scholarly journals Expression of fnr Is Constrained by an Upstream IS5 Insertion in Certain Escherichia coli K-12 Strains

2005 ◽  
Vol 187 (8) ◽  
pp. 2609-2617 ◽  
Author(s):  
R. Gary Sawers
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Transcription Initiation ◽  
Regulatory Region ◽  
Initiation Site ◽  
Strain Mg1655 ◽  
Transcript Levels ◽  
E Coli ◽  
K 12 ◽  
Strain Mc4100

ABSTRACT FNR is a global transcriptional regulator that controls anaerobic gene expression in Escherichia coli. Through the use of a number of approaches it was shown that fnr gene expression is reduced approximately three- to fourfold in E. coli strain MC4100 compared with the results seen with strain MG1655. This reduction in fnr expression is due to the insertion of IS5 (is5F) in the regulatory region of the gene at position −41 relative to the transcription initiation site. Transcription of the fnr gene nevertheless occurs from its own promoter in strain MC4100, but transcript levels are reduced approximately fourfold compared with those seen with strain MG1655. Remarkably, in strains bearing is5F the presence of Hfq prevents IS5-dependent transcriptional silencing of fnr expression. Thus, an hfq mutant of MC4100 is devoid of FNR protein and has the phenotype of an fnr mutant. In strain MG1655, or a derivative of MC4100 lacking is5F, mutation of hfq had no effect on fnr transcript levels. This finding indicates that IS5 mediates the effect of Hfq on fnr expression in MC4100. Western blot analysis revealed that cellular levels of FNR were reduced threefold in strain MC4100 compared with strain MG1655 results. A selection of FNR-dependent genes fused to lacZ were analyzed for the effects of reduced FNR levels on anaerobic gene expression. Expression of some operons, e.g., focA-pfl and fdnGHJI, was unaffected by reduction in the level of FNR, while the expression of other genes such as ndh and nikA was clearly affected.

scholarly journals The Escherichia coli K-12 NarL and NarP Proteins Insulate the nrf Promoter from the Effects of Integration Host Factor

2006 ◽  
Vol 188 (21) ◽  
pp. 7449-7456 ◽  
Author(s):  
Douglas F. Browning ◽  
David J. Lee ◽  
Alan J. Wolfe ◽  
Jeffrey A. Cole ◽  
Stephen J. W. Busby
Keyword(s):  
Escherichia Coli ◽  
Response Regulator ◽  
Regulatory Region ◽  
Enteric Bacteria ◽  
Integration Host Factor ◽  
Host Factor ◽  
Receptor Protein ◽  
E Coli ◽  
Serovar Typhimurium ◽  
K 12

ABSTRACT The Escherichia coli K-12 nrf operon promoter can be activated fully by the FNR protein (regulator of fumarate and nitrate reduction) binding to a site centered at position −41.5. FNR-dependent transcription is suppressed by integration host factor (IHF) binding at position −54, and this suppression is counteracted by binding of the NarL or NarP response regulator at position −74.5. The E. coli acs gene is transcribed from a divergent promoter upstream from the nrf operon promoter. Transcription from the major acsP2 promoter is dependent on the cyclic AMP receptor protein and is modulated by IHF and Fis binding at multiple sites. We show that IHF binding to one of these sites, located at position −127 with respect to the nrf promoter, has a positive effect on nrf promoter activity. This activation is dependent on the face of the DNA helix, independent of IHF binding at other locations, and found only when NarL/NarP are not bound at position −74.5. Binding of NarL/NarP appears to insulate the nrf promoter from the effects of IHF. The acs-nrf regulatory region is conserved in other pathogenic E. coli strains and related enteric bacteria but differs in Salmonella enterica serovar Typhimurium.

scholarly journals Regulation of P-Fimbrial Phase Variation Frequencies in Escherichia coli CFT073

