The histone‐like nucleoid structuring protein H‐NS represses the <i>Escherichia coli bgl</i> operon downstream of the promoter

Dole, Sudhanshu; Nagarajavel, V.; Schnetz, Karin

doi:10.1111/j.1365-2958.2004.04001.x

Cited by 72 publications

(116 citation statements)

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“…2C and 2D, multiple bands of nucleoprotein complexes were obtained for the reaction mixtures with low concentrations of MvaT or MvaU, and a complete shift of F501 to a low-mobility complex was observed with higher concentrations of the proteins. These patterns of multinucleoprotein complex formation were similar to what has been reported previously for H-NS of E. coli (6,36).…”

Section: Resultssupporting

confidence: 76%

The Multifaceted Proteins MvaT and MvaU, Members of the H-NS Family, Control Arginine Metabolism, Pyocyanin Synthesis, and Prophage Activation in Pseudomonas aeruginosa PAO1

Wally

Miller

et al. 2009

J Bacteriol

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The MvaT and MvaU proteins belonging to the H-NS family were identified as DNA-binding proteins that interact with the regulatory region of the aotJQMOP-argR operon for arginine uptake and regulation. Recombinant MvaT and MvaU proteins were purified, and binding of these purified proteins to the aotJ regulatory region was demonstrated using electromobility shift assays. Polyclonal antibodies against purified MvaT and MvaU were prepared and employed in supershift assays to support these observations. Knockout mutations resulting in a single lesion in mvaT or mvaU, as well as knockout mutations resulting in double lesions, were constructed using biparental conjugation, and the absence of MvaT and MvaU in the resulting mutants was confirmed by immunoblot analysis. Using measurements of the ␤-galactosidase activities from aotJ::lacZ fusions in the mutants and the parental strain, it was found that MvaT and MvaU serve as repressors in control of aotJ expression. The effects of MvaT and MvaU on pyocyanin synthesis and CupA fimbrial expression in these mutants were also analyzed. Pyocyanin synthesis was induced in the single mutants but was completely abolished in the double mutant, suggesting that there is a complicated regulatory scheme in which MvaT and MvaU are essential elements. In comparison, MvaT had a more profound role than MvaU as a repressor of cupA expression; however, a combination of MvaT depletion and MvaU depletion had a strong synergistic effect on cupA. Moreover, prophage Pf4 integrated into the chromosome of Pseudomonas aeruginosa PAO1 was activated in an mvaT mvaU double mutant but not in a single mutant. These results were supported by purification and nucleotide sequencing of replicative-form DNA and by the release of phage particles in plaque assays. In summary, the mvaT mvaU double mutant was viable, and depletion of MvaT and MvaU had serious effects on a variety of physiological functions in P. aeruginosa.The MvaT and MvaU proteins of Pseudomonas aeruginosa belong to the H-NS family of small DNA-binding proteins. MvaT was initially identified in P. mevalonii as a positive regulator for mevalonate catabolism (25). Subsequently, MvaT homologues have been identified in other pseudomonads based on structural and functional similarities; five homologues have been identified in P. putida, three homologues have been identified in P. fluorescens, four homologues have been identified in P. syringae, and two homologues have been identified in P. aeruginosa. In P. putida, the TurA protein represses the Pu promoter of the TOL plasmid in a temperature-dependent manner (24). In P. fluorescens, the MvaT and MvaV proteins regulate the expression of two biocontrol exoproducts, 2,4-diacetyl phloroglucinol and pyoluteorin (1). In P. aeruginosa, MvaT is involved in quorum-sensing responses and biofilm formation. Inactivation of mvaT resulted in increased production of PA-IL lectin and the toxic exoproduct pyocyanin, reduced biofilm formation, increased drug resistance, and reduced swarming motility (5, 33). In ad...

