title: Role of Src kinases in mobilization of glycosylphosphatidylinositol-anchored decay-accelerating factor by Dr fimbria-positive adhering bacteria.
abstract: Afa/Dr fimbriae constitute the major virulence factor of diffusely adhering Escherichia coli (Afa/Dr DAEC). After recognizing membrane-bound signaling receptors, they trigger cell responses. One of these receptors is the human decay-accelerating factor (hDAF). It has previously been reported that the binding of Afa/Dr fimbriae to hDAF quickly induces recruitment of hDAF around adhering bacteria. The aim of our study is to analyze the role of Src kinases in the Dr fimbria-induced recruitment of hDAF. Using biochemical methods and confocal microscopy followed by 3-dimensional (3D) analysis, we have shown that the activation and cell membrane targeting of Src kinases are necessary for the recruitment and organization of hDAF around adhering bacteria. We identified c-Src to be the specific kinase involved in this process. Using a set of Src-green fluorescent protein mutants, we showed that the catalytic activity and the Src homology 2 (SH2) and SH3 domains of the Src kinases are necessary for Dr fimbria-induced hDAF mobilization to occur. In addition, using mutated Dr fimbriae and a set of mutated hDAFs in which each of the complement control protein (CCP) domains had successively been deleted, we found that the aspartic acids at position 54 in the Dr fimbriae and in CCP domain 4 of hDAF played pivotal roles in the mobilization of the Src kinases and hDAF, respectively.
authors.affiliation: INSERM UMR 984, Réponses Cellulaires aux Microorganismes Pathogènes, 92296 Châtenay-Malabry, France.
authors.forename: Christophe J
authors.initials: CJ
authors.lastname: Queval
authors.forename: Valérie
authors.initials: V
authors.lastname: Nicolas
authors.forename: Isabelle
authors.initials: I
authors.lastname: Beau
mesh_terms: D018830:Adhesins, Escherichia coli / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001422:Bacterial Adhesion; D018958:CD55 Antigens / Q000737:chemistry / Q000378:metabolism*; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D004926:Escherichia coli / Q000472:pathogenicity* / Q000502:physiology; D029968:Escherichia coli Proteins / Q000378:metabolism; D036781:Fimbriae Proteins / Q000378:metabolism*; D010861:Fimbriae, Bacterial / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D021962:Membrane Microdomains; D034741:RNA, Small Interfering; D015398:Signal Transduction; D018909:src Homology Domains; D019061:src-Family Kinases / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018830:Adhesins, Escherichia coli; D015703:Antigens, CD; D018958:CD55 Antigens; C069847:CD66 antigens; D015815:Cell Adhesion Molecules; C465564:Dr adhesin, E coli; D029968:Escherichia coli Proteins; D017261:Glycosylphosphatidylinositols; D034741:RNA, Small Interfering; D049452:Green Fluorescent Proteins; D036781:Fimbriae Proteins; D019061:src-Family Kinases