Proteus mirabilis mutants defective in swarmer cell differentiation and multicellular behavior.

ABSTRACT In this study, we identified a transposon insertion in a novel gene, designated disA, that restored swarming motility to a putrescine-deficient speA mutant of Proteus mirabilis. A null allele in disA also increased swarming in a wild-type background. The DisA gene product was homologous to amino acid decarboxylases, and its role in regulating swarming was investigated by examining the expression of genes in the flagellar cascade. In a disA mutant background, we observed a 1.4-fold increase in the expression of flhDC, which encodes FlhD2C2, the master regulator of the flagellar gene cascade. However, the expressions of class 2 (fliA, flgM) and class 3 (flaA) genes were at least 16-fold higher in the disA background during swarmer cell differentiation. Overexpression of DisA on a high-copy-number plasmid did not significantly decrease flhDC mRNA accumulation but resulted in a complete block in mRNA accumulation for both fliA and flaA. DisA overexpression also blocked swarmer cell differentiation. The disA gene was regulated during the swarming cycle, and a single-copy disA::lacZ fusion exhibited a threefold increase in expression in swarmer cells. Given that DisA was similar to amino acid decarboxylases, a panel of decarboxylated amino acids was tested for effects similar to DisA overexpression, and phenethylamine, the product of phenylalanine decarboxylation, was capable of inhibiting both swarming and the expression of class 2 and class 3 genes in the flagellar regulon. A DisA-dependent decarboxylated amino acid may inhibit the formation of active FlhD2C2 heterotetramers or inhibit FlhD2C2 binding to DNA.

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Characterization of a Novel Gene, wosA, Regulating FlhDC Expression in Proteus mirabilis

Journal of Bacteriology ◽

10.1128/jb.01010-07 ◽

2008 ◽

Vol 190 (6) ◽

pp. 1946-1955 ◽

Cited By ~ 9

Author(s):

Janet K. Hatt ◽

Philip N. Rather

Keyword(s):

Cell Differentiation ◽

Liquid Medium ◽

Proteus Mirabilis ◽

Solid Surfaces ◽

Swarming Motility ◽

Swarmer Cell ◽

Fourfold Increase ◽

Nonpermissive Condition

ABSTRACT In this study, we describe wosA, a Proteus mirabilis gene identified by its ability to increase swarming motility when overexpressed. At various times during the swarming cycle, the increased expression of wosA resulted in a 4- to 16-fold upregulation of the transcription of flhDC, encoding the master regulator of the flagellar cascade. In turn, the expression of flaA, encoding flagellin, was substantially increased in wosA-overexpressing strains. The overexpression of wosA also resulted in constitutive swarmer cell differentiation in liquid medium, a normally nonpermissive condition. However, in wosA-overexpressing strains, the onset of swarming was not altered. A null wosA allele resulted in a slight decrease in swarming motility. The expression of wosA was growth phase dependent during growth in liquid and on agar plates during swarmer cell differentiation. Increasing the viscosity of liquid medium by the addition of polyvinylpyrrolidone induced swarmer cell differentiation and resulted in a fourfold increase in wosA transcription. A fliL mutation that results in constitutive swarmer cell elongation also increased wosA transcription. In this study, we discuss the possible role of the wosA gene product in signal transduction from solid surfaces to induce swarmer cell differentiation, possibly via alterations in the motor switch complex. This study also suggests that despite constitutive swarmer cell differentiation in wosA-overexpressing strains, there are additional regulatory and/or environmental conditions that may control the onset of swarming migration.

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Regulation of the Min Cell Division Inhibition Complex by the Rcs Phosphorelay in Proteus mirabilis

Journal of Bacteriology ◽

10.1128/jb.00094-15 ◽

2015 ◽

Vol 197 (15) ◽

pp. 2499-2507 ◽

Cited By ~ 18

Author(s):

Kristen E. Howery ◽

Katy M. Clemmer ◽

Emrah Şimşek ◽

Minsu Kim ◽

Philip N. Rather

Keyword(s):

