scholarly journals Roles of rpoN, fliA,and flgR in Expression of Flagella inCampylobacter jejuni

2001 ◽  
Vol 183 (9) ◽  
pp. 2937-2942 ◽  
Author(s):  
Aparna Jagannathan ◽  
Chrystala Constantinidou ◽  
Charles W. Penn
Keyword(s):  
Campylobacter Jejuni ◽  
Genome Sequence ◽  
Sigma Factor ◽  
Transcriptional Activator ◽  
Alternative Sigma Factor ◽  
Wild Type ◽  
Electron Microscopic ◽  
Global Regulation ◽  
Mutant Strains ◽  
Microscopic Studies

ABSTRACT Three potential regulators of flagellar expression present in the genome sequence of Campylobacter jejuni NCTC 11168, the genes rpoN, flgR, andfliA, which encode the alternative sigma factor ς54, the ς54-associated transcriptional activator FlgR, and the flagellar sigma factor ς28, respectively, were investigated for their role in global regulation of flagellar expression. The three genes were insertionally inactivated inC. jejuni strains NCTC 11168 and NCTC 11828. Electron microscopic studies of the wild-type and mutant strains showed that therpoN and flgR mutants were nonflagellate and that the fliA mutant had truncated flagella. Immunoblotting experiments with the three mutants confirmed the roles of rpoN, flgR, and fliA in the expression of flagellin.

scholarly journals Enhancing Isoprene Production by Genetic Modification of the 1-Deoxy-d-Xylulose-5-Phosphate Pathway inBacillus subtilis

2011 ◽  
Vol 77 (7) ◽  
pp. 2399-2405 ◽  
Author(s):  
Junfeng Xue ◽  
Birgitte K. Ahring
Keyword(s):  
Sigma Factor ◽  
Type Strain ◽  
Wild Type Strain ◽  
Stress Factors ◽  
Alternative Sigma Factor ◽  
Wild Type ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Overexpression Strain ◽  
As Stress

ABSTRACTTo enhance the production of isoprene, a volatile 5-carbon hydrocarbon, in the Gram-positive spore-forming rod-shaped bacteriumBacillus subtilis, 1-deoxy-d-xylulose-5-phosphate synthase (Dxs) and 1-deoxy-d-xylulose-5-phosphate reductoisomerase (Dxr) were overexpressed inB. subtilisDSM 10. For the strain that overexpresses Dxs, the yield of isoprene was increased 40% over that by the wild-type strain. In the Dxr overexpression strain, the level of isoprene production was unchanged. Overexpression of Dxr together with Dxs showed an isoprene production level similar to that of the Dxs overproduction strain. The effects of external factors, such as stress factors including heat (48°C), salt (0.3 M NaCl), ethanol (1%), and oxidative (0.005% H2O2) stress, on isoprene production were further examined. Heat, salt, and H2O2induced isoprene production; ethanol inhibited isoprene production. In addition, induction and repression effects are independent of SigB, which is the general stress-responsive alternative sigma factor of Gram-positive bacteria.

scholarly journals Mutagenic strategies against luxS gene affect the early stage of biofilm formation of Campylobacter jejuni

2021 ◽  
Author(s):  
Martin Tereň ◽  
Ekaterina Shagieva ◽  
Lucie Vondrakova ◽  
Jitka Viktorova ◽  
Viviana Svarcova ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Biofilm Formation ◽  
Early Stage ◽  
Wild Type ◽  
Genetic Determinants ◽  
Autoinducer 2 ◽  
Naturally Occurring ◽  
Mutant Strains ◽  
The Impact

Abstract Currently, it is clear that the luxS gene has an impact on the process of biofilm formation in Campylobacter jejuni. However, even within the species naturally occurring strains of Campylobacter lacking the luxS gene exist, which can form biofilms. In order to better understand the genetic determinants and the role of quorum sensing through the LuxS/AI-2 pathway in biofilm formation, a set of mutant/complemented strains of C. jejuni 81–176 were prepared. Additionally, the impact of the mutagenic strategy used against the luxS gene was investigated. Biofilm formation was affected by both the presence and absence of the luxS gene, and by the mutagenic strategy used. Analysis by CLSM showed that all mutant strains formed significantly less biofilm mass when compared to the wild-type. Interestingly, the deletion mutant (∆luxS) showed a larger decrease in biofilm mass than the substitution (∙luxS) and insertional inactivated (⸬luxS) mutants, even though all the mutant strains lost the ability to produce autoinducer-2 molecules. Moreover, the biofilm of the ∆luxS mutant lacked the characteristic microcolonies observed in all other strains. The complementation of all mutant strains resulted in restored ability to produce AI-2, to form a complex biofilm, and to develop microcolonies at the level of the wild-type.

