scholarly journals The LysR-Type Transcriptional Regulator QseD Alters Type Three Secretion in Enterohemorrhagic Escherichia coli and Motility in K-12 Escherichia coli

2010 ◽  
Vol 192 (14) ◽  
pp. 3699-3712 ◽  
Author(s):  
Benjamin J. Habdas ◽  
Jennifer Smart ◽  
James B. Kaper ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Transcriptional Regulator ◽  
Dominant Negative ◽  
Enterohemorrhagic Escherichia Coli ◽  
Lesion Formation ◽  
E Coli ◽  
Component Systems ◽  
Two Component Systems ◽  
Enterocyte Effacement ◽  
K 12

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) O157:H7 responds to the host-produced epinephrine and norepinephrine, and bacterially produced autoinducer 3 (AI-3), through two-component systems. Further integration of multiple regulatory signaling networks, involving regulators such as the LysR-type transcriptional regulator (LTTR) QseA, promotes effective regulation of virulence factors. These include the production of flagella, a phage-encoded Shiga toxin, and genes within the locus of enterocyte effacement (LEE) responsible for attaching and effacing (AE) lesion formation. Here, we describe a new member of this signaling cascade, an LTTR heretofore renamed QseD (quorum-sensing E. coli regulator D). QseD is present in all enterobacteria but exists almost exclusively in O157:H7 isolates as a helix-turn-helix (HTH) truncated isoform. This “short” isoform (sQseD) is still able to regulate gene expression through a different mechanism than the full-length K-12 E. coli “long” QseD isoform (lQseD). The EHEC ΔqseD mutant exhibits increased expression of all LEE operons and deregulation of AE lesion formation. The loss of qseD in EHEC does not affect motility, but the K-12 ΔqseD mutant is hypermotile. While the lQseD directly binds to the ler promoter, encoding the LEE master regulator, to repress LEE transcription, the sQseD isoform does not. LTTRs bind to DNA as tetramers, and these data suggest that sQseD regulates ler by forming heterotetramers with another LTTR. The LTTRs known to regulate LEE transcription, QseA and LrhA, do not interact with sQseD, suggesting that sQseD acts as a dominant-negative partner with a yet-unidentified LTTR.

scholarly journals Evolution of Enterohemorrhagic Escherichia coli Hemolysin Plasmids and the Locus for Enterocyte Effacement in Shiga Toxin-Producing E. coli

1998 ◽  
Vol 66 (6) ◽  
pp. 2553-2561 ◽  
Author(s):  
Patrick Boerlin ◽  
Shu Chen ◽  
John K. Colbourne ◽  
Roger Johnson ◽  
Stephanie De Grandis ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Restriction Pattern ◽  
Enterohemorrhagic Escherichia Coli ◽  
E Coli ◽  
Virulence Markers ◽  
Reference Isolate ◽  
Enterocyte Effacement ◽  
Escherichia Coli Hemolysin ◽  
K 12

ABSTRACT This study assessed the diversity of the enterohemorrhagicEscherichia coli (EHEC) hemolysin gene (ehxA) in a variety of Shiga toxin-producing E. coli (STEC) serotypes and the relationship between ehxA types and virulence markers on the locus for enterocyte effacement (LEE). Restriction fragment length polymorphism of the ehxA gene and flanking sequences and of the E. coli attaching and effacing (eae) gene was determined for 79 EHEC hemolysin-positive STEC isolates of 37 serotypes. Two main groups of EHEC hemolysin sequences and associated plasmids, which corresponded to the eae-positive and the eae-negative isolates, were delineated. Comparisons of the ehxA gene sequences of representative isolates of each group showed that this gene and the rest of the EHEC hemolysin operon are highly conserved. Digestion of anehxA PCR product with the restriction endonucleaseTaqI showed a unique restriction pattern foreae-negative isolates and another one for isolates of serotypes O157:H7 and O157:NM. A conserved fragment of 5.6 kb with four potential open reading frames was identified on the EHEC hemolysin plasmid of eae-positive STEC. Phylogenetic analysis of a subset of 27 STEC isolates, one enteropathogenic E. coliisolate, and a K-12 reference isolate showed thateae-positive STEC isolates all belong to a single evolutionary lineage and that the EHEC hemolysin plasmid and theehxA gene evolved within this lineage without recent horizontal transfer. However, the eae gene and the LEE appear to have been transferred horizontally within this STEC lineage on several occasions. The reasons for the lack of transfer or maintenance of the LEE in other STEC lineages are not clear and require further study.