2007 ◽  
Vol 75 (7) ◽  
pp. 3325-3334 ◽  
Author(s):  
Nicola Holden ◽  
Makrina Totsika ◽  
Lynn Dixon ◽  
Kirsteen Catherwood ◽  
David L. Gally
Keyword(s):  
Escherichia Coli ◽  
Phase Transition ◽  
Regulatory Region ◽  
Phase Variation ◽  
Culture Conditions ◽  
Host Tissue ◽  
Clinical Isolates ◽  
E Coli ◽  
K 12 ◽  
First Time

ABSTRACT Adherence of uropathogenic Escherichia coli to host tissue is required for infection and is mediated by fimbriae, such as pyelonephritis-associated pili (Pap). Expression of P fimbriae is regulated by phase variation, and to date, phase transition frequencies have been measured only for pap regulatory region constructs integrated into the E. coli K-12 chromosome. The aim of this work was to measure P phase transition frequencies in clinical isolates for the first time, including frequencies for the sequenced strain E. coli CFT073. P fimbriation and associated phase transition frequencies were measured for two E. coli clinical isolates and compared with levels for homologous pap constructs in E. coli K-12. Fimbriation and off-to-on transition frequencies were always higher in the clinical isolate. It was concluded that the regulatory inputs controlling papI expression are likely to be different in E. coli CFT073 and E. coli K-12 as (i) phase variation could be stimulated in E. coli K-12 by induction of papI and (ii) the level of expression of a papI::gfp + fusion was higher in E. coli CFT073 than in E. coli K-12. Furthermore, phase transition frequencies for the two E. coli CFT073 pap clusters were shown to be different depending on the culture conditions, indicating that there is a hierarchy of expression depending on signal inputs.

scholarly journals Affinity Isolation and I-DIRT Mass Spectrometric Analysis of the Escherichia coli O157:H7 Sakai RNA Polymerase Complex

2007 ◽  
Vol 190 (4) ◽  
pp. 1284-1289 ◽  
Author(s):  
David J. Lee ◽  
Stephen J. W. Busby ◽  
Lars F. Westblade ◽  
Brian T. Chait
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Transcription Initiation ◽  
Laboratory Strain ◽  
Effector Proteins ◽  
Spectrometric Analysis ◽  
Isolation Technique ◽  
E Coli ◽  
Affinity Isolation ◽  
K 12

ABSTRACT Bacteria contain a single multisubunit RNA polymerase that is responsible for the synthesis of all RNA. Previous studies of the Escherichia coli K-12 laboratory strain identified a group of effector proteins that interact directly with RNA polymerase to modulate the efficiency of transcription initiation, elongation, or termination. Here we used a rapid affinity isolation technique to isolate RNA polymerase from the pathogenic Escherichia coli strain O157:H7 Sakai. We analyzed the RNA polymerase enzyme complex using mass spectrometry and identified associated proteins. Although E. coli O157:H7 Sakai contains more than 1,600 genes not present in the K-12 strain, many of which are predicted to be involved in transcription regulation, all of the identified proteins in this study were encoded on the “core” E. coli genome.

scholarly journals The Transcriptional Antiterminator RfaH Represses Biofilm Formation in Escherichia coli

2006 ◽  
Vol 188 (4) ◽  
pp. 1316-1331 ◽  
Author(s):  
Christophe Beloin ◽  
Kai Michaelis ◽  
Karin Lindner ◽  
Paolo Landini ◽  
Jörg Hacker ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Wild Type Strain ◽  
Wild Type ◽  
Strain Mg1655 ◽  
Upec Strain ◽  
E Coli ◽  
Initial Adhesion ◽  
State Production ◽  
K 12

