Identification of genes encoding components of the swarmer cell flagellar motor and propeller and a sigma factor controlling differentiation of Vibrio parahaemolyticus.

ABSTRACT Vibrio parahaemolyticus possesses two types of flagella, polar and lateral, powered by distinct energy sources, which are derived from the sodium and proton motive forces, respectively. Although proton-powered flagella in Escherichia coli andSalmonella enterica serovar Typhimurium have been extensively studied, the mechanism of torque generation is still not understood. Molecular knowledge of the structure of the sodium-driven motor is only now being developed. In this work, we identify the switch components, FliG, FliM, and FliN, of the sodium-type motor. This brings the total number of genes identified as pertinent to polar motor function to seven. Both FliM and FliN possess charged domains not found in proton-type homologs; however, they can interact with the proton-type motor of E. coli to a limited extent. Residues known to be critical for torque generation in the proton-type motor are conserved in the sodium-type motor, suggesting a common mechanism for energy transfer at the rotor-stator interface regardless of the driving force powering rotation. Mutants representing a complete panel of insertionally inactivated switch and motor genes were constructed. All of these mutants were defective in sodium-driven swimming motility. Alkaline phosphatase could be fused to the C termini of MotB and MotY without abolishing motility, whereas deletion of the unusual, highly charged C-terminal domain of FliM disrupted motor function. All of the mutants retained proton-driven, lateral motility over surfaces. Thus, although central chemotaxis genes are shared by the polar and lateral systems, genes encoding the switch components, as well as the motor genes, are distinct for each motility system.

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Bacterial Swarming Reduces Proteus mirabilis and Vibrio parahaemolyticus Cell Stiffness and Increases β-Lactam Susceptibility

mBio ◽

10.1128/mbio.00210-19 ◽

2019 ◽

Vol 10 (5) ◽

Cited By ~ 2

Author(s):

George K. Auer ◽

Piercen M. Oliver ◽

Manohary Rajendram ◽

Ti-Yu Lin ◽

Qing Yao ◽

...

Keyword(s):

Cell Wall ◽

Osmotic Pressure ◽

Proteus Mirabilis ◽

Vibrio Parahaemolyticus ◽

Cell Stiffness ◽

Bending Rigidity ◽

Vegetative Cells ◽

Swarmer Cell ◽

Content Type ◽

Peptidoglycan Layer

ABSTRACT Swarmer cells of the Gram-negative uropathogenic bacteria Proteus mirabilis and Vibrio parahaemolyticus become long (>10 to 100 μm) and multinucleate during their growth and motility on polymer surfaces. We demonstrated that the increasing cell length is accompanied by a large increase in flexibility. Using a microfluidic assay to measure single-cell mechanics, we identified large differences in the swarmer cell stiffness (bending rigidity) of P. mirabilis (5.5 × 10−22 N m2) and V. parahaemolyticus (1.0 × 10−22 N m2) compared to vegetative cells (1.4 × 10−20 N m2 and 2.2 × 10−22 N m2, respectively). The reduction in bending rigidity (∼2-fold to ∼26-fold) was accompanied by a decrease in the average polysaccharide strand length of the peptidoglycan layer of the cell wall from 28 to 30 disaccharides to 19 to 22 disaccharides. Atomic force microscopy revealed a reduction in P. mirabilis peptidoglycan thickness from 1.5 nm (vegetative cells) to 1.0 nm (swarmer cells), and electron cryotomography indicated changes in swarmer cell wall morphology. P. mirabilis and V. parahaemolyticus swarmer cells became increasingly sensitive to osmotic pressure and susceptible to cell wall-modifying antibiotics (compared to vegetative cells)—they were ∼30% more likely to die after 3 h of treatment with MICs of the β-lactams cephalexin and penicillin G. The adaptive cost of “swarming” was offset by the increase in cell susceptibility to physical and chemical changes in their environment, thereby suggesting the development of new chemotherapies for bacteria that leverage swarming for the colonization of hosts and for survival. IMPORTANCE Proteus mirabilis and Vibrio parahaemolyticus are bacteria that infect humans. To adapt to environmental changes, these bacteria alter their cell morphology and move collectively to access new sources of nutrients in a process referred to as “swarming.” We found that changes in the composition and thickness of the peptidoglycan layer of the cell wall make swarmer cells of P. mirabilis and V. parahaemolyticus more flexible (i.e., reduce cell stiffness) and that they become more sensitive to osmotic pressure and cell wall-targeting antibiotics (e.g., β-lactams). These results highlight the importance of assessing the extracellular environment in determining antibiotic doses and the use of β-lactam antibiotics for treating infections caused by swarmer cells of P. mirabilis and V. parahaemolyticus.

