Characteristics and prevalence within serogroup O4 of a J96-like clonal group of uropathogenic Escherichia coli O4:H5 containing the class I and class III alleles of papG.

ABSTRACT In the murine model of urinary tract infections (UTI), cystitis by uropathogenic Escherichia coli (UPEC) occurs through an intimate relationship with the bladder superficial umbrella cell entailing cycles of adherence, invasion, intracellular bacterial community (IBC) formation, and dispersal (fluxing) from the intracellular environment. IBC dispersal is a key step that results in the spread of bacteria over the epithelial surface to initiate additional rounds of IBC formation. We investigated the role of flagella in mediating adherence and motility during UTI, hypothesizing that the dispersion of the IBC would be incomplete in the absence of motility, thus interrupting the IBC pathway and attenuating the infection. Using gfp reporter fusions, the expression of the flagellar class I flhDC and class III fliC genes was monitored to track key points of regulation throughout the pathogenic cascade. In vitro, growth under conditions promoting motility resulted in the robust expression of both fusions. In contrast, only the class I fusion produced significant expression throughout early stages of IBC development including the dispersion stage. Thus, unlike in vitro modeling of motility, the regulatory cascade appeared incomplete in vivo. Throughout IBC formation, nonmotile ΔfliC mutants achieved the same number of IBCs as the wild-type (wt) strain, demonstrating that flagella are neither essential nor required for first- or second-generation IBC formation. However, in competition experiments between wt and ΔfliC strains, the wt was shown to have a fitness advantage in persisting throughout the urinary tract for 2 weeks, demonstrating a subtle but measurable role for flagella in virulence.

Download Full-text

Cyclic AMP Receptor Protein-Dependent Activation of the Escherichia coli acsP2 Promoter by a Synergistic Class III Mechanism

Journal of Bacteriology ◽

10.1128/jb.185.17.5148-5157.2003 ◽

2003 ◽

Vol 185 (17) ◽

pp. 5148-5157 ◽

Cited By ~ 62

Author(s):

Christine M. Beatty ◽

Douglas F. Browning ◽

Stephen J. W. Busby ◽

Alan J. Wolfe

Keyword(s):

Escherichia Coli ◽

Cyclic Amp ◽

Rna Polymerase ◽

Class I ◽

Receptor Protein ◽

Class Iii ◽

Α Subunit ◽

Cyclic Amp Receptor Protein ◽

Rna Polymerase Holoenzyme

ABSTRACT The cyclic AMP receptor protein (CRP) activates transcription of the Escherichia coli acs gene, which encodes an acetate-scavenging enzyme required for fitness during periods of carbon starvation. Two promoters direct transcription of acs, the distal acsP1 and the proximal acsP2. In this study, we demonstrated that acsP2 can function as the major promoter and showed by in vitro studies that CRP facilitates transcription by “focusing” RNA polymerase to acsP2. We proposed that CRP activates transcription from acsP2 by a synergistic class III mechanism. Consistent with this proposal, we showed that CRP binds two sites, CRP I and CRP II. Induction of acs expression absolutely required CRP I, while optimal expression required both CRP I and CRP II. The locations of these DNA sites for CRP (centered at positions −69.5 and −122.5, respectively) suggest that CRP interacts with RNA polymerase through class I interactions. In support of this hypothesis, we demonstrated that acs transcription requires the surfaces of CRP and the C-terminal domain of the α subunit of RNA polymerase holoenzyme (α-CTD), which is known to participate in class I interactions: activating region 1 of CRP and the 287, 265, and 261 determinants of the α-CTD. Other surface-exposed residues in the α-CTD contributed to acs transcription, suggesting that the α-CTD may interact with at least one protein other than CRP.

Download Full-text

CD18-dependent activation of the neutrophil NADPH oxidase during phagocytosis of Escherichia coli or Staphylococcus aureus is regulated by class III but not class I or II PI3Ks

Blood ◽

10.1182/blood-2008-04-149450 ◽

2008 ◽

Vol 112 (13) ◽

pp. 5202-5211 ◽

Cited By ~ 65

Author(s):

Karen E. Anderson ◽

Keith B. Boyle ◽

Keith Davidson ◽

Tamara A. M. Chessa ◽

Suhasini Kulkarni ◽

...

