Mechanism of Shigella entry into epithelial cells

Nhieu, Guy Tran Van; Sansonetti, Philippe J.

doi:10.1016/s1369-5274(99)80009-5

Cited by 146 publications

(71 citation statements)

References 21 publications

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“…The bacterium then enters macrophages and induces them to undergo apoptosis (4), which leads to release of the pathogen at the basolateral side of the colonic epithelium. Shigella then promotes its own uptake into epithelial cells by inducing cytoskeletal rearrangements in these cells (5,6). The pathogenesis of S. flexneri requires a functional type III secretion system (6).…”

Section: From the Department Of Molecular Biosciences University Of mentioning

confidence: 99%

Identification of the MxiH Needle Protein Residues Responsible for Anchoring Invasion Plasmid Antigen D to the Type III Secretion Needle Tip

Zhang

Wang

Olive

et al. 2007

Journal of Biological Chemistry

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The pathogenesis of Shigella flexneri requires a functional type III secretion apparatus to serve as a conduit for injecting host-altering effector proteins into the membrane and cytoplasm of the targeted cell. The type III secretion apparatus is composed of a basal body and an exposed needle that is an extended polymer of MxiH with a 2.0-nm inner channel. Invasion plasmid antigen D (IpaD) resides at the tip of the needle to control type III secretion. The atomic structures of MxiH and IpaD have been solved. MxiH (8.3 kDa) is a helix-turn-helix, whereas IpaD (36.6 kDa) has a dumbbell shape with two globular domains flanking a central coiled-coil that stabilizes the protein. These structures alone, however, have not been sufficient to produce a workable in silico model by which IpaD docks at the needle tip. Thus, the work presented here provides an initial step in understanding this important protein-protein interaction. We have identified key MxiH residues located in its PSNP loop and the contiguous surface that uniquely contribute to the formation of the IpaD-needle interface as determined by NMR chemical shift mapping. Mutation of Asn-43, Leu-47, and Tyr-50 residues severely affects the stable maintenance of IpaD at the Shigella surface and thus compromises the invasive phenotype of S. flexneri. Other residues could be mutated to give rise to intermediate phenotypes, suggesting they have a role in tip complex stabilization while not being essential for tip complex formation. Initial in vitro fluorescence polarization studies confirmed that specific amino acid changes adversely affect the MxiH-IpaD interaction. Meanwhile, none of the mutations appeared to have a negative effect on the MxiH-MxiH interactions required for efficient needle assembly.

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Section: From the Department Of Molecular Biosciences University Of mentioning

confidence: 99%

Identification of the MxiH Needle Protein Residues Responsible for Anchoring Invasion Plasmid Antigen D to the Type III Secretion Needle Tip

Zhang

Wang

Olive

et al. 2007

Journal of Biological Chemistry

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“…IpaD localizes to the tip of the needle prior to secretion and likely serves as a scaffold for the assembly of the hydrophobic IpaBC translocon (Espina et al, 2006). IpaB and IpaC also have effector functions: while microinjected IpaC triggers actin polymerization and filopodia formation in epithelial cells (Tran Van Nhieu et al, 1999), microinjected IpaB triggers cell death in macrophages (Chen et al, 1996). Moreover, upon contact with epithelial cells IpaB binds to the hyaluron receptor CD44 (Skoudy et al, 2000).…”

Section: Introductionmentioning

confidence: 99%

Intracellular type III secretion by cytoplasmic Shigella flexneri promotes caspase-1-dependent macrophage cell death

