scholarly journals Slipped-Strand Mispairing Can Function as a Phase Variation Mechanism in Escherichia coli

2003 ◽  
Vol 185 (23) ◽  
pp. 6990-6994 ◽  
Author(s):  
Joshua Torres-Cruz ◽  
Marjan W. van der Woude
Keyword(s):  
Escherichia Coli ◽  
Haemophilus Influenzae ◽  
Reporter Gene ◽  
Phase Variation ◽  
Biologically Relevant ◽  
E Coli ◽  
Variation Mechanism ◽  
Relevant Range

ABSTRACT Slipped-strand mispairing (SSM) has not been identified as a mechanism of phase variation in Escherichia coli. Using a reporter gene, we show that sequences that cause phase variation by SSM in Haemophilus influenzae also lead to phase variation when introduced onto the chromosome of E. coli, and the frequencies of switching are in the biologically relevant range. Thus, the absence of SSM-mediated phase variation in E. coli does not appear to be due to a mechanistic constraint.

scholarly journals Fnr-, NarP- and NarL-Dependent Regulation of Transcription Initiation from the Haemophilus influenzae Rd napF (Periplasmic Nitrate Reductase) Promoter in Escherichia coli K-12

2005 ◽  
Vol 187 (20) ◽  
pp. 6928-6935 ◽  
Author(s):  
Valley Stewart ◽  
Peggy J. Bledsoe
Keyword(s):  
Escherichia Coli ◽  
Nitrate Reductase ◽  
Haemophilus Influenzae ◽  
Control Region ◽  
Binding Sites ◽  
Transcription Initiation ◽  
Transcriptional Control ◽  
Class I ◽  
E Coli ◽  
Fnr Protein

ABSTRACT Periplasmic nitrate reductase (napFDAGHBC operon product) functions in anaerobic respiration. Transcription initiation from the Escherichia coli napF operon control region is activated by the Fnr protein in response to anaerobiosis and by the NarQ-NarP two-component regulatory system in response to nitrate or nitrite. The binding sites for the Fnr and phospho-NarP proteins are centered at positions −64.5 and −44.5, respectively, with respect to the major transcription initiation point. The E. coli napF operon is a rare example of a class I Fnr-activated transcriptional control region, in which the Fnr protein binding site is located upstream of position −60. To broaden our understanding of napF operon transcriptional control, we studied the Haemophilus influenzae Rd napF operon control region, expressed as a napF-lacZ operon fusion in the surrogate host E. coli. Mutational analysis demonstrated that expression required binding sites for the Fnr and phospho-NarP proteins centered at positions −81.5 and −42.5, respectively. Transcription from the E. coli napF operon control region is activated by phospho-NarP but antagonized by the orthologous protein, phospho-NarL. By contrast, expression from the H. influenzae napF-lacZ operon fusion in E. coli was stimulated equally well by nitrate in both narP and narL null mutants, indicating that phospho-NarL and -NarP are equally effective regulators of this promoter. Overall, the H. influenzae napF operon control region provides a relatively simple model for studying synergistic transcription by the Fnr and phospho-NarP proteins acting from class I and class II locations, respectively.

scholarly journals A Genomic Islet Mediates Flagellar Phase Variation in Escherichia coli Strains Carrying the Flagellin-Specifying Locus flk

2008 ◽  
Vol 190 (13) ◽  
pp. 4470-4477 ◽  
Author(s):  
Lu Feng ◽  
Bin Liu ◽  
Yanqun Liu ◽  
Yuli A. Ratiner ◽  
Bo Hu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Phase Variation ◽  
Complementation Test ◽  
Direct Repeats ◽  
Integrase Gene ◽  
E Coli ◽  
Chromosomal Genes ◽  
Integration Sites ◽  
First Time ◽  
Transfer Potential

ABSTRACT The occurrence of unilateral flagellar phase variation was previously demonstrated in Escherichia coli strains carrying the non-fliC flagellin-specifying locus flk. In this study, we investigated the mechanism involved in this process. By using sequencing and sequence analysis, the flk region between the chromosomal genes yhaC and rnpB was characterized in all described flk-positive E. coli strains, including the H35 strain identified in this study (the other strains used are H3, H36, H47, and H53 strains), and this region was found to contain a putative integrase gene and flanking direct repeats in addition to the flk flagellin-specifying gene flkA and a fliC repressor gene, flkB, indicating that there is a typical genomic islet (GI), which was designated the flk GI. The horizontal transfer potential of the flk GI was indicated by detection of the excised extrachromosomal circular form of the flk GI. By generating fliC-expressing variants of H3 and H47 strains, unilateral flagellar phase variation in flk-positive strains was shown to be mediated by excision of the flk GI. The function of the proposed integrase gene was confirmed by deletion and a complementation test. The potential integration sites of the flk GI were identified. A general model for flagellar phase variation in flk-positive E. coli strains can be expressed as fliC off + flkA on → fliC on + flkA none. This is the first time that a molecular mechanism for flagellar phase variation has been reported for E. coli.