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Cloning and expression of two genes encoding highly homologous hemolysins from a Kanagawa-phenomenon-positive Vibrio parahaemolyticus T4750 strain

Gene ◽

10.1016/0378-1119(90)90129-f ◽

1990 ◽

Vol 93 (1) ◽

pp. 9-15 ◽

Cited By ~ 21

Author(s):

Tetsuya Iida ◽

Koichiro Yamamoto

Keyword(s):

Vibrio Parahaemolyticus ◽

Cloning And Expression ◽

Genes Encoding

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Occurrence of mutations impairing sigma factor B (SigB) function upon inactivation of Listeria monocytogenes genes encoding surface proteins

Microbiology ◽

10.1099/mic.0.067744-0 ◽

2013 ◽

Vol 159 (Pt_7) ◽

pp. 1328-1339 ◽

Cited By ~ 10

Author(s):

Juan J. Quereda ◽

M. Graciela Pucciarelli ◽

Laura Botello-Morte ◽

Enrique Calvo ◽

Filipe Carvalho ◽

...

Keyword(s):

Listeria Monocytogenes ◽

Sigma Factor ◽

Surface Proteins ◽

Factor B ◽

Genes Encoding ◽

Sigma Factor B

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Chitin Heterodisaccharide, Released from Chitin by Chitinase and Chitin Oligosaccharide Deacetylase, Enhances the Chitin-Metabolizing Ability of Vibrio parahaemolyticus

Journal of Bacteriology ◽

10.1128/jb.00270-19 ◽

2019 ◽

Vol 201 (20) ◽

Cited By ~ 1

Author(s):

Takako Hirano ◽

Manabu Okubo ◽

Hironobu Tsuda ◽

Masahiro Yokoyama ◽

Wataru Hakamata ◽

...

Keyword(s):

Vibrio Parahaemolyticus ◽

Pcr Analysis ◽

Content Type ◽

Chitin Degradation ◽

Expression Of Genes ◽

Peptide Mass ◽

Degrading Bacteria ◽

Genes Encoding ◽

Chitinase Production

ABSTRACT Vibrio parahaemolyticus RIMD2210633 secretes both chitinase and chitin oligosaccharide deacetylase and produces β-N-acetyl-d-glucosaminyl-(1,4)-d-glucosamine (GlcNAc-GlcN) from chitin. Previously, we reported that GlcNAc-GlcN induces chitinase production by several strains of Vibrio harboring chitin oligosaccharide deacetylase genes (T. Hirano, K. Kadokura, T. Ikegami, Y. Shigeta, et al., Glycobiology 19:1046–1053, 2009). The metabolism of chitin by Vibrio was speculated on the basis of the findings of previous studies, and the role of chitin oligosaccharide produced from chitin has been well studied. However, the role of GlcNAc-GlcN in the Vibrio chitin degradation system, with the exception of the above-mentioned function as an inducer of chitinase production, remains unclear. N,N′-Diacetylchitobiose, a homodisaccharide produced from chitin, is known to induce the expression of genes encoding several proteins involved in chitin metabolism in Vibrio strains (K. L. Meibom, X. B. Li, A. Nielsen, C. Wu, et al., Proc Natl Acad Sci U S A 101:2524–2529, 2004). We therefore hypothesized that GlcNAc-GlcN also affects the expression of enzymes involved in chitin metabolism in the same manner. In this study, we examined the induction of protein expression by several sugars released from chitin using peptide mass fingerprinting and confirmed the expression of genes encoding enzymes involved in chitin metabolism using real-time quantitative PCR analysis. We then confirmed that GlcNAc-GlcN induces the expression of genes encoding many soluble enzymes involved in chitin degradation in Vibrio parahaemolyticus. Here, we demonstrate that GlcNAc-GlcN enhances the chitin-metabolizing ability of V. parahaemolyticus. IMPORTANCE We demonstrate that β-N-acetyl-d-glucosaminyl-(1,4)-d-glucosamine (GlcNAc-GlcN) enhances the chitin-metabolizing ability of V. parahaemolyticus. Members of the genus Vibrio are chitin-degrading bacteria, and some species of this genus are associated with diseases affecting fish and animals, including humans (F. L. Thompson, T. Iida, and J. Swings, Microbiol Mol Biol Rev 68:403–431, 2004; M. Y. Ina-Salwany, N. Al-Saari, A. Mohamad, F.-A. Mursidi, et al., J Aquat Anim Health 31:3–22, 2019). Studies on Vibrio are considered important, as they may facilitate the development of solutions related to health, food, and aquaculture problems attributed to this genus. This report enhances the current understanding of chitin degradation by Vibrio bacteria.