ABSTRACT We investigated the influence of regulatory and pathogenicity island-associated factors (Hha, RpoS, LuxS, EvgA, RfaH, and tRNA5 Leu) on biofilm formation by uropathogenic Escherichia coli (UPEC) strain 536. Only inactivation of rfaH, which encodes a transcriptional antiterminator, resulted in increased initial adhesion and biofilm formation by E. coli 536. rfaH inactivation in nonpathogenic E. coli K-12 isolate MG1655 resulted in the same phenotype. Transcriptome analysis of wild-type strain 536 and an rfaH mutant of this strain revealed that deletion of rfaH correlated with increased expression of flu orthologs. flu encodes antigen 43 (Ag43), which mediates autoaggregation and biofilm formation. We confirmed that deletion of rfaH leads to increased levels of flu and flu-like transcripts in E. coli K-12 and UPEC. Supporting the hypothesis that RfaH represses biofilm formation through reduction of the Ag43 level, the increased-biofilm phenotype of E. coli MG1655rfaH was reversed upon inactivation of flu. Deletion of the two flu orthologs, however, did not modify the behavior of mutant 536rfaH. Our results demonstrate that the strong initial adhesion and biofilm formation capacities of strain MG1655rfaH are mediated by both increased steady-state production of Ag43 and likely increased Ag43 presentation due to null rfaH-dependent lipopolysaccharide depletion. Although the roles of rfaH in the biofilm phenotype are different in UPEC strain 536 and K-12 strain MG1655, this study shows that RfaH, in addition to affecting the expression of bacterial virulence factors, also negatively controls expression and surface presentation of Ag43 and possibly another Ag43-independent factor(s) that mediates cell-cell interactions and biofilm formation.

scholarly journals IraL Is an RssB Anti-adaptor That Stabilizes RpoS during Logarithmic Phase Growth in Escherichia coli and Shigella

mBio ◽  
2014 ◽  
Vol 5 (3) ◽  
Author(s):  
Andrew J. Hryckowian ◽  
Aurelia Battesti ◽  
Justin J. Lemke ◽  
Zachary C. Meyer ◽  
Rodney A. Welch
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Stress Response ◽  
Sigma Factor ◽  
Logarithmic Phase ◽  
Phase Growth ◽  
Content Type ◽  
General Stress Response ◽  
E Coli ◽  
K 12

ABSTRACTRpoS (σS), the general stress response sigma factor, directs the expression of genes under a variety of stressful conditions. Control of the cellular σSconcentration is critical for appropriately scaled σS-dependent gene expression. One way to maintain appropriate levels of σSis to regulate its stability. Indeed, σSdegradation is catalyzed by the ClpXP protease and the recognition of σSby ClpXP depends on the adaptor protein RssB. Three anti-adaptors (IraD, IraM, and IraP) exist inEscherichia coliK-12; each interacts with RssB andinhibitsRssBactivity under different stress conditions, thereby stabilizing σS. Unlike K-12, someE. coliisolates, including uropathogenicE. colistrain CFT073, show comparable cellular levels of σSduring the logarithmic and stationary growth phases, suggesting that there are differences in the regulation of σSlevels amongE. colistrains. Here, we describe IraL, an RssB anti-adaptor that stabilizes σSduring logarithmic phase growth in CFT073 and otherE. coliandShigellastrains. By immunoblot analyses, we show that IraL affects the levels and stability of σSduring logarithmic phase growth. By computational and PCR-based analyses, we reveal thatiraLis found in manyE. colipathotypes but not in laboratory-adapted strains. Finally, by bacterial two-hybrid and copurification analyses, we demonstrate that IraL interacts with RssB by a mechanism distinct from that used by other characterized anti-adaptors. We introduce a fourth RssB anti-adaptor found inE. colispecies and suggest that differences in the regulation of σSlevels may contribute to host and niche specificity in pathogenic and nonpathogenicE. colistrains.IMPORTANCEBacteria must cope with a variety of environmental conditions in order to survive. RpoS (σS), the general stress response sigma factor, directs the expression of many genes under stressful conditions in both pathogenic and nonpathogenicEscherichia colistrains. The regulation of σSlevels and activity allows appropriately scaled σS-dependent gene expression. Here, we describe IraL, an RssB anti-adaptor that, unlike previously described anti-adaptors, stabilizes σSduring the logarithmic growth phase in the absence of additional stress. We also demonstrate thatiraLis found in a large number ofE. coliandShigellaisolates. These data suggest that strains containingiraLare able to initiate σS-dependent gene expression under conditions under which strains withoutiraLcannot. Therefore, IraL-mediated σSstabilization may contribute to host and niche specificity inE. coli.