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 Pilus-Mediated Adherence of Haemophilus influenzae to Human Respiratory Mucins

2000 ◽  
Vol 68 (6) ◽  
pp. 3362-3367 ◽  
Author(s):  
Martin Kubiet ◽  
Reuben Ramphal ◽  
Allan Weber ◽  
Arnold Smith
Keyword(s):  
Escherichia Coli ◽  
Haemophilus Influenzae ◽  
Lung Disease ◽  
Gene Cluster ◽  
Otitis Media ◽  
Structural Gene ◽  
Chronic Lung Disease ◽  
Type B ◽  
E Coli

ABSTRACT Haemophilus influenzae, especially the nontypeable strains, are among the most common pathogens encountered in patients with chronic lung disease and otitis media. We and others have demonstrated that respiratory isolates of nontypeable H. influenzae bind to human mucins, but the mechanism of binding is not entirely clear. We have therefore examined the role of pili in the adherence of both type b and nontypeable H. influenzae to human respiratory mucins. We used isogenic H. influenzaestrains with a mutation in the structural gene for pilin (hifA), a laboratory H. influenzae strain transformed with a type b pilus gene cluster (from strain C54), antibodies raised against H. influenzae HifA, andEscherichia coli strains carrying a cloned type b pilus gene cluster (from strain AM30) in these studies. All bacteria lacking HifA or the pilus gene cluster had decreased adherence of piliatedH. influenzae to mucins, and Fab fragments of anti-HifA antibodies inhibited the adherence. E. coli strains carrying the cloned type b pilus gene cluster were six to seven times more adhesive than strains carrying the vector. The role of other putative adhesins was not examined and thus cannot be excluded, but these studies support a role for pili in the binding of H. influenzae to human respiratory mucins.

Induction of the SOS regulon of Haemophilus influenzae does not affect phase variation rates at tetranucleotide or dinucleotide repeats

Microbiology ◽  
2005 ◽  
Vol 151 (8) ◽  
pp. 2751-2763 ◽  
Author(s):  
Wendy A. Sweetman ◽  
E. Richard Moxon ◽  
Christopher D. Bayliss
Keyword(s):  
Haemophilus Influenzae ◽  
Uv Irradiation ◽  
Phase Variation ◽  
Sos Response ◽  
Site Directed Mutagenesis ◽  
Start Codon ◽  
Dinucleotide Repeats ◽  
Tetranucleotide Repeat ◽  
E Coli ◽  
Lexa Binding

Haemophilus influenzae has microsatellite repeat tracts in 5′ coding regions or promoters of several genes that are important for commensal and virulence behaviour. Changes in repeat number lead to switches in expression of these genes, a process referred to as phase variation. Hence, the virulence behaviour of this organism may be influenced by factors that alter the frequency of mutations in these repeat tracts. In Escherichia coli, induction of the SOS response destabilizes dinucleotide repeat tracts. H. influenzae encodes a homologue of the E. coli SOS repressor, LexA. The H. influenzae genome sequence was screened for the presence of the minimal consensus LexA-binding sequence from E. coli, CTG(N)10CAG, in order to identify genes with the potential to be SOS regulated. Twenty-five genes were identified that had LexA-binding sequences within 200 bp of the start codon. An H. influenzae non-inducible LexA mutant (lexA NI) was generated by site-directed mutagenesis. This mutant showed increased sensitivity, compared with wild-type (WT) cells, to both UV irradiation and mitomycin C (mitC) treatment. Semi-quantitative RT-PCR studies confirmed that H. influenzae mounts a LexA-regulated SOS response following DNA assault. Transcript levels of lexA, recA, recN, recX, ruvA and impA were increased in WT cells following DNA damage but not in lexA NI cells. Induction of the H. influenzae SOS response by UV irradiation or mitC treatment did not lead to any observable SOS-dependent changes in phase variation rates at either dinucleotide or tetranucleotide repeat tracts. Treatment with mitC caused a small increase in phase variation rates in both repeat tracts, independently of an SOS response. We suggest that the difference between H. influenzae and E. coli with regard to the effect of the SOS response on dinucleotide phase variation rates is due to the absence of any of the known trans-lesion synthesis DNA polymerases in H. influenzae.

scholarly journals The N-Acetyltransferase RimJ Responds to Environmental Stimuli To Repress pap Fimbrial Transcription in Escherichia coli