Keyword(s):

Escherichia Coli ◽

Staphylococcus Aureus ◽

Nadph Oxidase ◽

Cell Line ◽

Microbial Pathogens ◽

Class I ◽

Human Neutrophils ◽

Class Iii ◽

Single Class ◽

E Coli

Abstract Phagocytosis and activation of the NADPH oxidase are important mechanisms by which neutrophils and macrophages engulf and kill microbial pathogens. We investigated the role of PI3K signaling pathways in the regulation of the oxidase during phagocytosis of Staphylococcus aureus and Escherichia coli by mouse and human neutrophils, a mouse macrophage-like cell line and a human myeloid-like cell line. Phagocytosis of these bacteria was promoted by serum, independent of serum-derived antibodies, and effectively abolished in mouse neutrophils lacking the β2-integrin common chain, CD18. A combination of PI3K isoform-selective inhibitors, mouse knock-outs, and RNA-interference indicated CD18-dependent activation of the oxidase was independent of class I and II PI3Ks, but substantially dependent on the single class III isoform (Vps34). Class III PI3K was responsible for the synthesis of PtdIns(3)P on phagosomes containing either bacteria. The use of mouse neutrophils carrying an appropriate knock-in mutation indicated that PtdIns(3)P binding to the PX domain of their p40phox oxidase subunit is important for oxidase activation in response to both S aureus and E coli. This interaction does not, however, account for all the PI3K sensitivity of these responses, particularly the oxidase response to E coli, suggesting that additional mechanisms for PtdIns(3)P-regulation of the oxidase must exist.

Download Full-text

Prevalence of Known P-Fimbrial G Alleles inEscherichia coli and Identification of a New Adhesin Class

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.8.3.637-640.2001 ◽

2001 ◽

Vol 8 (3) ◽

pp. 637-640 ◽

Cited By ~ 8

Author(s):

Shannon D. Manning ◽

Lixin Zhang ◽

Betsy Foxman ◽

Angela Spindler ◽

Patricia Tallman ◽

...

Keyword(s):

Escherichia Coli ◽

Class Ii ◽

Class I ◽

The Novel ◽

Inescherichia Coli ◽

Class Iii ◽

Cloning And Sequencing ◽

Fimbrial Adhesin

ABSTRACT Screening a large Escherichia coli collection for P-fimbrial adhesin classes identified 20 unclassifiable strains. Cloning and sequencing of papG from an unclassifiable strain identified another G allele. The novel adhesin gene has 65% identity to the class I adhesin gene, 44% identity to the class II adhesin gene, and 43% identity to the class III adhesin gene.

Download Full-text

Antibiotic resistance patterns and characterization of class I integron in uropathogenic escherichia coli in Lebanon. (c2008)

10.26756/th.2008.44 ◽

2008 ◽

Author(s):

Maya J. Farah

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Uropathogenic Escherichia Coli ◽

Class I ◽

Resistance Patterns ◽

Antibiotic Resistance Patterns

Download Full-text

Structure–function relationships in Escherichia coli adenylate cyclase

Biochemical Journal ◽

10.1042/bj20080350 ◽

2008 ◽

Vol 415 (3) ◽

pp. 449-454 ◽

Cited By ~ 8

Author(s):

Jürgen U. Linder

Keyword(s):

Escherichia Coli ◽

Adenylate Cyclase ◽

Metal Ion ◽

Catalytic Domain ◽

Vibrio Vulnificus ◽

Class I ◽

Class Iii ◽

E Coli ◽

Metal Cofactor ◽

Binding Residues

Class I adenylate cyclases are found in γ- and δ-proteobacteria. They play central roles in processes such as catabolite repression in Escherichia coli or development of full virulence in pathogens such as Yersinia enterocolitica and Vibrio vulnificus. The catalytic domain (residues 2–446) of the adenylate cyclase of E. coli was overexpressed and purified. It displayed a Vmax of 665 nmol of cAMP·mg−1·min−1 and a Km of 270 μM. Titration of the metal cofactor Mg2+ against the substrate ATP showed a requirement for free metal ions in addition to the MgATP complex, suggesting a two-metal-ion mechanism as is known for class II and class III adenylate cyclases. Twelve residues which are essential for catalysis were identified by mutagenesis of a total of 20 polar residues conserved in all class I adenylate cyclases. Five essential residues (Ser103, Ser113, Asp114, Asp116 and Trp118) were part of a region which is found in all members of the large DNA polymerase β-like nucleotidyltransferase superfamily. Alignment of the E. coli adenylate cyclase with the crystal structure of a distant member of the superfamily, archaeal tRNA CCA-adding enzyme, suggested that Asp114 and Asp116 are the metal-cofactor-ion-binding residues. The S103A mutant had a 17-fold higher Km than wild-type, demonstrating its important role in substrate binding. In comparison with the tRNA CCA-adding enzyme, Ser103 of the E. coli adenylate cyclase apparently binds the γ-phosphate group of ATP. Consistent with this function, the S103A mutation caused a marked reduction of discrimination between ATP- and ADP- or AMP-derived inhibitors.

Download Full-text