scholarly journals Adaptation of Sucrose Metabolism in the Escherichia coli Wild-Type Strain EC3132†

2002 ◽  
Vol 184 (19) ◽  
pp. 5307-5316 ◽  
Author(s):  
Knut Jahreis ◽  
Lars Bentler ◽  
Jürgen Bockmann ◽  
Stephan Hans ◽  
Astrid Meyer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Hot Spot ◽  
Chromosomal Rearrangements ◽  
Enteric Bacteria ◽  
Transcriptional Level ◽  
Wild Type ◽  
New Host ◽  
E Coli ◽  
K 12

ABSTRACT Although Escherichia coli strain EC3132 possesses a chromosomally encoded sucrose metabolic pathway, its growth on low sucrose concentrations (5 mM) is unusually slow, with a doubling time of 20 h. In this report we describe the subcloning and further characterization of the corresponding csc genes and adjacent genes. The csc regulon comprises three genes for a sucrose permease, a fructokinase, and a sucrose hydrolase (genes cscB, cscK, and cscA, respectively). The genes are arranged in two operons and are negatively controlled at the transcriptional level by the repressor CscR. Furthermore, csc gene expression was found to be cyclic AMP-CrpA dependent. A comparison of the genomic sequences of the E. coli strains EC3132, K-12, and O157:H7 in addition to Salmonella enterica serovar Typhimurium LT2 revealed that the csc genes are located in a hot spot region for chromosomal rearrangements in enteric bacteria. The comparison further indicated that the csc genes might have been transferred relatively recently to the E. coli wild-type EC3132 at around the time when the different strains of the enteric bacteria diverged. We found evidence that a mobile genetic element, which used the gene argW for site-specific integration into the chromosome, was probably involved in this horizontal gene transfer and that the csc genes are still in the process of optimal adaptation to the new host. Selection for such adaptational mutants growing faster on low sucrose concentrations gave three different classes of mutants. One class comprised cscR(Con) mutations that expressed all csc genes constitutively. The second class constituted a cscKo operator mutation, which became inducible for csc gene expression at low sucrose concentrations. The third class was found to be a mutation in the sucrose permease that caused an increase in transport activity.

scholarly journals Dual Function of Aar, a Member of the New AraC Negative Regulator Family, in Escherichia coli Gene Expression

2020 ◽  
Vol 88 (6) ◽  
Author(s):  
Abigail S. Mickey ◽  
James P. Nataro
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Negative Regulator ◽  
Dual Function ◽  
Content Type ◽  
Virulence Gene Expression ◽  
E Coli ◽  
K 12

ABSTRACT Enteroaggregative Escherichia coli (EAEC) is an E. coli pathotype associated with diarrhea and growth faltering. EAEC virulence gene expression is controlled by the autoactivated AraC family transcriptional regulator, AggR. AggR activates transcription of a large number of virulence genes, including Aar, which in turn acts as a negative regulator of AggR itself. Aar has also been shown to affect expression of E. coli housekeeping genes, including H-NS, a global regulator that acts at multiple promoters and silences AT-rich genes (such as those in the AggR regulon). Although Aar has been shown to bind both AggR and H-NS in vitro, functional significance of these interactions has not been shown in vivo. In order to dissect this regulatory network, we removed the complex interdependence of aggR and aar by placing the genes under the control of titratable promoters. We measured phenotypic and genotypic changes on downstream genes in EAEC strain 042 and E. coli K-12 strain DH5α, which lacks the AggR regulon. In EAEC, we found that low expression of aar increases aafA fimbrial gene expression via H-NS; however, when aar is more highly expressed, it acts as a negative regulator via AggR. In DH5α, aar affected expression of E. coli genes in some cases via H-NS and in some cases independent of H-NS. Our data support the model that Aar interacts in concert with AggR, H-NS, and possibly other regulators and that these interactions are likely to be functionally significant in vivo.