2002 ◽  
Vol 184 (16) ◽  
pp. 4334-4342 ◽  
Author(s):  
Christine A. White-Ziegler ◽  
Alia M. Black ◽  
Stacie H. Eliades ◽  
Sarah Young ◽  
Kimberly Porter
Keyword(s):  
Escherichia Coli ◽  
Low Temperature ◽  
Synthetic Lethality ◽  
Phase Variation ◽  
Luria Bertani ◽  
Thermoregulatory Response ◽  
Gene Encoding ◽  
Environmental Stimuli ◽  
Variation Mechanism ◽  
Insertion Mutations

ABSTRACT In uropathogenic Escherichia coli, P pili (Pap) facilitate binding to host epithelial cells and subsequent colonization. Whereas P pili can be produced at 37°C, the expression of these fimbriae is suppressed at 23°C. Previously, insertion mutations in rimJ, a gene encoding the N-terminal acetyltransferase of ribosomal protein S5, were shown to disrupt this thermoregulatory response, allowing papBA transcription at low temperature. In this study, we created an in-frame deletion of rimJ. This deletion relieved the repressive effects not only of low temperature but also of rich (Luria-Bertani [LB]) medium and glucose on papBA transcription, indicating that RimJ modulates papBA transcription in response to multiple environmental stimuli. papI transcription was also shown to be regulated by RimJ. papBA transcription is also controlled by a phase variation mechanism. We demonstrated that the regulators necessary to establish a phase ON state—PapI, PapB, Dam, Lrp, and cyclic AMP-CAP-are still required for papBA transcription in a rimJ mutant strain. rimJ mutations increase the rate at which bacteria transition into the phase ON state, indicating that RimJ inhibits the phase OFF→ON transition. A ΔrimJ hns651 mutant is viable on LB medium but not on minimal medium. This synthetic lethality, along with transcriptional analyses, indicates that RimJ and H-NS work through separate pathways to control papBA transcription. Mutations in rimJ do not greatly influence the transcription of the fan, daa, or fim operon, suggesting that RimJ may be a pap-specific regulator. Overexpression of rimJ under conditions repressive for papBA transcription complements the ΔrimJ mutation but has little effect on transcription under activating conditions, indicating that the ability of RimJ to regulate transcription is environmentally controlled.

Role of fimbrial adhesins in the pathogenesis of Escherichia coli infections

1992 ◽  
Vol 38 (7) ◽  
pp. 720-727 ◽  
Author(s):  
Jörg Hacker
Keyword(s):  
Escherichia Coli ◽  
Phase Variation ◽  
Gene Clusters ◽  
E Coli ◽  
Colonization Factor ◽  
Colonization Factors ◽  
Fimbrial Adhesins ◽  
Tract Infections ◽  
Fimbrial Adhesin

Escherichia coli strains are able to cause intestinal (enteritis, diarrhoeal diseases) and extraintestinal (urinary tract infections, sepsis, meningitis) infections. Most pathogenic E. coli strains produce specific fimbrial adhesins, which represent essential colonization factors: intestinal E. coli strains very often carry transferable plasmids with gene clusters specific for fimbrial adhesins, like K88 and K99, or colonization factor antigens (CFA) I and II. In contrast, the fimbrial gene clusters of extraintestinal E. coli strains, such as P, S, or F1C fimbriae, are located on the chromosomes. The fimbrial adhesin complexes consist of major and minor subunit proteins. Their binding specificity can generally be assayed in hemagglutination tests. In the case of fimbrial adhesins of intestinal E. coli strains, the major subunit proteins preferentially represent the hemagglutinating adhesins, whereas minor subunit proteins are the hemagglutinins of extraintestinal E. coli strains. Recently "alternative" adhesin proteins were identified, which have the capacity to bind to eukaryotic structures different from the receptors of the erythrocytes. Fimbrial adhesins are not constitutively expressed but are stringently regulated on the molecular level. Extraintestinal E. coli wild-type strains normally carry three or more fimbrial adhesin determinants, which have the capacity to influence the expression of one another (cross talk). Furthermore the fimbrial gene clusters undergo phase variation, which seems to be important for their contribution to pathogenesis of E. coli. Key words: fimbriae, adhesins, Escherichia coli, pathogenicity.

scholarly journals Crystal structures of the amino-terminal domain of LpoA from Escherichia coli and Haemophilus influenzae

2019 ◽  
Vol 75 (5) ◽  
pp. 368-376 ◽  
Author(s):  
Aaron Kelley ◽  
J. Vijayalakshmi ◽  
Mark A. Saper
Keyword(s):  
Crystal Structure ◽  
Escherichia Coli ◽  
Haemophilus Influenzae ◽  
Rotation Axis ◽  
Quantitative Description ◽  
Nmr Structure ◽  
Monoclinic Crystal ◽  
Orthorhombic Crystal Structure ◽  
E Coli ◽  
Amino Terminal