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Characterization of Vibrio parahaemolyticus genes encoding the systems for utilization of enterobactin as a xenosiderophore

Microbiology ◽

10.1099/mic.0.059568-0 ◽

2012 ◽

Vol 158 (8) ◽

pp. 2039-2049 ◽

Cited By ~ 12

Author(s):

Tomotaka Tanabe ◽

Tatsuya Funahashi ◽

Keiichi Shiuchi ◽

Noriyuki Okajima ◽

Hiroshi Nakao ◽

...

Keyword(s):

Vibrio Parahaemolyticus ◽

Genes Encoding

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Regulation of σB by an Anti- and an Anti-Anti-Sigma Factor in Streptomyces coelicolor in Response to Osmotic Stress

Journal of Bacteriology ◽

10.1128/jb.186.24.8490-8498.2004 ◽

2004 ◽

Vol 186 (24) ◽

pp. 8490-8498 ◽

Cited By ~ 37

Author(s):

Eun-Jin Lee ◽

You-Hee Cho ◽

Hyo-Sub Kim ◽

Bo-Eun Ahn ◽

Jung-Hye Roe

Keyword(s):

Osmotic Stress ◽

Sigma Factor ◽

Target Genes ◽

Sequence Similarity ◽

Streptomyces Coelicolor ◽

Dependent Manner ◽

Its Gene ◽

Genes Encoding

ABSTRACT σB, a homolog of stress-responsive σB of Bacillus subtilis, controls both osmoprotection and differentiation in Streptomyces coelicolor A3 (2). Its gene is preceded by rsbA and rsbB genes encoding homologs of an anti-sigma factor, RsbW, and its antagonist, RsbV, of B. subtilis, respectively. Purified RsbA bound to σB and prevented σB-directed transcription from the sigBp1 promoter in vitro. An rsbA-null mutant exhibited contrasting behavior to the sigB mutant, with elevated sigBp1 transcription, no actinorhodin production, and precocious aerial mycelial formation, reflecting enhanced activity of σB in vivo. Despite sequence similarity to RsbV, RsbB lacks the conserved phosphorylatable serine residue and its gene disruption produced no distinct phenotype. RsbV (SCO7325) from a putative six-gene operon (rsbV-rsbR-rsbS-rsbT-rsbU1-rsbU) was strongly induced by osmotic stress in a σB-dependent manner. It antagonized the inhibitory action of RsbA on σB-directed transcription and was phosphorylated by RsbA in vitro. These results support the hypothesis that the rapid induction of σB target genes by osmotic stress results from modulation of σB activity by the kinase-anti-sigma factor RsbA and its phosphorylatable antagonist RsbV, which function by a partner-switching mechanism. Amplified induction could result from a rapid increase in the synthesis of both σB and its inhibitor antagonist.

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Multiple groESL Operons Are Not Key Targets of RpoH1 and RpoH2 in Sinorhizobium meliloti

Journal of Bacteriology ◽

10.1128/jb.188.10.3507-3515.2006 ◽

2006 ◽

Vol 188 (10) ◽

pp. 3507-3515 ◽

Cited By ~ 20

Author(s):

Alycia N. Bittner ◽

Valerie Oke

Keyword(s):

