Unipolar localization and ATPase activity of IcsA, a Shigella flexneri protein involved in intracellular movement

Goldberg, Marcia B.; Bârzu, Octavian; Parsot, Claude; Sansonetti, Philippe J.

doi:10.1128/jb.175.8.2189-2196.1993

Cited by 272 publications

(318 citation statements)

References 52 publications

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“…Wild-type S. flexneri expresses IcsA at one pole of the bacterium and thus forms polar actin tails in the cytosol of host cells (Ogawa et al, 1968;Bernardini et al, 1989;Pal et al, 1989;Goldberg et al, 1993;Goldberg and Theriot, 1995). Several factors contribute to the polar localization of IcsA, including the rate of diffusion of outer membrane IcsA, which is affected by the O-antigen polysaccharide chain length of the LPS molecules (Sandlin et al, 1995;Sandlin and Maurelli, 1999;Robbins et al, 2001), and the specific cleavage of IcsA in the outer membrane by the protease IcsP (SopA) (d 'Hauteville et al, 1996;Egile et al, 1997;Steinhauer et al, 1999).…”

Section: Resultsmentioning

confidence: 99%

“…IcsA, a 120 kDa outer membrane protein, is located at a single pole on the surface of the bacterium, at the junction with the actin tail (Goldberg et al, 1993). Polar localization of IcsA requires polar protein secretion through a pathway that is common to the four Gramnegative bacteria S. flexneri, Escherichia coli, Yersinia pseudotuberculosis and Salmonella typhimurium Steinhauer et al, 1999;Robbins et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

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Actin-based motility is sufficient for bacterial membrane protrusion formation and host cell uptake

Monack

Theriot

2001

Cellular Microbiology

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SummaryShigella flexneri replicates in the cytoplasm of host cells, where it nucleates host cell actin filaments at one pole of the bacterial cell to form a`comet tail' that propels the bacterium through the host's cytoplasm. To determine whether the ability to move by actinbased motility is sufficient for subsequent formation of membrane-bound protrusions and intercellular spread, we conferred the ability to nucleate actin on a heterologous bacterium, Escherichia coli. Previous work has shown that IcsA (VirG), the molecule that is necessary and sufficient for actin nucleation and actin-based motility, is distributed in a unipolar fashion on the surface of S. flexneri. Maintenance of the unipolar distribution of IcsA depends on both the S. flexneri outer membrane protease IcsP (SopA) and the structure of the lipopolysaccharide (LPS) in the outer membrane. We co-expressed IcsA and IcsP in two strains of E. coli that differed in their LPS structures. The E. coli were engineered to invade host cells by expression of invasin from Yersinia pseudotuberculosis and to escape the phagosome by incubation in purified listeriolysin O (LLO) from Listeria monocytogenes. All E. coli strains expressing IcsA replicated in host cell cytoplasm and moved by actin-based motility. Actin-based motility alone was sufficient for the formation of membrane protrusions and uptake by recipient host cells. The presence of IcsP and an elaborate LPS structure combined to enhance the ability of E. coli to form protrusions at the same frequency as S. flexneri, quantitatively reconstituting this step in pathogen intercellular spread in a heterologous organism. The frequency of membrane protrusion formation across all strains tested correlates with the efficiency of unidirectional actin-based movement, but not with bacterial speed.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Actin-based motility is sufficient for bacterial membrane protrusion formation and host cell uptake

Monack

Theriot

2001

Cellular Microbiology

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“…Wells were then washed with washing solution [phosphate-buffered saline (PBS) containing 0.05% Tween 20] and blocked with blocking buffer (3% bovine serum albumin in PBS). Wells were incubated with affinity-purified IcsA antiserum (Goldberg et al, 1993) for 2 h at room temperature, washed with washing solution and then incubated with a 1:10 000 dilution of horseradish peroxidase-conjugated goat anti-rabbit antiserum for 1 h at room temperature. Visualization was performed by adding to each well 50 l of substrate, which is a solution of 0.4 mg ml ¹1 o-phenylenediamine dihydrochloride (Sigma) in 0.0514 M Na 2 PO 4 , 0.0243 M citric acid, pH 5.0, to which 1.2 × 10 ¹4 % (v/v) H 2 O 2 was added just before use.…”

