Genetic and molecular analyses of the interaction between the flagellum-specific sigma and anti-sigma factors in Salmonella typhimurium.

Kutsukake, Kazuhiro; Iyoda, Sunao; Ohnishi, Kouhei; Iino, Tetsuo

doi:10.1002/j.1460-2075.1994.tb06778.x

Cited by 77 publications

(71 citation statements)

References 39 publications

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“…Besides the necessity of FliA for the transcription of class III genes, FliA may also act as an activator in the transcription of class II genes (25). The activity of FliA in S. typhimurium is negatively regulated by FlgM, which acts as an anti-sigma factor, binding to FliA and preventing its association with the RNA polymerase core (26).…”

Section: Discussionmentioning

confidence: 99%

The alternative sigma factor sigma28 of Legionella pneumophila restores flagellation and motility to an Escherichia coli fliA mutant

1997

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Gene expression in Legionella pneumophila, the etiological agent of Legionnaires' disease, can be controlled by alternative forms of RNA polymerase programmed by distinct factors. To understand the regulation of L. pneumophila flagellin expression, we cloned the factor (FliA) of RNA polymerase responsible for the transcription of the flagellin gene, flaA. FliA is a member of the 28 class of alternative factors identified in several bacterial genera. The gene fliA has been isolated from an expression library of L. pneumophila isolate Corby in Escherichia coli K-12. This library was transformed into a fliA mutant of E. coli K-12 containing a plasmid carrying the L. pneumophila-specific flaA promoter fused to the reporter gene luxAB. Screening the obtained transformants for luciferase activity, we isolated the major part of the fliA gene on a 1.64-kb fragment. This fragment was sequenced and used for reverse PCR in order to recover the complete fliA gene. The resulting 1.03-kb fragment was shown to contain the entire fliA gene. L. pneumophila FliA has 55 and 43% amino acid identity with the homologous sequences of Pseudomonas aeruginosa and E. coli. Furthermore, the L. pneumophila fliA gene was able to restore the flagellation and the motility defect of an E. coli fliA mutant. This result suggests that the L. pneumophila 28 protein can bind to the E. coli core RNA polymerase to direct transcription initiation from the flaA-specific promoter.Legionella pneumophila, a ubiquitous microorganism inhabiting freshwater biotopes and man-made water systems, is also a pathogen of humans causing severe pneumonia termed Legionnaires' disease. In the environment, the bacteria are able to replicate intracellularly in amoebae and other protozoa (14,39,46). Legionella infection occurs after inhalation of aerosolized bacteria. The bacteria enter the human lungs, where they are capable of invading and proliferating in alveolar macrophages and blood monocytes.A growing number of virulence factors elaborated by L. pneumophila contribute to the pathogenicity of the organism. These include a number of genetic loci that have been implicated in being important for intracellular growth (for a review, see reference 6). The icm locus is essential for the intracellular multiplication of L. pneumophila within human macrophages and is also required for virulence in guinea pigs (5, 30). Adjacent to icm is dot, a gene essential for intracellular replication and proper trafficking of host organelles during macrophage infection (3-5). Several surface-associated cellular components have been implicated in virulence. The Mip (macrophage infectivity potentiator) protein contributes to intracellular survival of L. pneumophila, and invasion experiments using a null mutation in mip revealed that Mip is required for early survival processes of L. pneumophila in eukaryotic cells (9, 10, 47). Convincing evidence suggests that the major outer membrane protein (encoded by ompS) is involved in the uptake of bacteria by the host cell (22). The major outer membr...

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Section: Discussionmentioning

confidence: 99%

The alternative sigma factor sigma28 of Legionella pneumophila restores flagellation and motility to an Escherichia coli fliA mutant

1997

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“…Standard DNA procedures, including DNA sequencing and PCR, were performed as described previously (17)(18)(19)33).…”

Section: Methodsmentioning

confidence: 99%

ClpXP Protease Controls Expression of the Type III Protein Secretion System through Regulation of RpoS and GrlR Levels in EnterohemorrhagicEscherichia coli