scholarly journals Elucidation of Regulatory Modes for Five Two-Component Systems in Escherichia coli Reveals Novel Relationships

mSystems ◽  
2020 ◽  
Vol 5 (6) ◽  
Author(s):  
Kumari Sonal Choudhary ◽  
Julia A. Kleinmanns ◽  
Katherine Decker ◽  
Anand V. Sastry ◽  
Ye Gao ◽  
...  
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Target Gene ◽  
Target Genes ◽  
Rna Seq ◽  
Content Type ◽  
E Coli ◽  
Component Systems ◽  
Two Component ◽  
Two Component Systems

ABSTRACT Escherichia coli uses two-component systems (TCSs) to respond to environmental signals. TCSs affect gene expression and are parts of E. coli’s global transcriptional regulatory network (TRN). Here, we identified the regulons of five TCSs in E. coli MG1655: BaeSR and CpxAR, which were stimulated by ethanol stress; KdpDE and PhoRB, induced by limiting potassium and phosphate, respectively; and ZraSR, stimulated by zinc. We analyzed RNA-seq data using independent component analysis (ICA). ChIP-exo data were used to validate condition-specific target gene binding sites. Based on these data, we do the following: (i) identify the target genes for each TCS; (ii) show how the target genes are transcribed in response to stimulus; and (iii) reveal novel relationships between TCSs, which indicate noncognate inducers for various response regulators, such as BaeR to iron starvation, CpxR to phosphate limitation, and PhoB and ZraR to cell envelope stress. Our understanding of the TRN in E. coli is thus notably expanded. IMPORTANCE E. coli is a common commensal microbe found in the human gut microenvironment; however, some strains cause diseases like diarrhea, urinary tract infections, and meningitis. E. coli’s two-component systems (TCSs) modulate target gene expression, especially related to virulence, pathogenesis, and antimicrobial peptides, in response to environmental stimuli. Thus, it is of utmost importance to understand the transcriptional regulation of TCSs to infer bacterial environmental adaptation and disease pathogenicity. Utilizing a combinatorial approach integrating RNA sequencing (RNA-seq), independent component analysis, chromatin immunoprecipitation coupled with exonuclease treatment (ChIP-exo), and data mining, we suggest five different modes of TCS transcriptional regulation. Our data further highlight noncognate inducers of TCSs, which emphasizes the cross-regulatory nature of TCSs in E. coli and suggests that TCSs may have a role beyond their cognate functionalities. In summary, these results can lead to an understanding of the metabolic capabilities of bacteria and correctly predict complex phenotype under diverse conditions, especially when further incorporated with genome-scale metabolic models.

scholarly journals The Locus of Enterocyte Effacement-Encoded Effector Proteins All Promote Enterohemorrhagic Escherichia coli Pathogenicity in Infant Rabbits

2005 ◽  
Vol 73 (3) ◽  
pp. 1466-1474 ◽  
Author(s):  
Jennifer M. Ritchie ◽  
Matthew K. Waldor
Keyword(s):  
Escherichia Coli ◽  
Effector Proteins ◽  
Enterohemorrhagic Escherichia Coli ◽  
Lesion Formation ◽  
Specific Effects ◽  
Genes Encoding ◽  
Organ Specific ◽  
Rabbit Intestine ◽  
Enterocyte Effacement