Escherichia Coli ◽

Nitrogen Fixation ◽

High Temperature ◽

Sigma Factor ◽

Host Plants ◽

Sinorhizobium Meliloti ◽

Free Living ◽

Genes Encoding ◽

Multiple Copies ◽

Free Living Conditions

ABSTRACT Among the rhizobia that establish nitrogen-fixing nodules on the roots of host plants, many contain multiple copies of genes encoding the sigma factor RpoH and the chaperone GroEL/GroES. In Sinorhizobium meliloti there are two rpoH genes, four groESL operons, and one groEL gene. rpoH1 mutants are defective for growth at high temperature and form ineffective nodules, rpoH1 rpoH2 double mutants are unable to form nodules, and groESL1 mutants form ineffective nodules. To explore the roles of RpoH1 and RpoH2, we identified mutants that suppress both the growth and nodulation defects. These mutants do not suppress the nitrogen fixation defect. This implies that the functions of RpoH1 during growth and RpoH1/RpoH2 during the initiation of symbiosis are similar but that there is a different function of RpoH1 needed later during symbiosis. We showed that, unlike in Escherichia coli, overexpression of groESL is not sufficient to bypass any of the RpoH defects. Under free-living conditions, we determined that RpoH2 does not control expression of the groE genes, and RpoH1 only controls expression of groESL5. Finally, we completed the series of groE mutants by constructing groESL3 and groEL4 mutants and demonstrated that they do not display symbiotic defects. Therefore, the only groESL operon required by itself for symbiosis is groESL1. Taken together, these results suggest that GroEL/GroES production alone cannot explain the requirements for RpoH1 and RpoH2 in S. meliloti and that there must be other crucial targets.

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The RpoH-Mediated Stress Response in Neisseria gonorrhoeae Is Regulated at the Level of Activity

Journal of Bacteriology ◽

10.1128/jb.186.24.8443-8452.2004 ◽

2004 ◽

Vol 186 (24) ◽

pp. 8443-8452 ◽

Cited By ~ 19

Author(s):

Lina Laskos ◽

Catherine S. Ryan ◽

Janet A. M. Fyfe ◽

John K. Davies

Keyword(s):

Stress Response ◽

Neisseria Gonorrhoeae ◽

Molecular Chaperones ◽

Sigma Factor ◽

Mutational Analysis ◽

Normal Growth ◽

Gene Encoding ◽

General Stress Response ◽

Level Of Activity ◽

Genes Encoding

ABSTRACT The general stress response in Neisseria gonorrhoeae was investigated. Transcriptional analyses of the genes encoding the molecular chaperones DnaK, DnaJ, and GrpE suggested that they are transcribed from σ32 (RpoH)-dependent promoters upon exposure to stress. This was confirmed by mutational analysis of the σ32 promoter of dnaK. The gene encoding the gonococcal RpoH sigma factor appears to be essential, as we could not isolate viable mutants. Deletion of an unusually long rpoH leader sequence resulted in elevated levels of transcription, suggesting that this region is involved in negative regulation of RpoH expression during normal growth. Transcriptional analyses and protein studies determined that regulation of the RpoH-mediated stress response is different from that observed in most other species, in which regulation occurs predominantly at the transcriptional and translational levels. We suggest that an increase in the activity of preformed RpoH is primarily responsible for induction of the stress response in N. gonorrhoeae.

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The Complex Flagellar Torque Generator of Pseudomonas aeruginosa

Journal of Bacteriology ◽

10.1128/jb.186.19.6341-6350.2004 ◽

2004 ◽

Vol 186 (19) ◽

pp. 6341-6350 ◽

Cited By ~ 104

Author(s):

Timothy B. Doyle ◽

Andrew C. Hawkins ◽

Linda L. McCarter

Keyword(s):

Pseudomonas Aeruginosa ◽

Membrane Proteins ◽

Motor Function ◽

Swimming Speed ◽

Flagellar Motor ◽

Loss Of Function ◽

Wild Type ◽

Aqueous Environments ◽

Genes Encoding ◽

Rotary Motors

ABSTRACT Flagella act as semirigid helical propellers that are powered by reversible rotary motors. Two membrane proteins, MotA and MotB, function as a complex that acts as the stator and generates the torque that drives rotation. The genome sequence of Pseudomonas aeruginosa PAO1 contains dual sets of motA and motB genes, PA1460-PA1461 (motAB) and PA4954-PA4953 (motCD), as well as another gene, motY (PA3526), which is known to be required for motor function in some bacteria. Here, we show that these five genes contribute to motility. Loss of function of either motAB-like locus was dispensable for translocation in aqueous environments. However, swimming could be entirely eliminated by introduction of combinations of mutations in the two motAB-encoding regions. Mutation of both genes encoding the MotA homologs or MotB homologs was sufficient to abolish motility. Mutants carrying double mutations in nonequivalent genes (i.e., motA motD or motB motC) retained motility, indicating that noncognate components can function together. motY appears to be required for motAB function. The combination of motY and motCD mutations rendered the cells nonmotile. Loss of function of motAB, motY, or motAB motY produced similar phenotypes; although the swimming speed was only reduced to ∼85% of the wild-type speed, translocation in semisolid motility agar and swarming on the surface of solidified agar were severely impeded. Thus, the flagellar motor of P. aeruginosa represents a more complex configuration than the configuration that has been studied in other bacteria, and it enables efficient movement under different circumstances.

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