Neural Wiskott-Aldrich syndrome protein is implicated in the actin-based motility of Shigella flexneri

Suzuki, Toshihiko; Miki, Hiroaki; Takenawa, Tadaomi; Sasakawa, Chihiro

doi:10.1093/emboj/17.10.2767

Cited by 214 publications

(218 citation statements)

References 49 publications

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“…To date, it has been shown that WASP is not essential for actin tail formation of VirG-expressing E. coli in human platelet extracts in vitro (Egile et al, 1999). Recent reports using N-WASP-deficient mouse embryonic fibroblasts have confirmed our previous work showing that N-WASP is required for Shigella motility in mammalian cells (Suzuki et al, 1998). However, it has been shown that the N-WASP-deficient cells lack WASP, and the expression of WAVEs has not been examined.…”

Section: Discussionmentioning

confidence: 90%

“…Recent reports have shown that N-WASP and WASP are involved in actin rearrangements induced by several pathogens. N-WASP is required for the formation of an actin tail by Shigella in the cytoplasm of epithelial cells (Suzuki et al, 1998;Egile et al, 1999), whereas N-WASP (or WASP) is involved in the actin tail formation of vaccinia virus and in the formation of actin pedestal structures beneath enteropathogenic Escherichia coli (EPEC) attached to the host cell (Frischknecht et al, 1999;Kalman et al, 1999). Although the precise mechanisms underlying pathogen-directed cytoskeletal rearrangements remain to be elucidated, studies have clearly indicated that pathogens share similar mechanisms of inducing actin polymerization both to each other and to the host systems, as typically exemplified by the status of N-WASP or Arp2/3 complex in mammalian cells.…”

Section: Discussionmentioning

confidence: 99%

“…The cell lysates were subjected to a pull-down assay using glutathione sepharose beads ligated to GST-a3, the VirG portion encompassing Thr-53-Thr-508 including the N-WASP-binding domain (Suzuki et al, 1998). The proteins co-precipitated with GST-a3 beads were analysed by Western blotting using anti-FLAG antibody.…”

Section: Shigella Virg Binds To N-wasp But Not To the Other Wasp Famimentioning

confidence: 99%

“…In contrast, N-WASP is thought to be ubiquitously expressed (Miki et al, 1996;Suzuki et al, 1998). However, whether the level of N-WASP expression in haematopoietic cells would be the same or not as that in other epithelial or fibroblastic cells has never been seriously investigated yet.…”

Section: Human Macrophages Do Not Support Actin-based Motility Of Intmentioning

confidence: 99%

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Neural Wiskott-Aldrich syndrome protein (N-WASP) is the specific ligand for Shigella VirG among the WASP family and determines the host cell type allowing actin-based spreading

et al. 2002

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SummaryShigella, the causative agent of bacillary dysentery, is capable of directing its movement within host cells by forming an actin comet tail. The VirG (IcsA) protein 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, (V) domain, a domain (C) with homology to the actindepolymerizing protein cofilin and, finally, a C-terminal acidic (A) segment (Miki et al., 1996). The C-terminal VCA domain interacts with the actin-related protein (Arp) 2/3 complex after the stimulation of actin polymerization by the complex (Rohatgi et al., 1999). N-WASP exists in two conformations: an inactive form in which the VCA domain is masked, and an active form in which it is exposed to promote actin polymerization by interaction with the Arp2/3 complex. The activation of N-WASP requires Cdc42 or PtdIns(4,5)P2 bound to N-WASP (Miki et al., 1998b;Rohatgi et al., 1999;2000;Prehoda et al., 2000). It has been reported that N-WASP is required for generating the actin tail from motile S. flexneri (Suzuki et al., 1998;Egile et al., 1999). Arp2/3 complex is also required for actin-based motility of Shigella, in which N-WASP stimulates the actin nucleation activity of the Arp2/3 complex in vitro (Egile et al., 1999).In mammalian cells, five WASP family members, N-WASP, WASP and WAVE1-3, have been identified and characterized to date (Suetsugu et al., 1999); however, none of them except N-WASP has been elucidated for involvement in the actin-based motility of Shigella in infected cells. It has been shown that human platelet extracts supported Shigella actin-based motility as added N-WASP in vitro (Egile et al., 1999). Recent reports using N-WASP-deficient mouse embryonic fibroblasts (Lommel et al., 2001;Snapper et al., 2001) confirmed our previous work indicating that N-WASP is required for Shigella motility in mammalian cells (Suzuki et al., 1998). However...