2007

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The Gram-negative bacterium Shigella flexneri triggers pro-inflammatory apoptotic cell death in macrophages, which is crucial for the onset of an acute inflammatory diarrhoea termed bacillary dysentery. The Mxi-Spa type III secretion system promotes bacterial uptake and escape into the cytoplasm, where, dependent on the translocator/effector protein IpaB, caspase-1 [interleukin (IL)-1b-converting enzyme] and its substrate IL-1b are activated. Here, we show that in the course of a macrophage infection, IpaB is secreted intracellularly for more than 1 h post-infection and progressively accumulates in aggregates on the bacterial surface. Concomitantly, the bacterial pool of IpaB is gradually depleted. The protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) dose-dependently inhibited the Mxi-Spa-dependent secretion of IpaB triggered by the dye Congo red in vitro and abolished translocation of IpaB into the host-cell cytoplasm of S. flexneri-infected macrophages. CCCP specifically inhibited S. flexneri-triggered macrophage death in a dose-dependent manner, even if added up to 60 min post-infection. Addition of CCCP 15 min after infection blocked macrophage cell death, the activation of caspase-1 and the maturation of IL-1b, without affecting uptake or escape of S. flexneri from the phagosome. By contrast, CCCP used at the same concentration had no effect on ATP-induced caspase-1 activation or staurosporine-induced apoptosis. Our results indicate that under the conditions used, CCCP rapidly and specifically blocks bacterial type III secretion, and thus, intracellular type III secretion promotes cytotoxicity of S. flexneri. INTRODUCTIONMany symbiotic or pathogenic Gram-negative bacteria employ a type III secretion system (T3SS) to establish a replicative niche (Cornelis, 2006;Galan & Wolf-Watz, 2006). The complex T3SSs are evolutionarily related to flagellar secretion systems and composed of a basal structure spanning the two bacterial membranes, a hollow needle (or pilus/filament) and a hydrophobic translocator complex inserted into eukaryotic target membranes. Some T3SSs deliver more than 20 different 'effector' proteins into target cells, where they subvert cellular functions in favour of the bacteria. In the eukaryotic host cells, the bacterial effectors target phosphoinositide metabolism, GTP cycles, phosphoprotein signalling, cytoskeletal and vesicle dynamics, or apoptotic pathways (Cossart & Sansonetti, 2004;Hilbi, 2006;Rosenberger & Finlay, 2003;Schlumberger & Hardt, 2006). Shigella flexneri, the causative agent of a severe diarrhoea termed bacillary dysentery (shigellosis), employs a T3SS to invade non-phagocytic epithelial cells and to kill macrophages (Cossart & Sansonetti, 2004). The S. flexneri T3SS is encoded by the mxi and spa genes on the 214 kb virulence plasmid (Buchrieser et al., 2000). Investigations by electron microscopy revealed that the Mxi-Spa needle complex is a canonical T3SS composed of a basal body and a needle, the major subunit of which (MxiH) is required for the secretion of eff...

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“…Estudos mostraram que o escape da Shigella do vacúolo fagocitico das células epiteliais é dependente do sistema de secreção e das proteínas ipaB e ipaC (HIGH et al, 1992;TRAN VAN NHIEU et al, 1999). Avaliamos a transcrição relativa dos genes ipaB e ipaC das três cepas de EIEC nas duas primeiras horas de infecção pós-fagocitose.…”

Section: Discussionunclassified

“…Dentre as proteínas efetoras secretadas por EIEC e Shigella, a ipaB foi descrita como essencial na invasão bacteriana na célula epitelial intestinal (HIGH et al, 1992; TRAN VAN NHIEU et al, 1999), e nos macrófagos foi demonstrado que a ipaB e a ipaC desempenham um papel crucial na sobrevivência bacteriana intracelular após a fagocitose (KUWAE et al, 2001 …”

Section: E Coli Enteroinvasora (Eiec) E Coli Enteropatogênica (Epeunclassified

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O papel da flagelina e do sistema de secreção de Escherichia coli enteroinvasora na resposta imune inata dos macrófagos

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Mechanism of Shigella entry into epithelial cells

Cited by 146 publications

References 21 publications

Identification of the MxiH Needle Protein Residues Responsible for Anchoring Invasion Plasmid Antigen D to the Type III Secretion Needle Tip

Identification of the MxiH Needle Protein Residues Responsible for Anchoring Invasion Plasmid Antigen D to the Type III Secretion Needle Tip

Intracellular type III secretion by cytoplasmic Shigella flexneri promotes caspase-1-dependent macrophage cell death

O papel da flagelina e do sistema de secreção de Escherichia coli enteroinvasora na resposta imune inata dos macrófagos

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