scholarly journals Regulation of the Bacillus subtilis Extracytoplasmic Function Protein σY and Its Target Promoters

2003 ◽  
Vol 185 (16) ◽  
pp. 4883-4890 ◽  
Author(s):  
Min Cao ◽  
Letal Salzberg ◽  
Ching Sung Tsai ◽  
Thorsten Mascher ◽  
Carla Bonilla ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Sigma Factor ◽  
Negative Regulation ◽  
Wild Type ◽  
Induced Mutations ◽  
Mutant Strains ◽  
Elevated Expression ◽  
Direct Target ◽  
Extracytoplasmic Function Sigma Factor ◽  
Target Promoters

ABSTRACT The Bacillus subtilis extracytoplasmic function sigma factor σY is of unknown function. We demonstrate that the sigY operon is expressed from an autoregulatory promoter site, PY. We selected for transposon-induced mutations that upregulate PY transcription in an attempt to identify genes involved in σY regulation. The resulting insertions disrupted yxlC, the gene immediately downstream of sigY. However, the phenotype of the yxlC::Tn10 insertion was due to polarity on the downstream genes of the sigY operon; a nonpolar insertion in yxlC did not lead to derepression of PY. Further analyses revealed that both yxlD and yxlE encoded proteins important for the negative regulation of σY activity. A comparison of the transcriptomes of wild-type and yxlC::Tn10 mutant strains revealed elevated expression of several operons. However, only one additional gene, ybgB, was unambiguously identified as a direct target for σY. This was supported by analysis of direct targets for σY transcription with whole-genome runoff transcription followed by macroarray analysis.

scholarly journals RpoS-Regulated Genes of Escherichia coli Identified by Random lacZ Fusion Mutagenesis

2004 ◽  
Vol 186 (24) ◽  
pp. 8499-8507 ◽  
Author(s):  
Somalinga R. V. Vijayakumar ◽  
Mark G. Kirchhof ◽  
Cheryl L. Patten ◽  
Herb E. Schellhorn
Keyword(s):  
Escherichia Coli ◽  
Sigma Factor ◽  
Genetic Screen ◽  
The Other ◽  
Alternative Sigma Factor ◽  
Gene Fusions ◽  
Wild Type ◽  
Stress Response Genes ◽  
Independent Gene ◽  
Bacterial Genes

ABSTRACT RpoS is a conserved alternative sigma factor that regulates the expression of many stress response genes in Escherichia coli. The RpoS regulon is large but has not yet been completely characterized. In this study, we report the identification of over 100 RpoS-dependent fusions in a genetic screen based on the differential expression of an operon-lacZ fusion bank in rpoS mutant and wild-type backgrounds. Forty-eight independent gene fusions were identified, including several in well-characterized RpoS-regulated genes, such as osmY, katE, and otsA. Many of the other fusions mapped to genes of unknown function or to genes that were not previously known to be under RpoS control. Based on the homology to other known bacterial genes, some of the RpoS-regulated genes of unknown functions are likely important in nutrient scavenging.

scholarly journals Haemophilus influenzaeMicroarrays: Virulence and Vaccines

2002 ◽  
Vol 3 (4) ◽  
pp. 358-361
Author(s):  
Tahir R. Ali ◽  
J. Simon Kroll ◽  
Paul R. Langford
Keyword(s):  
Genome Sequence ◽  
Regulatory Networks ◽  
Living Organism ◽  
Dna Arrays ◽  
Wild Type ◽  
Free Living ◽  
Mutant Strains ◽  
Genome Array ◽  
Global Strategies