Cell Division ◽

Cell Differentiation ◽

Proteus Mirabilis ◽

Cell Elongation ◽

Response Regulator ◽

Swarmer Cell ◽

Content Type ◽

Expression Of Genes ◽

Rapid Amplification

ABSTRACTA key regulator of swarming inProteus mirabilisis the Rcs phosphorelay, which repressesflhDC, encoding the master flagellar regulator FlhD4C2. Mutants inrcsB, the response regulator in the Rcs phosphorelay, hyperswarm on solid agar and differentiate into swarmer cells in liquid, demonstrating that this system also influences the expression of genes central to differentiation. To gain a further understanding of RcsB-regulated genes involved in swarmer cell differentiation, transcriptome sequencing (RNA-Seq) was used to examine the RcsB regulon. Among the 133 genes identified,minCandminD, encoding cell division inhibitors, were identified as RcsB-activated genes. A third gene,minE, was shown to be part of an operon withminCD. To examineminCDEregulation, theminpromoter was identified by 5′ rapid amplification of cDNA ends (5′-RACE), and both transcriptionallacZfusions and quantitative real-time reverse transcriptase (qRT) PCR were used to confirm that theminCDEoperon was RcsB activated. Purified RcsB was capable of directly binding theminCpromoter region. To determine the role of RcsB-mediated activation ofminCDEin swarmer cell differentiation, a polarminCmutation was constructed. This mutant formed minicells during growth in liquid, produced shortened swarmer cells during differentiation, and exhibited decreased swarming motility.IMPORTANCEThis work describes the regulation and role of the MinCDE cell division system inP. mirabilisswarming and swarmer cell elongation. Prior to this study, the mechanisms that inhibit cell division and allow swarmer cell elongation were unknown. In addition, this work outlines for the first time the RcsB regulon inP. mirabilis. Taken together, the data presented in this study begin to address howP. mirabiliselongates upon contact with a solid surface.

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The Ability of Proteus mirabilis To Sense Surfaces and Regulate Virulence Gene Expression Involves FliL, a Flagellar Basal Body Protein

Journal of Bacteriology ◽

10.1128/jb.187.19.6789-6803.2005 ◽

2005 ◽

Vol 187 (19) ◽

pp. 6789-6803 ◽

Cited By ~ 80

Author(s):

Robert Belas ◽

Rooge Suvanasuthi

Keyword(s):

Gene Expression ◽

Cell Differentiation ◽

Basal Body ◽

Proteus Mirabilis ◽

Virulence Gene ◽

Wild Type ◽

Swarmer Cell ◽

Virulence Gene Expression ◽

Flagellar Motors ◽

Flagellar Rotation

ABSTRACT Proteus mirabilis is a urinary tract pathogen that differentiates from a short swimmer cell to an elongated, highly flagellated swarmer cell. Swarmer cell differentiation parallels an increased expression of several virulence factors, suggesting that both processes are controlled by the same signal. The molecular nature of this signal is not known but is hypothesized to involve the inhibition of flagellar rotation. In this study, data are presented supporting the idea that conditions inhibiting flagellar rotation induce swarmer cell differentiation and implicating a rotating flagellar filament as critical to the sensing mechanism. Mutations in three genes, fliL, fliF, and fliG, encoding components of the flagellar basal body, result in the inappropriate development of swarmer cells in noninducing liquid media or hyperelongated swarmer cells on agar media. The fliL mutation was studied in detail. FliL− mutants are nonmotile and fail to synthesize flagellin, while complementation of fliL restores wild-type cell elongation but not motility. Overexpression of fliL + in wild-type cells prevents swarmer cell differentiation and motility, a result also observed when P. mirabilis fliL + was expressed in Escherichia coli. These results suggest that FliL plays a role in swarmer cell differentiation and implicate FliL as critical to transduction of the signal inducing swarmer cell differentiation and virulence gene expression. In concert with this idea, defects in fliL up-regulate the expression of two virulence genes, zapA and hpmB. These results support the hypothesis that P. mirabilis ascertains its location in the environment or host by assessing the status of its flagellar motors, which in turn control swarmer cell gene expression.

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Faculty Opinions recommendation of Ultrastructure of Proteus mirabilis swarmer cell rafts and role of swarming in catheter-associated urinary tract infection.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1020375.232550 ◽

2004 ◽

Author(s):

Charles Czuprynski

Keyword(s):

Urinary Tract Infection ◽

Urinary Tract ◽

Tract Infection ◽

Proteus Mirabilis ◽

Swarmer Cell

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Requirement for FlhA in flagella assembly and swarm-cell differentiation by Proteus mirabilis

Molecular Microbiology ◽

10.1111/j.1365-2958.1995.tb02383.x ◽

2006 ◽

Vol 15 (4) ◽

pp. 761-769 ◽

Cited By ~ 48

Author(s):

Daniel Gygi ◽

Marc J. Bailey ◽

Clive Allison ◽

Colin Hughes

Keyword(s):

Cell Differentiation ◽

Proteus Mirabilis ◽

Flagella Assembly

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Closely linked genetic loci required for swarm cell differentiation and multicellular migration by Proteus mirabilis

Molecular Microbiology ◽

10.1111/j.1365-2958.1991.tb00819.x ◽

1991 ◽

Vol 5 (8) ◽

pp. 1975-1982 ◽

Cited By ~ 32

Author(s):

C. Allison ◽

C. Hughes

Keyword(s):

Cell Differentiation ◽

Proteus Mirabilis ◽

Genetic Loci

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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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