%0 Journal Article
%T Increased Cell-Matrix Adhesion upon Constitutive Activation of Rho Proteins by Cytotoxic Necrotizing Factors from E. coli and Y. pseudotuberculosis
%A Martin May
%A Tanja Kolbe
%A Tianbang Wang
%A Gudula Schmidt
%A Harald Genth
%J Journal of Signal Transduction
%D 2012
%I Hindawi Publishing Corporation
%R 10.1155/2012/570183
%X Cytotoxic necrotizing factors (CNFs) encompass a class of autotransporter toxins produced by uropathogenic E. coli (CNF1) or Y. pseudotuberculosis (CNFy). CNF toxins deamidate and thereby constitutively activate RhoA, Rac1, and Cdc42. In this study, the effects of CNF1 on cell-matrix adhesion are analysed using functional cell-adhesion assays. CNF1 strongly increased cell-matrix binding of suspended Hela cells and decreased the susceptibly of cells to trypsin-induced cell detachment. Increased cell-matrix binding was also observed upon treatment of Hela cells with isomeric CNFy, that specifically deamidates RhoA. Increased cell-matrix binding thus appears to depend on RhoA deamidation. In contrast, increased cell spreading was specifically observed upon CNF1 treatment, suggesting that it rather depended on Rac1/Cdc42 deamidation. Increased cell-matrix adhesion is further presented to result in reduced cell migration of adherent cells. In contrast, migration of suspended cells was not affected upon treatment with CNF1 or CNFy. CNF1 and CNFy thus reduced cell migration specifically under the condition of pre-established cell-matrix adhesion. 1. Introduction Cell-matrix adhesion involves several processes including integrin binding, cell spreading, and flattening against the substrate. Cultured cells, that spread out on ligand coated surfaces, rearrange their cytoskeleton and begin to move. Integrins thereby cluster together in ¡°focal complexes¡± at the leading edge. These focal complexes grow into mature focal contacts, also called focal adhesions (FAs) [1]. Focal adhesions contain over 100 different proteins, including integrins, adapter proteins, and intracellular signaling proteins. Clustered integrins anchor actin filaments to the cell membrane and link them with the extracellular matrix (ECM) through adapter proteins such as talin and vinculin. The adapter protein paxillin links integrins to signaling proteins, forming a scaffold for Src kinases, the focal adhesion kinase (FAK), or the p21-activated kinase (PAK) [2¨C5]. The turnover of FAs in moving cells is driven by small GTPases of the Rho subfamily. FA formation and disassembly at the leading edge is driven by Rac1 and the localized suppression of Rho activity. Disassembly of FAs at the cell rear requires RhoA activity [6]. The activity of Rho proteins is regulated by the GTPase cycle. Rho proteins are active in the GTP-bound state and inactive in the GDP-bound state. In their active conformation Rho proteins interact with effector proteins to transmit downstream signaling. The cycling between
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