ABSTRACT The genes encoding the enterohemorrhagic Escherichia coli (EHEC) type III secretion system (TTSS) and five effector proteins secreted by the TTSS are located on the locus of enterocyte effacement (LEE) pathogenicity island. Deletion of tir, which encodes one of these effector proteins, results in a profound reduction (∼10,000-fold) in EHEC colonization of the infant rabbit intestine, but the in vivo phenotypes of other LEE genes are unknown. Here, we constructed in-frame deletions in escN, the putative ATPase component of the TTSS, and the genes encoding the four other LEE-encoded effector proteins, EspH, Map, EspF, and EspG, to investigate the contributions of the TTSS and the translocated effector proteins to EHEC pathogenicity in infant rabbits. We found that the TTSS is required for EHEC colonization and attaching and effacing (A/E) lesion formation in the rabbit intestine. Deletion of escN reduced EHEC recovery from the rabbit intestine by ∼10,000-fold. Although EspH, Map, EspF, and EspG were not required for A/E lesion formation in the rabbit intestine or in HeLa cells, these effector proteins promote EHEC colonization. Colonization by the espH and espF mutants was reduced throughout the intestine. In contrast, colonization by the map and espG mutants was reduced only in the small intestine, indicating that Map and EspG have organ-specific effects. EspF appears to down-regulate the host response to EHEC, since we observed increased accumulation of polymorphonuclear leukocytes in the colonic mucosa of rabbits infected with the EHEC espF mutant. Thus, all the known LEE-encoded effector proteins influence EHEC pathogenicity.

scholarly journals Contribution of the Twin Arginine Translocation System to the Virulence of Enterohemorrhagic Escherichia coli O157:H7

2003 ◽  
Vol 71 (9) ◽  
pp. 4908-4916 ◽  
Author(s):  
Nathalie Pradel ◽  
Changyun Ye ◽  
Valérie Livrelli ◽  
Jianguo Xu ◽  
Bernard Joly ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Immunoblot Analysis ◽  
Vero Cells ◽  
Soft Agar ◽  
Enterohemorrhagic Escherichia Coli ◽  
E Coli ◽  
Twin Arginine Translocation ◽  
Tat System ◽  
K 12

ABSTRACT Shiga toxin-producing Escherichia coli O157:H7 is a major food-borne infectious pathogen. In order to analyze the contribution of the twin arginine translocation (TAT) system to the virulence of E. coli O157:H7, we deleted the tatABC genes of the O157:H7 EDL933 reference strain. The mutant displayed attenuated toxicity on Vero cells and completely lost motility on soft agar plates. Further analyses revealed that the ΔtatABC mutation impaired the secretion of the Shiga toxin 1 (Stx1) and abolished the synthesis of H7 flagellin, which are two major known virulence factors of enterohemorrhagic E. coli O157:H7. Expression of the EDL933 stxAB 1 genes in E. coli K-12 conferred verotoxicity on this nonpathogenic strain. Remarkably, cytotoxicity assay and immunoblot analysis showed, for the first time, an accumulation of the holotoxin complex in the periplasm of the wild-type strain and that a much smaller amount of StxA1 and reduced verotoxicity were detected in the ΔtatC mutant cells. Together, these results establish that the TAT system of E. coli O157:H7 is an important virulence determinant of this enterohemorrhagic pathogen.

scholarly journals Quorum-Sensing Escherichia coli Regulator A: a Regulator of the LysR Family Involved in the Regulation of the Locus of Enterocyte Effacement Pathogenicity Island in Enterohemorrhagic E. coli

2002 ◽  
Vol 70 (6) ◽  
pp. 3085-3093 ◽  
Author(s):  
Vanessa Sperandio ◽  
Caiyi C. Li ◽  
James B. Kaper
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Type Iii Secretion ◽  
Pathogenicity Island ◽  
The Other ◽  
E Coli ◽  
Type Iii ◽  
Enterocyte Effacement ◽  
K 12 ◽  
Lysr Family