In 1995 the genome sequence of theHaemophilus influenzaeKW20 (Rd) strain was published, the first available for a free-living organism. The genome has been invaluable in global strategies to identify certain virulence-related genes, e.g. those involved in LPS synthesis, and also essential genes, but there is a paucity of wholegenome transcriptome studies. We have now constructed a whole-genome array consisting of genes from Rd, additional genes identified in other strains ofH. influenzaeand controls (from eukaryotic sources and other bacteria). We intend to use this array in studies aimed at understanding the bacterium’s basic metabolism and its response to changing environments; deciphering global regulatory networks (by comparison of wild-type and mutant strains); and identifying genes expressedin vivo. The use ofH. influenzaeDNA arrays combined with proteomic approaches will enhance our understanding of the metabolism and virulence of the organism. Additionally, the genome sequence of a non-typableH. influenzaestrain is in progress. The sequence from this isolate will be invaluable not only in identifying potential novel antibiotic targets and putative vaccine candidates but also in the design of a microarray for genome-typing purposes.

scholarly journals fleN, a Gene That Regulates Flagellar Number in Pseudomonas aeruginosa

2000 ◽  
Vol 182 (2) ◽  
pp. 357-364 ◽  
Author(s):  
Nandini Dasgupta ◽  
Shiwani K. Arora ◽  
Reuben Ramphal
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Binding Motif ◽  
Gene Promoters ◽  
Wild Type ◽  
Electron Microscopic ◽  
Genome Database ◽  
Reading Frame ◽  
Mutant Strains ◽  
In The Wild ◽  
Flagellar Genes

ABSTRACT The single polar flagellum of Pseudomonas aeruginosaplays an important role in the pathogenesis of infection by this organism. However, regulation of the assembly of this organelle has not been delineated. In analyzing the sequence available at thePseudomonas genome database, an open reading frame (ORF), flanked by flagellar genes flhF and fliA, that coded for a protein (280 amino acids) with an ATP-binding motif at its N terminus was found. The ORF was inactivated by inserting a gentamicin cassette in P. aeruginosa PAK and PAO1. The resulting mutants were nonmotile on motility agar plates, but under a light microscope they exhibited random movement and tumbling behavior. Electron microscopic studies of the wild-type and mutant strains revealed that the mutants were multiflagellate, with three to six polar flagella per bacterium as rather than one as in the wild type, indicating that this ORF was involved in regulating the number of flagella and chemotactic motility in P. aeruginosa. The ORF was named fleN. An intact copy of fleN on a plasmid complemented the mutant by restoring motility and monoflagellate status. The β-galactosidase activities of eight flagellar operon or gene promoters in the wild-type andfleN mutant strains revealed a direct correlation between six promoters that were upregulated in the fleN mutant (fliLMNOPQ, flgBCDE, fliEFG,fliDS orf126, fleSR, and fliC) and positive regulation by FleQ, an NtrC-like transcriptional regulator for flagellar genes. Based on these results, we propose a model where FleN influences FleQ activity (directly or indirectly) in regulating flagellar number in P. aeruginosa.

scholarly journals RpoN Modulates Carbapenem Tolerance in Pseudomonas aeruginosa through Pseudomonas Quinolone Signal and PqsE

2016 ◽  
Vol 60 (10) ◽  
pp. 5752-5764 ◽  
Author(s):  
Darija Viducic ◽  
Keiji Murakami ◽  
Takashi Amoh ◽  
Tsuneko Ono ◽  
Yoichiro Miyake
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Sigma Factor ◽  
Alternative Sigma Factor ◽  
Regulatory Effect ◽  
Rich Medium ◽  
Wild Type ◽  
Transcript Levels ◽  
Content Type ◽  
Pseudomonas Quinolone Signal ◽  
Nutrient Rich Medium

