Identification and characterization of the products of six region III flagellar genes (flaAII.3 through flaQII) of Salmonella typhimurium

Homma, Mitsuo; Iino, Tetsuo; Macnab, Robert M.

doi:10.1128/jb.170.5.2221-2228.1988

Cited by 29 publications

(28 citation statements)

References 32 publications

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“…Inconsistencies tended to occur at the vicinity of a segment boundary, or where a segment was small, and probably reflect ambiguities in the recombination analysis. Two mutations that were originally mapped to segment 1 of fliM (41) actually lie infliL, a gene that was discovered later (7,12); analysis of thefliL gene will be presented separately.…”

Section: Resultsmentioning

confidence: 99%

Molecular analysis of the flagellar switch protein FliM of Salmonella typhimurium

et al. 1992

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Previous studies have led to the conclusion that in Salmonella typhimurium (and also in Escherichia coli) there are three flagellar proteins, FliG, FliM, and FliN, which function together in enabling motor rotation and in controlling its direction of rotation (2,4,10,35,41). The part of the flagellum constructed from these proteins is called the flagellar switch. The state of the switch, counterclockwise (CCW) or clockwise (CW), is influenced by the chemotaxis sensory system, such that CCW rotation is enhanced by favorable stimuli and CW rotation is enhanced by unfavorable stimuli (14). On the basis of intergenic suppression analysis in E. coli (24) and the effect of CheY on cell envelopes lacking any of the other chemotaxis proteins (25), CheY is thought to bind to the switch and bias it to the CW state. CheZ has the opposite effect on switch bias (13), and intergenic suppression analysis (23,24) suggests that it, too, binds to the switch; however, in this case no biochemical evidence is available to support the hypothesis. CheY is activated by CheA (by phosphorylation of aspartate 57) and deactivated by CheZ (by dephosphorylation) (1,6,27,38).

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Section: Resultsmentioning

confidence: 99%

Molecular analysis of the flagellar switch protein FliM of Salmonella typhimurium

et al. 1992

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“…Both fliI and orf4 are parts of a flagellar operon, and the hypothetical ATPase could be involved in some steps of the assembly of flagellar components. It may be relevant that the S. typhimurium Flil protein was found to be associated mostly with the membrane fraction (12).…”

Section: Discussionmentioning

confidence: 99%

“…For strain DP50, the medium was supplemented with diaminopimelic acid (100 p,g/ml) and thymidine (50 ,ug/ml). TY medium was solidified by adding 12 Scoring for motility. Colonies were transferred with toothpicks to swarm plates prepared with semisolid medium B (Difco Bacto-Peptone, 10 g; Difco gelatin, 80 g; Difco agar, 4 g; NaCl, 5 g; water to 1 liter).…”

Section: Methodsmentioning

confidence: 99%

The flaA locus of Bacillus subtilis is part of a large operon coding for flagellar structures, motility functions, and an ATPase-like polypeptide

et al. 1991

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We cloned and sequenced 8.3 kb of Bacillus subtilis DNA corresponding to theflaA locus involved in flagellar biosynthesis, motility, and chemotaxis. The DNA sequence revealed the presence of 10 complete and 2 incomplete open reading frames. Comparison of the deduced amino acid sequences to data banks showed similarities of nine of the deduced products to a number of proteins of Escherichia coli and Salmonella typhimurium for which a role in flagellar functioning has been directly demonstrated. In particular, the sequence data suggest that theflaA operon codes for the M-ring protein, components of the motor switch, and the distal part of the basal-body rod. The gene order is remarkably similar to that described for region III of the enterobacterial flagellar regulon. One of the open reading frames was translated into a protein with 48% amino acid identity to S. typhimurium Fliu and 29% identity to the 0 subunit of E. coli ATP synthase.Many bacterial species swim actively by means of flagella. Swimming tends to move the bacteria towards attractants and away from repellents. The flagellar organelle is made of three parts: the basal body, the hook, and the filament, with more than 10 different proteins involved in flagellar formation. In addition, there are the motor apparatus and the switch, necessary for rotation of the flagellar filaments and control of the sense (clockwise or counterclockwise) of rotation. A number of sensory elements and components for transduction of the signal to the motor switch are also part of the motility system. A considerable body of data is available regarding the structure and functioning of the flagella of Escherichia coli and Salmonella typhimurium (reviewed in references 21 and 22).About the gram-positive bacterium Bacillus subtilis much less is known. A limited number of mutants affected in motility have been isolated, and a few have been characterized (8, 32). More advanced are the studies related to the chemotactic mechanisms that operate in B. subtilis. A large number of mutants defective in chemotaxis have been isolated, 21 different complementation groups have been identified, and part of a chemotactic locus has been cloned (28, 29). Better knowledge of the genes and gene products of B. subtilis involved in flagellar assembly, motility, and taxis seemed desirable in view of the fact that B. subtilis is a gram-positive bacterium and that its chemotactic response is more complex than that of E. coli (37). From a B. subtilis chromosomal DNA library in XCharon 4A (5) we have isolated one clone that contains the wild-type alleles of two known flaA mutations, flaA4 and flaA15 (9).In this report, we describe the nucleotide sequence of 8,302 contiguous nucleotides of the flaA locus. Insertional inactivation with integrative plasmids showed that the region sequenced is part of a single operon involved in motility (10, 44). Nine of the deduced gene products show similarity to E. coli and S. typhimurium proteins known to be involved in * Corresponding author. flagellar structures and fu...