ABSTRACT The locus of enterocyte effacement (LEE) is a chromosomal pathogenicity island that encodes the proteins involved in the formation of the attaching and effacing lesions by enterohemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC). The LEE comprises 41 open reading frames organized in five major operons, LEE1, LEE2, LEE3, tir (LEE5), and LEE4, which encode a type III secretion system, the intimin adhesin, the translocated intimin receptor (Tir), and other effector proteins. The first gene of LEE1 encodes the Ler regulator, which activates all the other genes within the LEE. We previously reported that the LEE genes were activated by quorum sensing through Ler (V. Sperandio, J. L. Mellies, W. Nguyen, S. Shin, and J. B. Kaper, Proc. Natl. Acad. Sci. USA 96:15196-15201, 1999). In this study we report that a putative regulator in the E. coli genome is itself activated by quorum sensing. This regulator is encoded by open reading frame b3243; belongs to the LysR family of regulators; is present in EHEC, EPEC, and E. coli K-12; and shares homology with the AphB and PtxR regulators of Vibrio cholerae and Pseudomonas aeruginosa, respectively. We confirmed the activation of b3243 by quorum sensing by using transcriptional fusions and renamed this regulator quorum-sensing E. coli regulator A (QseA). We observed that QseA activated transcription of ler and therefore of the other LEE genes. An EHEC qseA mutant had a striking reduction of type III secretion activity, which was complemented when qseA was provided in trans. Similar results were also observed with a qseA mutant of EPEC. The QseA regulator is part of the regulatory cascade that regulates EHEC and EPEC virulence genes by quorum sensing.

scholarly journals Cloning, Expression, and Characterization of Fimbrial Operon F9 from Enterohemorrhagic Escherichia coli O157:H7

2006 ◽  
Vol 74 (4) ◽  
pp. 2233-2244 ◽  
Author(s):  
Alison S. Low ◽  
Francis Dziva ◽  
Alfredo G. Torres ◽  
Jessenya L. Martinez ◽  
Tracy Rosser ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epithelial Cells ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Enterohemorrhagic Escherichia Coli ◽  
Wild Type ◽  
Induced Expression ◽  
E Coli ◽  
Extracellular Matrix Components ◽  
K 12

ABSTRACT Recent transposon mutagenesis studies with two enterohemorrhagic Escherichia coli (EHEC) strains, a sero- type O26:H- strain and a serotype O157:H7 strain, led to identification of a putative fimbrial operon that promotes colonization of young calves (1 to 2 weeks old). The distribution of the gene encoding the major fimbrial subunit present in O-island 61 of EHEC O157:H7 in a characterized set of 78 diarrheagenic E. coli strains was determined, and this gene was found in 87.2% of the strains and is therefore not an EHEC-specific region. The cluster was amplified by long-range PCR and cloned into the inducible expression vector pBAD18. Induced expression in E. coli K-12 led to production of fimbriae, as demonstrated by transmission electron microscopy and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. The fimbriae were purified, and sera to the purified major subunit were raised and used to demonstrate expression from wild-type E. coli O157:H7 strains. Induced expression of the fimbriae, designated F9 fimbriae, was used to characterize binding to bovine epithelial cells, bovine gastrointestinal tissue explants, and extracellular matrix components. The fimbriae promoted increases in the levels of E. coli K-12 binding only to bovine epithelial cells. In contrast, induced expression of F9 fimbriae in E. coli O157:H7 significantly reduced adherence of the bacteria to bovine gastrointestinal explant tissue. This may have been due to physical hindrance of type III secretion-dependent attachment. The main F9 subunit gene was deleted in E. coli O157:H7, and the resulting mutant was compared with the wild-type strain for colonization in weaned cattle. While the shedding levels of the mutant were reduced, the animals were still colonized at the terminal rectum, indicating that the adhesin is not responsible for the rectal tropism observed but may contribute to colonization at other sites, as demonstrated previously with very young animals.

scholarly journals Cross-Reactive Protection against Enterohemorrhagic Escherichia coli Infection by Enteropathogenic E. coli in a Mouse Model

2011 ◽  
Vol 79 (6) ◽  
pp. 2224-2233 ◽  
Author(s):  
Carla Calderon Toledo ◽  
Ida Arvidsson ◽  
Diana Karpman
Keyword(s):  
Escherichia Coli ◽  
Weight Loss ◽  
Mouse Model ◽  
Enterohemorrhagic Escherichia Coli ◽  
Epec Strain ◽  
Content Type ◽  
E Coli ◽  
Renal Histology ◽  
Escherichia Coli Infection ◽  
Enterocyte Effacement

