title: Ultrastructural analysis of IpaD at the tip of the nascent MxiH type III secretion apparatus of Shigella flexneri.
abstract: Shigella flexneri is a Gram-negative enteric pathogen that is the predominant cause of bacillary dysentery. Shigella uses a type III secretion system to deliver effector proteins that alter normal target cell functions to promote pathogen invasion. The type III secretion apparatus (T3SA) consists of a basal body, an extracellular needle, and a tip complex that is responsible for delivering effectors into the host cell cytoplasm. IpaD [Ipa (invasion plasmid antigen)] is the first protein to localize to the T3SA needle tip, where it prevents premature effector secretion and serves as an environmental sensor for triggering recruitment of the translocator protein IpaB to the needle tip. Thus, IpaD would be expected to form a stable structure whose overall architecture supports its functions. It is not immediately obvious from the published IpaD crystal structure (Protein Data Bank ID 2j0o) how a multimer of IpaD would be incorporated at the tip of the first static T3SA intermediate, nor what its functional role would be in building a mature T3SA. Here, we produce three-dimensional reconstructions from transmission electron microscopy images of IpaD localized at the Shigella T3SA needle tip for comparison to needle tips from a Shigella ipaD-null mutant. The results demonstrate that IpaD resides as a homopentamer at the needle tip of the T3SA. Furthermore, comparison to tips assembled from the distal domain IpaD(Δ192-267) mutation shows that IpaD adopts an elongated conformation that facilitates its ability to control type III secretion and stepwise assembly of the T3SA needle tip complex.
authors.affiliation: Department of Microbiology and Molecular Genetics, Oklahoma State University, Stillwater, OK 74078, USA.
authors.forename: Chelsea R
authors.initials: CR
authors.lastname: Epler
authors.forename: Nicholas E
authors.initials: NE
authors.lastname: Dickenson
authors.forename: Esther
authors.initials: E
authors.lastname: Bullitt
authors.forename: Wendy L
authors.initials: WL
authors.lastname: Picking
mesh_terms: D000942:Antigens, Bacterial / Q000737:chemistry / Q000648:ultrastructure*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000648:ultrastructure*; D058947:Bacterial Secretion Systems; D021621:Imaging, Three-Dimensional; D008855:Microscopy, Electron, Scanning; D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D012763:Shigella flexneri / Q000737:chemistry* / Q000472:pathogenicity / Q000648:ultrastructure*
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D058947:Bacterial Secretion Systems; C095770:IpaD protein, Shigella flexneri; C413572:MxiH protein, Shigella