The bacterial periplasmic protein LpoA is an outer membrane lipoprotein and an activator for the cross-linking activity of PBP1A, a bifunctional peptidoglycan synthase. Previous structures of the amino-terminal (N) domain of LpoA showed it to consist entirely of helices and loops, with at least four tetratricopeptide-like repeats. Although the previously determined orthorhombic crystal structure of the N domain of Haemophilus influenzae LpoA showed a typical curved structure with a concave groove, an NMR structure of the same domain from Escherichia coli was relatively flat. Here, a crystal structure of the N domain of E. coli LpoA was determined to a resolution of 2.1 Å and was found to be more similar to the H. influenzae crystal structure than to the E. coli NMR structure. To provide a quantitative description for these comparisons, the various structures were superimposed pairwise by fitting the first half of each structure to its pairwise partner and then calculating the rotation axis that would optimally superimpose the second half. Differences in both the magnitude of the rotation and the direction of the rotation axis were observed between different pairs of structures. A 1.35 Å resolution structure of a monoclinic crystal form of the N domain of H. influenzae LpoA was also determined. In this structure, the subdomains rotate 10° relative to those in the original orthorhombic H. influenzae crystal structure to further narrow the groove between the subdomains. To accommodate this, a bound chloride ion (in place of sulfate) allowed the closer approach of a helix that forms one side of the groove.

scholarly journals Continuous Control in Bacterial Regulatory Circuits

2004 ◽  
Vol 186 (22) ◽  
pp. 7618-7625 ◽  
Author(s):  
Eric Batchelor ◽  
Thomas J. Silhavy ◽  
Mark Goulian
Keyword(s):  
Escherichia Coli ◽  
Tetracycline Resistance ◽  
Open Loop ◽  
Expression Level ◽  
Continuous Control ◽  
Biologically Relevant ◽  
E Coli ◽  
Naturally Occurring ◽  
Regulatory Circuit ◽  
Regulatory Circuits

ABSTRACT We show that for two well-characterized regulatory circuits in Escherichia coli, Tn10 tetracycline resistance and porin osmoregulation, the transcriptional outputs in individual cells are graded functions of the applied stimuli. These systems are therefore examples of naturally occurring regulatory circuits that exhibit continuous control of transcription. Surprisingly, however, we find that porin osmoregulation is open loop; i.e., the porin expression level does not feed back into the regulatory circuit. This mode of control is particularly interesting for an organism such as E. coli, which proliferates in diverse environments, and raises important questions regarding the biologically relevant inputs and outputs for this system.

scholarly journals Mutations in Haemophilus influenzae Mismatch Repair Genes Increase Mutation Rates of Dinucleotide Repeat Tracts but Not Dinucleotide Repeat-Driven Pilin Phase Variation Rates

2004 ◽  
Vol 186 (10) ◽  
pp. 2928-2935 ◽  
Author(s):  
Christopher D. Bayliss ◽  
Wendy A. Sweetman ◽  
E. Richard Moxon
Keyword(s):  
Haemophilus Influenzae ◽  
Mismatch Repair ◽  
Bacterial Species ◽  
Phase Variation ◽  
Dinucleotide Repeat ◽  
Phase Variable ◽  
Upper Respiratory Tract ◽  
E Coli ◽  
Serotype B ◽  
Repair Genes

ABSTRACT High-frequency, reversible switches in expression of surface antigens, referred to as phase variation (PV), are characteristic of Haemophilus influenzae. PV enables this bacterial species, an obligate commensal and pathogen of the human upper respiratory tract, to adapt to changes in the host environment. Phase-variable hemagglutinating pili are expressed by many H. influenzae isolates. PV involves alterations in the number of 5′ TA repeats located between the −10 and −35 promoter elements of the overlapping, divergently orientated promoters of hifA and hifBCDE, whose products mediate biosynthesis and assembly of pili. Dinucleotide repeat tracts are destabilized by mismatch repair (MMR) mutations in Escherichia coli. The influence of mutations in MMR genes of H. influenzae strain Rd on dinucleotide repeat-mediated PV rates was investigated by using reporter constructs containing 20 5′ AT repeats. Mutations in mutS, mutL, and mutH elevated rates approximately 30-fold, while rates in dam and uvrD mutants were increased 14- and 3-fold, respectively. PV rates of constructs containing 10 to 12 5′ AT repeats were significantly elevated in mutS mutants of H. influenzae strains Rd and Eagan. An intact hif locus was found in 14 and 12% of representative nontypeable H. influenzae isolates associated with either otitis media or carriage, respectively. Nine or more tandem 5′ TA repeats were present in the promoter region. Surprisingly, inactivation of mutS in two serotype b H. influenzae strains did not alter pilin PV rates. Thus, although functionally analogous to the E. coli MMR pathway and active on dinucleotide repeat tracts, defects in H. influenzae MMR do not affect 5′ TA-mediated pilin PV.

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