ABSTRACTEnteropathogenicEscherichia coli(EPEC) and enterohemorrhagicE. coli(EHEC) are related attaching and effacing (A/E) pathogens. The genes responsible for the A/E pathology are carried on a chromosomal pathogenicity island termed the locus of enterocyte effacement (LEE). Both pathogens share a high degree of homology in the LEE and additional O islands. EHEC prevalence is much lower in areas where EPEC is endemic. This may be due to the development of antibodies against common EPEC and EHEC antigens. This study investigated the hypothesis that EPEC infections may protect against EHEC infections. We used a mouse model to inoculate BALB/c mice intragastrically, first with EPEC and then with EHEC (E. coliO157:H7). Four control groups received either a nonpathogenicE. coli(NPEC) strain followed by EHEC (NPEC/EHEC), phosphate-buffered saline (PBS) followed by EHEC (PBS/EHEC), EPEC/PBS, or PBS/PBS. Mice were monitored for weight loss and symptoms. EPEC colonized the intestine after challenge, and mice developed serum antibodies to intimin andE. colisecreted protein B (encoded in the LEE). Prechallenge with an EPEC strain had a protective effect after EHEC infection, as only a few mice developed mild symptoms, from which they recovered. These mice had an increase in body weight similar to that in control animals, and tissue morphology exhibited mild intestinal changes and normal renal histology. All mice that were not prechallenged with the EPEC strain developed mild to severe symptoms after EHEC infection, with weight loss as well as intestinal and renal histopathological changes. These data suggest that EPEC may protect against EHEC infection in this mouse model.

scholarly journals Global Analysis of Posttranscriptional Regulation by GlmY and GlmZ in Enterohemorrhagic Escherichia coli O157:H7

2015 ◽  
Vol 83 (4) ◽  
pp. 1286-1295 ◽  
Author(s):  
Charley C. Gruber ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Rna Sequencing ◽  
Posttranscriptional Regulation ◽  
Global Analysis ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Global Changes ◽  
Content Type ◽  
Enterocyte Effacement ◽  
K 12

EnterohemorrhagicEscherichia coli(EHEC) is a significant human pathogen and is the cause of bloody diarrhea and hemolytic-uremic syndrome. The virulence repertoire of EHEC includes the genes within the locus of enterocyte effacement (LEE) that are largely organized in five operons,LEE1toLEE5, which encode a type III secretion system, several effectors, chaperones, and regulatory proteins. In addition, EHEC also encodes several non-LEE-encoded effectors and fimbrial operons. The virulence genes of this pathogen are under a large amount of posttranscriptional regulation. The small RNAs (sRNAs) GlmY and GlmZ activate the translation of glucosamine synthase (GlmS) inE. coliK-12, and in EHEC they destabilize the 3′ fragments of theLEE4andLEE5operons and promote translation of the non-LEE-encoded effector EspFu. We investigated the global changes of EHEC gene expression governed by GlmY and GlmZ using RNA sequencing and gene arrays. This study extends the known effects of GlmY and GlmZ regulation to show that they promote expression of the curli adhesin, repress the expression of tryptophan metabolism genes, and promote the expression of acid resistance genes and the non-LEE-encoded effector NleA. In addition, seven novel EHEC-specific sRNAs were identified using RNA sequencing, and three of them—sRNA56, sRNA103, and sRNA350—were shown to regulate urease, fimbria, and the LEE, respectively. These findings expand the knowledge of posttranscriptional regulation in EHEC.

scholarly journals Evolutionary Silence of the Acid Chaperone Protein HdeB in Enterohemorrhagic Escherichia coli O157:H7