scholarly journals Stationary-Phase Quorum-Sensing Signals Affect Autoinducer-2 and Gene Expression in Escherichia coli

2004 ◽  
Vol 70 (4) ◽  
pp. 2038-2043 ◽  
Author(s):  
Dacheng Ren ◽  
Laura A. Bedzyk ◽  
Rick W. Ye ◽  
Stuart M. Thomas ◽  
Thomas K. Wood
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Quorum Sensing ◽  
Stationary Phase ◽  
Growth Stages ◽  
E Coli ◽  
Stationary Culture ◽  
Autoinducer 2 ◽  
K 12 ◽  
Quorum Sensing Signals

ABSTRACT Quorum sensing via autoinducer-2 (AI-2) has been identified in different strains, including those from Escherichia, Vibrio, Streptococcus, and Bacillus species, and previous studies have suggested the existence of additional quorum-sensing signals working in the stationary phase of Escherichia coli cultures. To investigate the presence and global effect of these possible quorum-sensing signals other than AI-2, DNA microarrays were used to study the effect of stationary-phase signals on the gene expression of early exponential-phase cells of the AI-2-deficient strain E. coli DH5α. For statistically significant differential gene expression (P < 0.05), 14 genes were induced by supernatants from a stationary culture and 6 genes were repressed, suggesting the involvement of indole (induction of tnaA and tnaL) and phosphate (repression of phoA, phoB, and phoU). To study the stability of the signals, the stationary-phase supernatant was autoclaved and was used to study its effect on E. coli gene expression. Three genes were induced by autoclaved stationary-phase supernatant, and 34 genes were repressed. In total, three genes (ompC, ptsA, and btuB) were induced and five genes (nupC, phoB, phoU, argT, and ompF) were repressed by both fresh and autoclaved stationary-phase supernatants. Furthermore, supernatant from E. coli DH5α stationary culture was found to repress E. coli K-12 AI-2 concentrations by 4.8-fold ± 0.4-fold, suggesting that an additional quorum-sensing system in E. coli exists and that gene expression is controlled as a network with different signals working at different growth stages.

scholarly journals Dual Overlapping Promoters Control napF (Periplasmic Nitrate Reductase) Operon Expression in Escherichia coli K-12

2003 ◽  
Vol 185 (19) ◽  
pp. 5862-5870 ◽  
Author(s):  
Valley Stewart ◽  
Peggy J. Bledsoe ◽  
Stanly B. Williams
Keyword(s):  
Escherichia Coli ◽  
Nitrate Reductase ◽  
Control Region ◽  
Transcription Initiation ◽  
Initiation Site ◽  
Periplasmic Nitrate Reductase ◽  
Fnr Protein ◽  
Operon Expression ◽  
K 12 ◽  
A Site

ABSTRACT Escherichia coli elaborates a flexible respiratory metabolism, involving differential synthesis of isoenzymes for many oxidation and reduction reactions. Periplasmic nitrate reductase, encoded by the napFDAGHBC operon, functions with concentrations of nitrate that are too low to support respiration by membrane-bound nitrate reductase. The napF operon control region exhibits unusual organization of DNA binding sites for the transcription regulators Fnr and NarP, which activate transcription in response to anaerobiosis and nitrate, respectively. Previous studies have shown that the napF operon control region directs synthesis of two transcripts whose 5′ ends differ by about 3 nucleotides. We constructed mutant control regions in which either of the two promoter −10 regions is inactivated. Results indicate that the downstream promoter (P1) was responsible for Fnr- and NarP-regulated napF operon expression, whereas transcription from the upstream promoter (P2) was activated only weakly by the Fnr protein and was inhibited by phospho-NarP and -NarL proteins. The physiological function of promoter P2 is unknown. These results establish the unconventional napF operon control region architecture, in which the major promoter P1 is activated by the Fnr protein bound to a site centered at −64.5 with respect to the transcription initiation site, working in conjunction with the phospho-NarP protein bound to a site centered at −44.5.

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