2005

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Expression of the type III protein secretion system (TTSS), encoded in the locus of enterocyte effacement (LEE) of enterohemorrhagic Escherichia coli (EHEC), has been shown to be controlled by various regulators. In a search for additional regulatory genes, we identified a DNA fragment containing clpX and clpP that has a positive regulatory effect on LEE expression in EHEC O157. The expression of LEE-encoded Esp proteins was significantly reduced in a clpXP deletion mutant. Deletion of grlR, a negative regulatory gene within LEE, markedly increased LEE expression even in the clpXP mutant. To verify the regulatory mechanism of GrlR expression, a chromosomal epitope-tagged allele of grlR (grlR-FLAG) was constructed. GrlR-FLAG expression was increased significantly in the clpXP deletion mutant, suggesting that the GrlR level is under the control of ClpXP, and this regulation is critical for the ClpXP-dependent expression of LEE in EHEC. Deletion of rpoS, the gene encoding a stationary-phase-inducing sigma factor that is a substrate for ClpXP protease, partially restored LEE expression in the clpXP mutant. A multicopy plasmid carrying rpoS strongly repressed expression of Esp proteins, suggesting that positive regulation by ClpXP is partially mediated through a negative effect of RpoS on LEE expression. We also found that rpoS deletion induces transcription of pchA, which encodes one of the positive regulators for LEE expression in EHEC. These results suggest that ClpXP controls expression of LEE through the regulation of RpoS and GrlR levels in EHEC.Enterohemorrhagic Escherichia coli (EHEC) strains are lifethreatening human pathogens and cause hemorrhagic colitis, bloody diarrhea, and hemolytic uremic syndrome (40). EHEC is a member of the attaching and effacing pathogens (40, 56), a group that includes enteropathogenic E. coli (EPEC) (39) and the mouse pathogen Citrobacter rodentium (45). The attaching/effacing lesion on intestinal epithelial cells is characterized by destruction of microvilli and formation of a pedestal-like structure, triggered by rearrangement of cytoskeletal proteins (30, 42). The genes essential for causing the attaching/ effacing lesion are encoded in a pathogenicity island designated the locus of enterocyte effacement (LEE). LEE consists of more than 40 genes organized into five major operons, designated LEE1 to LEE5 (6,8,43). LEE1, -2 and -3 operons contain mostly genes encoding the structural and auxiliary proteins necessary for formation of a dedicated type III protein secretion system (TTSS) (22). The LEE4 operon encodes several secreted proteins (e.g., EspA, EspB, EspD, and EspF), all of which are secreted through TTSS (28, 29, 34, 37). The LEE5 operon encodes an adhesion factor, designated intimin (23, 24), and Tir, a protein that is also translocated through TTSS and acts as a receptor for intimin at the host cell membrane (27).A transcriptional activator, Ler, is encoded by the first gene of the LEE1 operon and is essential for the expression of almost all LEE genes (9, 38). Deng et...

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“…A deletion analysis was carried out on both FlgM and 28 to assess which portions of these proteins were involved in direct interactions (46,69). Mutations in 28 were obtained that express a lac operon fusion to class 3 promoters under FlgMinhibitory conditions (hook-basal body mutant backgrounds) (69; Chadsey and Hughes, submitted).…”

Section: Genetic and Molecular Analysis Of Flgm-28 Interactionmentioning

confidence: 99%

Coupling of Flagellar Gene Expression to Flagellar Assembly in Salmonella enterica Serovar Typhimurium and Escherichia coli

Chilcott

Hughes

2000

Microbiol Mol Biol Rev

591

554

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SUMMARY How do organisms assess the degree of completion of a large structure, especially an extracellular structure such as a flagellum? Bacteria can do this. Mutants that lack key components needed early in assembly fail to express proteins that would normally be added at later assembly stages. In some cases, the regulatory circuitry is able to sense completion of structures beyond the cell surface, such as completion of the external hook structure. In Salmonella and Escherichia coli, regulation occurs at both transcriptional and posttranscriptional levels. One transcriptional regulatory mechanism involves a regulatory protein, FlgM, that escapes from the cell (and thus can no longer act) through a complete flagellum and is held inside when the structure has not reached a later stage of completion. FlgM prevents late flagellar gene transcription by binding the flagellum-specific transcription factor ς28. FlgM is itself regulated in response to the assembly of an incomplete flagellum known as the hook-basal body intermediate structure. Upon completion of the hook-basal body structure, FlgM is exported through this structure out of the cell. Inhibition of ς28-dependent transcription is relieved, and genes required for the later assembly stages are expressed, allowing completion of the flagellar organelle. Distinct posttranscriptional regulatory mechanisms occur in response to assembly of the flagellar type III secretion apparatus and of ring structures in the peptidoglycan and lipopolysaccharide layers. The entire flagellar regulatory pathway is regulated in response to environmental cues. Cell cycle control and flagellar development are codependent. We discuss how all these levels of regulation ensure efficient assembly of the flagellum in response to environmental stimuli.

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Genetic and molecular analyses of the interaction between the flagellum-specific sigma and anti-sigma factors in Salmonella typhimurium.

Cited by 77 publications

References 39 publications

The alternative sigma factor sigma28 of Legionella pneumophila restores flagellation and motility to an Escherichia coli fliA mutant

The alternative sigma factor sigma28 of Legionella pneumophila restores flagellation and motility to an Escherichia coli fliA mutant

ClpXP Protease Controls Expression of the Type III Protein Secretion System through Regulation of RpoS and GrlR Levels in EnterohemorrhagicEscherichia coli

Coupling of Flagellar Gene Expression to Flagellar Assembly in Salmonella enterica Serovar Typhimurium and Escherichia coli

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