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ATPase-independent type-III protein secretion in Salmonella enterica.

Erhardt M, Mertens ME, Fabiani FD, Hughes KT - PLoS Genet. (2014)

Bottom Line: Type-III protein secretion systems are utilized by gram-negative pathogens to secrete building blocks of the bacterial flagellum, virulence effectors from the cytoplasm into host cells, and structural subunits of the needle complex.We additionally show that increased proton motive force bypassed the requirement of the Salmonella pathogenicity island 1 virulence-associated type-III ATPase for secretion.Our data support a role for type-III ATPases in enhancing secretion efficiency under limited secretion substrate concentrations and reveal the dispensability of ATPase activity in the type-III protein export process.

View Article: PubMed Central - PubMed

Affiliation: Junior Research Group Infection Biology of Salmonella, Helmholtz Centre for Infection Research, Braunschweig, Germany.

ABSTRACT
Type-III protein secretion systems are utilized by gram-negative pathogens to secrete building blocks of the bacterial flagellum, virulence effectors from the cytoplasm into host cells, and structural subunits of the needle complex. The flagellar type-III secretion apparatus utilizes both the energy of the proton motive force and ATP hydrolysis to energize substrate unfolding and translocation. We report formation of functional flagella in the absence of type-III ATPase activity by mutations that increased the proton motive force and flagellar substrate levels. We additionally show that increased proton motive force bypassed the requirement of the Salmonella pathogenicity island 1 virulence-associated type-III ATPase for secretion. Our data support a role for type-III ATPases in enhancing secretion efficiency under limited secretion substrate concentrations and reveal the dispensability of ATPase activity in the type-III protein export process.

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Related in: MedlinePlus

Motility of fliHIJ mutants is increased by mutations in ΔatpA, ΔflgM, ΔclpX and fliAH14D.Null mutations in atpA, flgM, clpX, and the more stable FliAH14D variant increased motility of fliHIJ mutant strains in a swimming motility assay using 0.3% soft agar plates. (A) Representative soft agar motility plates after 4.5 hours incubation at 37°C of the FliC-phase locked wildtype and fliHIJ mutant strains. n.d., not determined. (B) Quantified relative motility of fliHIJ mutant strains. The diameter of the motility swarm relative to the wildtype was measured after 4.5 hours incubation. Biological replicates are shown as individual data points. Data were analyzed by the Student's t test. Stars indicate significantly different motility (*, P<0.05; **, P<0.01; ***, P<0.001).
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pgen-1004800-g002: Motility of fliHIJ mutants is increased by mutations in ΔatpA, ΔflgM, ΔclpX and fliAH14D.Null mutations in atpA, flgM, clpX, and the more stable FliAH14D variant increased motility of fliHIJ mutant strains in a swimming motility assay using 0.3% soft agar plates. (A) Representative soft agar motility plates after 4.5 hours incubation at 37°C of the FliC-phase locked wildtype and fliHIJ mutant strains. n.d., not determined. (B) Quantified relative motility of fliHIJ mutant strains. The diameter of the motility swarm relative to the wildtype was measured after 4.5 hours incubation. Biological replicates are shown as individual data points. Data were analyzed by the Student's t test. Stars indicate significantly different motility (*, P<0.05; **, P<0.01; ***, P<0.001).

Mentions: We constructed deletion mutants of clpX and atpA and tested the suppressor function in various fliHIJ deletion backgrounds by monitoring their swimming behavior in soft agar plates (Fig. 2A and Fig. S2). The absence of the FOF1 ATPase restored motility of a fliI deletion mutant to about 5% of the wildtype, of a fliHI deletion mutant to about 25% of the wildtype and of a fliHIJ deletion mutant to about 11% of the wildtype (Fig. 2B). In an otherwise wildtype background, deletion of atpA resulted in a pronounced growth defect (Fig. S3), however did not affect the free-swimming velocity (Fig. S4).