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Section: Discussionmentioning

confidence: 90%

Section: Discussionmentioning

confidence: 99%

Section: Shigella Virg Binds To N-wasp But Not To the Other Wasp Famimentioning

confidence: 99%

Section: Human Macrophages Do Not Support Actin-based Motility Of Intmentioning

confidence: 99%

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Neural Wiskott-Aldrich syndrome protein (N-WASP) is the specific ligand for Shigella VirG among the WASP family and determines the host cell type allowing actin-based spreading

et al. 2002

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“…When Shigella and Listeria bacteria infect cells, they move inside the cells by forming actin comets. The bacteria use host cell's actin reorganization machinery, including N-WASP and Arp2/3 complex [Suzuki et al, 1998;Welch et al, 1997]. These actin comets are not specific to bacteria.…”

Section: Simplified Model Of Actin Cytoskeleton Reorganization: Actinmentioning

confidence: 99%

Spatial and temporal regulation of actin polymerization for cytoskeleton formation through Arp2/3 complex and WASP/WAVE proteins

Suetsugu

Miki

Takenawa

2002

Cell Motil. Cytoskeleton

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Shigella actin-based motility in the presence of truncated vinculin

Southwick¹,

Adamson²,

Purich³

2000

Cell Motil. Cytoskeleton

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Mounting evidence supports the role of truncated vinculin in the intracellular actin‐based motility of Shigella flexneri. Vinculin's role was recently questioned by Goldberg [1997: Cell Motil Cytoskeleton 37:44–53] who observed Shigella motility in mouse embryonal carcinoma 5.51 cells, a genetically modified cell line that reputedly lacked vinculin. That challenge implicitly relied on the assumption that 5.51 cells had no detectable vinculin polypeptide and lacked full‐length vinculin mRNA. Despite the appearance of being an unambiguous test of vinculin's role in Shigella motility, 5.51 cells were shown to contain adequate amounts of truncated vinculin (as well as the corresponding mRNA transcript) to support bacterial locomotion. We also examined Shigella locomotion in γ229 cells, a related embryonal carcinoma cell line containing approximately one‐half the vinculin content found in 5.51 cells. We observed that there was a commensurate twofold decrease in the Shigella motility rate, as compared to 5.51 cells; this finding raises the possibility that vinculin can become a rate‐limiting factor under some circumstances. Immunofluorescence microscopy using vin 11‐5 monoclonal antibody directed against the vinculin head domain showed intense staining of Shigella rocket tails in both γ229 and 5.51 cells. Our findings clearly demonstrate that motility in 5.51 cells cannot be regarded as a valid criterion for evaluating the role of truncated vinculin in Shigella motility. Cell Motil. Cytoskeleton 45:272–278, 2000 © 2000 Wiley‐Liss, Inc.

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Neural Wiskott-Aldrich syndrome protein is implicated in the actin-based motility of Shigella flexneri

Cited by 214 publications

References 49 publications

Neural Wiskott-Aldrich syndrome protein (N-WASP) is the specific ligand for Shigella VirG among the WASP family and determines the host cell type allowing actin-based spreading

Neural Wiskott-Aldrich syndrome protein (N-WASP) is the specific ligand for Shigella VirG among the WASP family and determines the host cell type allowing actin-based spreading

Spatial and temporal regulation of actin polymerization for cytoskeleton formation through Arp2/3 complex and WASP/WAVE proteins

Shigella actin-based motility in the presence of truncated vinculin

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