Section: Methodsmentioning

confidence: 99%

Disruption of IcsP, the major Shigella protease that cleaves IcsA, accelerates actin‐based motility

Shere

Sallustio

Manessis

et al. 1997

Molecular Microbiology

121

136

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SummaryShigella pathogenesis involves bacterial invasion of colonic epithelial cells and movement of bacteria through the cytoplasm and into adjacent cells by means of actin-based motility. The Shigella protein IcsA (VirG) is unipolar on the bacterial surface and is both necessary and sufficient for actin-based motility. IcsA is inserted into the outer membrane as a 120-kDa polypeptide that is subsequently slowly cleaved, thereby releasing the 95-kDa amino-terminal portion into the culture supernatant. IcsP, the major Shigella protease that cleaves IcsA, was identified and cloned. It has significant sequence similarity to the E. coli serine proteases, OmpP and OmpT. Disruption of icsP in serotype 2a S. flexneri leads to a marked reduction in IcsA cleavage, increased amounts of IcsA associated with the bacterium and altered distribution of IcsA on the bacterial surface. The icsP mutant displays significantly increased rates of actin-based motility, with a mean speed 27% faster than the wild-type strain; moreover, a significantly greater percentage of the icsP mutant moves in the cytoplasm. Yet, plaque formation on epithelial monolayers by the mutant was not altered detectably. These data suggest that IcsA, and not a host protein, is limiting in the rate of actinbased motility of wild-type serotype 2a S. flexneri.

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“…One structure is the organized arrays of membrane receptor, which govern chemosensory behavior in swimming bacteria via a phosphor-relay system (26). In Shigella sp., the polar protein IcsA mediates assembly of an actin tail inside infected mammalian cells (27). Polar localization of this protein seems to depend on the cytoskeletal MreB filament (28,29).…”

Section: Dual Fluorescent Labeling Reveals a Mosaic Structure Of The mentioning

confidence: 99%

Biogenesis of actin-like bacterial cytoskeletal filaments destined for positioning prokaryotic magnetic organelles

Pradel

Santini

Bernadac

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

100

108

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Magnetosomes comprise a magnetic nanocrystal surrounded by a lipid bilayer membrane. These unique prokaryotic organelles align inside magnetotactic bacterial cells and serve as an intracellular compass allowing the bacteria to navigate along the geomagnetic field in aquatic environments. Cryoelectron tomography of Magnetospirillum strains has revealed that the magnetosome chain is surrounded by a network of filaments that may be composed of MamK given that the filaments are absent in the mamK mutant cells. The process of the MamK filament assembly is unknown. Here we prove the authenticity of the MamK filaments and show that MamK exhibits linear distribution inside Magnetospirillum sp. cells even in the area without magnetosomes. The mamK gene alone is sufficient to direct the synthesis of straight filaments in Escherichia coli, and one extremity of the MamK filaments is located at the cellular pole. By using dual fluorescent labeling of MamK, we found that MamK nucleates at multiple sites and assembles into mosaic filaments. Time-lapse experiments reveal that the assembly of the MamK filaments is a highly dynamic and kinetically asymmetrical process. MamK bundles might initiate the formation of a new filament or associate to one preexistent filament. Our results demonstrate the mechanism of biogenesis of prokaryotic cytoskeletal filaments that are structurally and functionally distinct from the known MreB and ParM filaments. In addition to positioning magnetosomes, other hypothetical functions of the MamK filaments in magnetotaxis might include anchoring magnetosomes and being involved in magnetic reception.assembly ͉ magnetosomes ͉ prokaryote ͉ magnetic reception

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Unipolar localization and ATPase activity of IcsA, a Shigella flexneri protein involved in intracellular movement

Cited by 272 publications

References 52 publications

Actin-based motility is sufficient for bacterial membrane protrusion formation and host cell uptake

Actin-based motility is sufficient for bacterial membrane protrusion formation and host cell uptake

Disruption of IcsP, the major Shigella protease that cleaves IcsA, accelerates actin‐based motility

Biogenesis of actin-like bacterial cytoskeletal filaments destined for positioning prokaryotic magnetic organelles

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