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“…FliL is found in the membrane fraction in minicell experiments (3,11), and on the basis of its sequence, it would be expected to span the membrane just once and have a C-terminal domain in the periplasm. Its role in the flagellar system is unknown.…”

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confidence: 99%

“…The flagellar regulons of Escherichia coli and Salmonella typhimurium contain exclusively genes concerned with flagellation, motility, and chemotaxis (10).fliL is the first gene in one of the flagellar operons (3,7) and is immediately followed by fliM and fliN, two of the three genes responsible for flagellar rotation and switching between the counterclockwise and clockwise directions (13); it was discovered as a result of cloning the adjacent gene, fliM (3,7), having escaped attention in searches for mutants. FliL is found in the membrane fraction in minicell experiments (3,11), and on the basis of its sequence, it would be expected to span the membrane just once and have a C-terminal domain in the periplasm.…”

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Characterization of the fliL gene in the flagellar regulon of Escherichia coli and Salmonella typhimurium

1994

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fliL is a small gene of unknown function that lies within the beginning of a large flagellar operon of Salmonella typhimurium and Escherichia coli. A spontaneousfliL mutant ofS. typhimurium, containing a frameshift mutation about 40% from the 3' end of the gene, was moderately motile but swarmed poorly, suggesting that FliL might be a component of the flagellar motor or switch. However, in-frame deletions of the E. coli gene, including an essentially total deletion, had little or no effect on motility or chemotaxis. Thus, FliL does not appear to have a major role in flagellar structure or function and is therefore unlikely to be a component of the motor or switch; the elfect on motility caused by truncation of the gene is probably an indirect one.The flagellar regulons of Escherichia coli and Salmonella typhimurium contain exclusively genes concerned with flagellation, motility, and chemotaxis (10).fliL is the first gene in one of the flagellar operons (3, 7) and is immediately followed by fliM and fliN, two of the three genes responsible for flagellar rotation and switching between the counterclockwise and clockwise directions (13); it was discovered as a result of cloning the adjacent gene, fliM (3, 7), having escaped attention in searches for mutants. FliL is found in the membrane fraction in minicell experiments (3, 11), and on the basis of its sequence, it would be expected to span the membrane just once and have a C-terminal domain in the periplasm. Its role in the flagellar system is unknown.Malakooti et al. (11) found that an E. coli strain, JM7623 (Fig. lb), whose fliL gene had been disrupted by a kanamycin resistance gene (Kmr) cassette, was nonflagellate; they concluded that the mutation was not polar on downstream genes and that the null phenotype for fliL was nonflagellate.We subsequently encountered in S. typhimurium the first example of a spontaneous fliL mutant, SJW2295 (12). This strain carried a severe truncation of the gene yet was motile. Its properties have led us to examine more closely the role of the fliL gene. MATERIALS AND METHODSBacterial strains and plasmids. GM2163 (New England Biolabs, Beverly, Mass.) was used for transformations with DNA containing Clal sites, to avoid dam methylation. UH869 (4) was used as a minicell-producing strain. AW330 (1) and EKK9 (8) are E. coli strains that are wild type for motility and chemotaxis; we used AW330 initially but later changed to EKK9 because we found it had better motility and a higher level of transformation efficiency. E. coli JM7623 (11) (2), with slight modification in the case of the BamHIClaI deletion allele of fliL in plasmid pMR14 (see Results), with which transformation was accomplished by electroporation, and the transformed cells (after 1 h of growth at 30°C to recover) were diluted into 5 ml of Luria broth plus 10 ,ug of chloramphenicol per ml and grown overnight at 44°C to enrich for recombinants before plating. RESULTSProperties of a S. typhimurium mutant with a truncatedfliL gene. Strain SJW2295 (13), a poorly swarming derivat...

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Identification and characterization of the products of six region III flagellar genes (flaAII.3 through flaQII) of Salmonella typhimurium

Cited by 29 publications

References 32 publications

Molecular analysis of the flagellar switch protein FliM of Salmonella typhimurium

Molecular analysis of the flagellar switch protein FliM of Salmonella typhimurium

The flaA locus of Bacillus subtilis is part of a large operon coding for flagellar structures, motility functions, and an ATPase-like polypeptide

Characterization of the fliL gene in the flagellar regulon of Escherichia coli and Salmonella typhimurium

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