2011 ◽  
Vol 78 (4) ◽  
pp. 1004-1014 ◽  
Author(s):  
Michelle Q. Carter ◽  
Jacqueline W. Louie ◽  
Clifton K. Fagerquist ◽  
Omar Sultan ◽  
William G. Miller ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acid Resistance ◽  
Significant Loss ◽  
Enterohemorrhagic Escherichia Coli ◽  
Survival Strategies ◽  
Start Codon ◽  
Divergent Evolution ◽  
Content Type ◽  
E Coli ◽  
K 12

ABSTRACTThe periplasmic chaperones HdeA and HdeB are known to be important for cell survival at low pH (pH < 3) inEscherichia coliandShigellaspp. Here we investigated the roles of HdeA and HdeB in the survival of various enterohemorrhagicE. coli(EHEC) following exposure to pH 2.0. Similar to K-12 strains, the acid protections conferred by HdeA and HdeB in EHEC O145 were significant: loss of HdeA and HdeB led to over 100- to 1,000-fold reductions in acid survival, depending on the growth condition of prechallenge cells. However, this protection was much less inE. coliO157:H7 strains. Deletion ofhdeBdid not affect the acid survival of cells, and deletion ofhdeAled to less than a 5-fold decrease in survival. Sequence analysis of thehdeABoperon revealed a point mutation at the putative start codon of thehdeBgene in all 26E. coliO157:H7 strains analyzed, which shifted the ATG start codon to ATA. This mutation correlated with the lack of HdeB inE. coliO157:H7; however, the plasmid-borne O157-hdeBwas able to restore partially the acid resistance in anE. coliO145ΔhdeABmutant, suggesting the potential function of O157-HdeB as an acid chaperone. We conclude thatE. coliO157:H7 strains have evolved acid survival strategies independent of the HdeA/B chaperones and are more acid resistant than nonpathogenic K-12 for cells grown under nonfavorable culturing conditions such as in Luria-Bertani no-salt broth at 28°C. These results suggest a divergent evolution of acid resistance mechanisms withinE. coli.

scholarly journals Global Expression of Prophage Genes in Escherichia coli O157:H7 Strain EDL933 in Response to Norfloxacin

2005 ◽  
Vol 49 (3) ◽  
pp. 931-944 ◽  
Author(s):  
Sylvia Herold ◽  
Jutta Siebert ◽  
Andrea Huber ◽  
Herbert Schmidt
Keyword(s):  
Escherichia Coli ◽  
Primary Metabolism ◽  
Low Concentration ◽  
E Coli ◽  
Specific Phage ◽  
Stress Functions ◽  
Enterocyte Effacement ◽  
K 12 ◽  
Upregulated Genes ◽  
Production Cell

ABSTRACT We investigated the influence of a low concentration of the gyrase inhibitor norfloxacin on the transcriptome of enterohemorrhagic Escherichia coli O157:H7 strain EDL933. For this purpose, we used a commercial DNA microarray containing oligonucleotides specific for E. coli O157:H7 strains EDL933 and RIMD0509952 and E. coli K-12 strain MG1655. Under the conditions applied, 5,963 spots (94% of all spots) could be analyzed. Among these, 118 spots (P < 0.05) indicated transcriptional upregulation and 122 spots (P < 0.05) indicated transcriptional downregulation of the E. coli genes present on the array. Eighty-five upregulated EDL933 genes were phage borne. Fifty-two of them could be ascribed to the Shiga toxin-encoding phages (Stx phages) BP-933W and CP-933V; the other 33 genes belonged to non-Stx prophage elements in the EDL933 genome. Genes present in the BP-933W prophage genome were induced most strongly up to 158-fold in the case of stxA2 upon induction with norfloxacin. Twenty-two additional upregulated genes appeared to be E. coli O157:H7 strain RIMD0509952-specific phage elements, and the remaining 11 genes were related mainly to recombination and stress functions. Downregulation was indicated predominantly for genes responsible for bacterial primary metabolism, such as energy production, cell division, and amino acid biosynthesis. Interestingly, some genes present in the locus of enterocyte effacement appeared to be downregulated. The results of the study have shown that a low concentration of norfloxacin has profound effects on the transcriptome of E. coli O157:H7.

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