Q000166:cytology; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008840:Microfilament Proteins / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D010641:Phenotype; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011554:Pseudopodia / Q000378:metabolism*; D015398:Signal Transduction; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism*; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: D054758:A Kinase Anchor Proteins; C491162:AKAP12 protein, human; C491161:Akap12 protein, mouse; D018797:Cell Cycle Proteins; C485165:Fgd4 protein, mouse; D020662:Guanine Nucleotide Exchange Factors; D008840:Microfilament Proteins; D018129:Phosphatidylinositol Phosphates; D019061:src-Family Kinases; D020764:cdc42 GTP-Binding Protein