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Section: Resultssupporting

confidence: 76%

The Multifaceted Proteins MvaT and MvaU, Members of the H-NS Family, Control Arginine Metabolism, Pyocyanin Synthesis, and Prophage Activation in Pseudomonas aeruginosa PAO1

Wally

Miller

et al. 2009

J Bacteriol

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“…The bgl operon from E. coli carries two silencers to which H-NS binds preferentially (Schnetz, 1995;Dole et al, 2004). One silencer is located upstream of the promoter and the other is located 600-700 bp downstream of the start site of transcription (Dole et al, 2004). The two silencers function independently in the repression of bgl expression.…”

Section: Discussionmentioning

confidence: 99%

“…Detailed analysis of the mechanism of repression of the proU promoter has shown that binding of H-NS to DREs inhibits open complex formation by RNA polymerase (Jordi & Higgins, 2000). The bgl operon from E. coli carries two silencers to which H-NS binds preferentially (Schnetz, 1995;Dole et al, 2004). One silencer is located upstream of the promoter and the other is located 600-700 bp downstream of the start site of transcription (Dole et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

The H-NS protein represses transcription of the eltAB operon, which encodes heat-labile enterotoxin in enterotoxigenic Escherichia coli, by binding to regions downstream of the promoter

et al. 2005

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Heat-labile enterotoxin, a major virulence determinant of enterotoxigenic Escherichia coli, is encoded by the eltAB operon. To elucidate the molecular mechanism by which the heat-stable nucleoid-structural (H-NS) protein controls transcription of eltAB, the authors constructed an eltAB-lacZ transcriptional fusion and performed b-galactosidase analysis. The results showed that H-NS protein exerts fivefold repression on transcription from the eltAB promoter at 37 6C and 10-fold repression at 22 6C. Two silencer regions that were required for H-NS-mediated repression of eltAB expression were identified, both of which were located downstream of the start site of transcription. One silencer was located between +31 and +110, the other between +460 and +556, relative to the start site of transcription, and they worked cooperatively in repression. DNA sequences containing the silencers were predicted to be curved by in silico analysis and bound H-NS protein directly in vitro. Repression of eltAB transcription by H-NS was independent of promoter strength, and the presence of H-NS protein did not affect promoter opening in vitro, indicating that repression was achieved by inhibiting promoter clearance or blocking transcription elongation, probably via DNA looping between the two silencers.

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“…The mechanism by which H-NS influences RNA polymerase activity is not fully understood. Its function as a transcription factor is, however, tightly connected with its ability to oligomerize [Rimsky et al, 2001] and condense DNA into stable nucleoprotein complexes [Schroder and Wagner, 2000;Dole et al, 2004;Prosseda et al, 2004], thereby occluding promoter regions or blocking transcription elongation. Recently, atomic force microscopy was used to visualize the effects of H-NS on DNA topology [Dame et al, 2000]: after initial binding at dispersed sites, H-NS assembles into patches of oligomers that interact with each other, thus bridging different DNA regions.…”

Section: Mechansims Of Nucleoid Organizationmentioning

confidence: 99%

The bacterial nucleoid: A highly organized and dynamic structure

Thanbichler

Wang

Shapiro

2005

J of Cellular Biochemistry

122

101

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Recent advances in bacterial cell biology have revealed unanticipated structural and functional complexity, reminiscent of eukaryotic cells. Particular progress has been made in understanding the structure, replication, and segregation of the bacterial chromosome. It emerged that multiple mechanisms cooperate to establish a dynamic assembly of supercoiled domains, which are stacked in consecutive order to adopt a defined higher-level organization. The position of genetic loci on the chromosome is thereby linearly correlated with their position in the cell. SMC complexes and histone-like proteins continuously remodel the nucleoid to reconcile chromatin compaction with DNA replication and gene regulation. Moreover, active transport processes ensure the efficient segregation of sister chromosomes and the faithful restoration of nucleoid organization while DNA replication and condensation are in progress.

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The histone‐like nucleoid structuring protein H‐NS represses the Escherichia coli bgl operon downstream of the promoter

Cited by 72 publications

References 48 publications

The Multifaceted Proteins MvaT and MvaU, Members of the H-NS Family, Control Arginine Metabolism, Pyocyanin Synthesis, and Prophage Activation in Pseudomonas aeruginosa PAO1

The Multifaceted Proteins MvaT and MvaU, Members of the H-NS Family, Control Arginine Metabolism, Pyocyanin Synthesis, and Prophage Activation in Pseudomonas aeruginosa PAO1

The H-NS protein represses transcription of the eltAB operon, which encodes heat-labile enterotoxin in enterotoxigenic Escherichia coli, by binding to regions downstream of the promoter

The bacterial nucleoid: A highly organized and dynamic structure

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