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Summary Genetic and functional studies have indicated that the type IV secretion system (TFSS) of Helicobacter pylori forms a secretion complex in the cell envelope that protrudes towards the outside in order to inject CagA protein into gastric epithelial cells. However, the proposed structural model is based on partial amino acid homology with the components of the Agrobacterium tumefaciens TFSS. Therefore, we undertook the identification of the structural features of the TFSS exposed on the surface of H. pylori and found that filamentous structures present on the bacterial surface are related to the secretion apparatus. Using immunofluorescence microscopy with antibodies directed to tyrosine-phosphorylated CagA (pY-CagA) and Hp0532 (VirB7) in the infection assay, pY-CagA signals were detected just below the host cell-attached bacteria, where Hp0532 (VirB7) signals were detected as co-localized, suggesting that the CagA injected into the host cell through the TFSS apparatus is still mostly confined to the areas just below the attached bacteria after being phosphorylated. Furthermore, the filamentous structures on bacterium were found to be associated with Hp0532 (VirB7) or Hp0528 (VirB9), the major components of TFSS, by immunogold electron microscopy. These results strongly suggest that the H. pylori TFSS apparatus is a filamentous macromolecular structure protruding from the bacterial envelope. [ABSTRACT FROM AUTHOR]

Summary Shigella , the causative agent of bacillary dysentery, is capable of directing its movement within host cells by forming an actin comet tail. The VirG (IcsA) pro-tein expressed at one pole of the bacterium recruits neural Wiskott–Aldrich syndrome protein (N-WASP), a member of the WASP family, which in turn stimulates actin-related protein (Arp) 2/3 complex-mediated actin polymerization. As all the WASP family proteins induce actin polymerization by recruiting Arp2/3 complex, we investigated their involvement in Shigella motility. Here, we show that VirG binds to N-WASP but not to the other WASP family proteins. Using a series of chimeras obtained by swapping N-WASP and WASP domains, we demonstrated that the specificity of VirG to interact with N-WASP lies in the N-terminal region containing the pleckstrin homology (PH) domain and calmodulin-binding IQ motif of N-WASP. A conformational change in N-WASP was important for the VirG–N-WASP interaction, as elimination of the C-terminal acidic region, which is responsible for the intramolecular interaction with the central basic region of N-WASP, affected the specific binding to VirG. We observed that, in haematopoietic cells such as macrophages, polymorphonuclear leucocytes (PMNs) and platelets, WASP was predominantly expressed, whereas the expression of N-WASP was greatly suppressed. Indeed, unlike Listeria , Shigella was unable to move in macrophages at all, although the movement was restored as N-WASP was expressed ectopically. Thus, our findings demonstrate that N-WASP is a specific ligand of VirG, which determines the host cell type allowing actin-based spreading of Shigella . [ABSTRACT FROM AUTHOR]

Subversion of antigen‐specific immune responses by intracellular pathogens is pivotal for successful colonisation. Bacterial pathogens, including Shigella, deliver effectors into host cells via the type III secretion system (T3SS) in order to manipulate host innate and adaptive immune responses, thereby promoting infection. However, the strategy for subverting antigen‐specific immunity is not well understood. Here, we show that Shigella flexneri invasion plasmid antigen H (IpaH) 4.5, a member of the E3 ubiquitin ligase effector family, targets the proteasome regulatory particle non‐ATPase 13 (RPN13) and induces its degradation via the ubiquitin–proteasome system (UPS). IpaH4.5‐mediated RPN13 degradation causes dysfunction of the 19S regulatory particle (RP) in the 26S proteasome, inhibiting guidance of ubiquitinated proteins to the proteolytically active 20S core particle (CP) of 26S proteasome and thereby suppressing proteasome‐catalysed peptide splicing. This, in turn, reduces antigen cross‐presentation to CD8+ T cells via major histocompatibility complex (MHC) class I in vitro. In RPN13 knockout mouse embryonic fibroblasts (MEFs), loss of RPN13 suppressed CD8+ T cell priming during Shigella infection. Our results uncover the unique tactics employed by Shigella to dampen the antigen‐specific cytotoxic T lymphocyte (CTL) response. [ABSTRACT FROM AUTHOR]