title: Absence of O antigen suppresses Shigella flexneri IcsA autochaperone region mutations.
abstract: The Shigella flexneri IcsA (VirG) protein is a polarly distributed autotransporter protein. IcsA functions as a virulence factor by interacting with the host actin regulatory protein N-WASP, which in turn activates the Arp2/3 complex, initiating actin polymerization. Formation of F-actin comet tails allows bacterial cell-to-cell spreading. Although various accessory proteins such as periplasmic chaperones and the β-barrel assembly machine (BAM) complex have been shown to be involved in the export of IcsA, the IcsA translocation mechanism remains to be fully elucidated. A putative autochaperone (AC) region (amino acids 634-735) located at the C-terminal end of the IcsA passenger domain, which forms part of the self-associating autotransporter (SAAT) domain, has been suggested to be required for IcsA biogenesis, as well as for N-WASP recruitment, based on mutagenesis studies. IcsA(i) proteins with linker insertion mutations within the AC region have a significant reduction in production and are defective in N-WASP recruitment when expressed in smooth LPS (S-LPS) S. flexneri. In this study, we have found that the LPS O antigen plays a role in IcsA(i) production based on the use of an rmlD (rfbD) mutant having rough LPS (R-LPS) and a novel assay in which O antigen is depleted using tunicamycin treatment and then regenerated. In addition, we have identified a new N-WASP binding/interaction site within the IcsA AC region.
authors.affiliation: Discipline of Microbiology and Immunology, School of Molecular and Biomedical Science, University of Adelaide, Adelaide 5005, Australia.
authors.forename: Min Yan
authors.initials: MY
authors.lastname: Teh
authors.affiliation: Discipline of Microbiology and Immunology, School of Molecular and Biomedical Science, University of Adelaide, Adelaide 5005, Australia.
authors.forename: Elizabeth Ngoc Hoa
authors.initials: ENH
authors.lastname: Tran
authors.affiliation: Discipline of Microbiology and Immunology, School of Molecular and Biomedical Science, University of Adelaide, Adelaide 5005, Australia.
authors.forename: Renato
authors.initials: R
authors.lastname: Morona
mesh_terms: D020816:Amino Acid Motifs; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D015536:Down-Regulation; D004405:Dysentery, Bacillary / Q000378:metabolism / Q000382:microbiology; D006367:HeLa Cells; D006801:Humans; D009154:Mutation; D019081:O Antigens / Q000096:biosynthesis*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D021381:Protein Transport; D012763:Shigella flexneri / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D019081:O Antigens; D014157:Transcription Factors; C495771:WASL protein, human; D051317:Wiskott-Aldrich Syndrome Protein, Neuronal; C058483:virG protein, Shigella flexneri