ATPase-independent type-III protein secretion in Salmonella enterica.

Erhardt M, Mertens ME, Fabiani FD, Hughes KT - PLoS Genet. (2014)

Motility of fliHIJ mutants is increased by mutations in ΔatpA, ΔflgM, ΔclpX and fliAH14D.Null mutations in atpA, flgM, clpX, and the more stable FliAH14D variant increased motility of fliHIJ mutant strains in a swimming motility assay using 0.3% soft agar plates. (A) Representative soft agar motility plates after 4.5 hours incubation at 37°C of the FliC-phase locked wildtype and fliHIJ mutant strains. n.d., not determined. (B) Quantified relative motility of fliHIJ mutant strains. The diameter of the motility swarm relative to the wildtype was measured after 4.5 hours incubation. Biological replicates are shown as individual data points. Data were analyzed by the Student's t test. Stars indicate significantly different motility (*, P<0.05; **, P<0.01; ***, P<0.001).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230889&req=5

pgen-1004800-g002: Motility of fliHIJ mutants is increased by mutations in ΔatpA, ΔflgM, ΔclpX and fliAH14D.Null mutations in atpA, flgM, clpX, and the more stable FliAH14D variant increased motility of fliHIJ mutant strains in a swimming motility assay using 0.3% soft agar plates. (A) Representative soft agar motility plates after 4.5 hours incubation at 37°C of the FliC-phase locked wildtype and fliHIJ mutant strains. n.d., not determined. (B) Quantified relative motility of fliHIJ mutant strains. The diameter of the motility swarm relative to the wildtype was measured after 4.5 hours incubation. Biological replicates are shown as individual data points. Data were analyzed by the Student's t test. Stars indicate significantly different motility (*, P<0.05; **, P<0.01; ***, P<0.001).
Mentions: We constructed deletion mutants of clpX and atpA and tested the suppressor function in various fliHIJ deletion backgrounds by monitoring their swimming behavior in soft agar plates (Fig. 2A and Fig. S2). The absence of the FOF1 ATPase restored motility of a fliI deletion mutant to about 5% of the wildtype, of a fliHI deletion mutant to about 25% of the wildtype and of a fliHIJ deletion mutant to about 11% of the wildtype (Fig. 2B). In an otherwise wildtype background, deletion of atpA resulted in a pronounced growth defect (Fig. S3), however did not affect the free-swimming velocity (Fig. S4).

Bottom Line: Type-III protein secretion systems are utilized by gram-negative pathogens to secrete building blocks of the bacterial flagellum, virulence effectors from the cytoplasm into host cells, and structural subunits of the needle complex.We additionally show that increased proton motive force bypassed the requirement of the Salmonella pathogenicity island 1 virulence-associated type-III ATPase for secretion.Our data support a role for type-III ATPases in enhancing secretion efficiency under limited secretion substrate concentrations and reveal the dispensability of ATPase activity in the type-III protein export process.

View Article: PubMed Central - PubMed

Affiliation: Junior Research Group Infection Biology of Salmonella, Helmholtz Centre for Infection Research, Braunschweig, Germany.

ABSTRACT
Type-III protein secretion systems are utilized by gram-negative pathogens to secrete building blocks of the bacterial flagellum, virulence effectors from the cytoplasm into host cells, and structural subunits of the needle complex. The flagellar type-III secretion apparatus utilizes both the energy of the proton motive force and ATP hydrolysis to energize substrate unfolding and translocation. We report formation of functional flagella in the absence of type-III ATPase activity by mutations that increased the proton motive force and flagellar substrate levels. We additionally show that increased proton motive force bypassed the requirement of the Salmonella pathogenicity island 1 virulence-associated type-III ATPase for secretion. Our data support a role for type-III ATPases in enhancing secretion efficiency under limited secretion substrate concentrations and reveal the dispensability of ATPase activity in the type-III protein export process.

Show MeSH
Related in: MedlinePlus