ABSTRACTThe ability ofPseudomonas aeruginosato rapidly modulate its response to antibiotic stress and persist in the presence of antibiotics is closely associated with the process of cell-to-cell signaling. The alternative sigma factor RpoN (σ54) is involved in the regulation of quorum sensing (QS) and plays an important role in the survival of stationary-phase cells in the presence of carbapenems. Here, we demonstrate that a ΔrpoNmutant grown in nutrient-rich medium has increased expression ofpqsA,pqsH, andpqsRthroughout growth, resulting in the increased production of thePseudomonasquinolone signal (PQS). The link betweenpqsAand its role in carbapenem tolerance was studied using a ΔrpoNΔpqsAmutant, in which the carbapenem-tolerant phenotype of the ΔrpoNmutant was abolished. In addition, we demonstrate that another mechanism leading to carbapenem tolerance in the ΔrpoNmutant is mediated throughpqsE. Exogenously supplied PQS abolished the biapenem-sensitive phenotype of the ΔrpoNΔpqsAmutant, and overexpression ofpqsEfailed to alter the susceptibility of the ΔrpoNΔpqsAmutant to biapenem. The mutations in the ΔrpoNΔrhlRmutant and the ΔrpoNΔpqsHmutant led to susceptibility to biapenem. Comparison of the changes in the expression of the genes involved in QS in wild-type PAO1 with their expression in the ΔrpoNmutant and the ΔrpoNmutant-derived strains demonstrated the regulatory effect of RpoN on the transcript levels ofrhlR,vqsR, andrpoS. The findings of this study demonstrate that RpoN negatively regulates the expression of PQS in nutrient-rich medium and provide evidence that RpoN interacts withpqsA,pqsE,pqsH, andrhlRin response to antibiotic stress.

scholarly journals Effects of Growth Phase and Temperature onσBActivity within aListeria monocytogenesPopulation: Evidence for RsbV-Independent Activation ofσBat Refrigeration Temperatures

2014 ◽  
Vol 2014 ◽  
pp. 1-11 ◽  
Author(s):  
Marta Utratna ◽  
Eoin Cosgrave ◽  
Claas Baustian ◽  
Rhodri H. Ceredig ◽  
Conor P. O’Byrne
Keyword(s):  
Growth Phase ◽  
Sigma Factor ◽  
Current Model ◽  
Exponential Phase ◽  
Alternative Sigma Factor ◽  
Alternative Route ◽  
Activation Pattern ◽  
Wild Type ◽  
Early Stationary Phase ◽  
Early Exponential Phase

The alternative sigma factorσBofListeria monocytogenesis responsible for regulating the transcription of many of the genes necessary for adaptation to both food-related stresses and to conditions found within the gastrointestinal tract of the host. The present study sought to investigate the influence of growth phase and temperature on the activation ofσBwithin populations ofL.monocytogenesEGD-e wild-type, ΔsigB,and ΔrsbVthroughout growth at both 4°C and 37°C, using a reporter fusion that couples expression of EGFP to the stronglyσB-dependent promoter oflmo2230. A similarσBactivation pattern within the population was observed in wt-egfpat both temperatures, with the highest induction ofσBoccurring in the early exponential phase of growth when the fluorescent population rapidly increased, eventually reaching the maximum in early stationary phase. Interestingly, induction ofσBactivity was heterogeneous, with only a proportion of the cells in the wt-egfppopulation being fluorescent above the background autofluorescence level. Moreover, significant RsbV-independent activation ofσBwas observed during growth at 4°C. This result suggests that an alternative route toσBactivation exists in the absence of RsbV, a finding that is not explained by the current model forσBregulation.

scholarly journals ClpC acts as a negative regulator of competence in Streptococcus thermophilus

Microbiology ◽  
2011 ◽  
Vol 157 (6) ◽  
pp. 1676-1684 ◽  
Author(s):  
Truls Johan Biørnstad ◽  
Leiv Sigve Håvarstein
Keyword(s):  
Sigma Factor ◽  
Wild Type Strain ◽  
Natural Transformation ◽  
Streptococcus Thermophilus ◽  
Negative Regulator ◽  
Wild Type ◽  
Environmental Stimulus ◽  
Mutant Strains ◽  
Low Levels

The alternative sigma factor ComX is a key regulator of natural transformation in members of the genus Streptococcus. ComX controls expression of the late competence genes, which are essential for DNA binding, uptake and recombination. In Streptococcus pneumoniae, it has been demonstrated that ComX is degraded by ClpEP at the end of the competence period. In the present study we show that a different Clp protease complex, ClpCP, contributes to ComX degradation in Streptococcus thermophilus. Mutant strains lacking the ClpC chaperone displayed significantly increased transformability compared with the wild-type strain under conditions where ComX was expressed at relatively low levels. At higher expression levels, ClpCP appears to become saturated and unable to prevent the accumulation of ComX. Together, our results suggest that the role of ClpC is to mediate degradation of ComX when the sigma factor is produced in low amounts, i.e. when the environmental stimulus promoting competence development is weak. This would prevent S. thermophilus from developing the competent state at an inappropriate time and/or place.

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