scholarly journals Genome-wide comparison of phage M13-infected vs. uninfectedEscherichia coli

2005 ◽  
Vol 51 (1) ◽  
pp. 29-35 ◽  
Author(s):  
Fredrik Karlsson ◽  
Ann-Christin Malmborg-Hager ◽  
Ann-Sofie Albrekt ◽  
Carl A.K Borrebaeck
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Acid Stress ◽  
Phage Infection ◽  
Transcriptional Analysis ◽  
Filamentous Phage ◽  
Phosphotransferase System ◽  
E Coli ◽  
Genome Wide

To identify Escherichia coli genes potentially regulated by filamentous phage infection, we used oligonucleotide microarrays. Genome-wide comparison of phage M13-infected and uninfected E. coli, 2 and 20 min after infection, was performed. The analysis revealed altered transcription levels of 12 E. coli genes in response to phage infection, and the observed regulation of phage genes correlated with the known in vivo pattern of M13 mRNA species. Ten of the 12 host genes affected could be grouped into 3 different categories based on cellular function, suggesting a coordinated response. The significantly upregulated genes encode proteins involved in reactions of the energy-generating phosphotransferase system and transcription processing, which could be related to phage transcription. No genes belonging to any known E. coli stress response pathways were scored as upregulated. Furthermore, phage infection led to significant downregulation of transcripts of the bacterial genes gadA, gadB, hdeA, gadE, slp, and crl. These downregulated genes are normally part of the host stress response mechanisms that protect the bacterium during conditions of acid stress and stationary phase transition. The phage-infected cells demonstrated impaired function of the oxidative and the glutamate-dependent acid resistance systems. Thus, global transcriptional analysis and functional analysis revealed previously unknown host responses to filamentous phage infection.Key words: filamentous phage infection, global transcriptional analysis, AR, Escherichia coli.

scholarly journals Inactivation of PadR, the Repressor of the Phenolic Acid Stress Response, by Molecular Interaction with Usp1, a Universal Stress Protein from Lactobacillus plantarum, in Escherichia coli

2009 ◽  
Vol 75 (16) ◽  
pp. 5273-5283 ◽  
Author(s):  
Jérôme Gury ◽  
Hélène Seraut ◽  
Ngoc Phuong Tran ◽  
Lise Barthelmebs ◽  
Stéphanie Weidmann ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Lactobacillus Plantarum ◽  
Phenolic Acid ◽  
Stress Protein ◽  
Acid Stress ◽  
Universal Stress Protein ◽  
E Coli ◽  
Gram Positive Bacteria ◽  
Acid Stress Response

ABSTRACT The phenolic acid decarboxylase gene padA is involved in the phenolic acid stress response (PASR) in gram-positive bacteria. In Lactobacillus plantarum, the padR gene encodes the negative transcriptional regulator of padA and is cotranscribed with a downstream gene, usp1, which encodes a putative universal stress protein (USP), Usp1, of unknown function. The usp1 gene is overexpressed during the PASR. However, the role and the mechanism of action of the USPs are unknown in gram-positive bacteria. Therefore, to gain insights into the role of USPs in the PASR; (i) a usp1 deletion mutant was constructed; (ii) the two genes padR and usp1 were coexpressed with padA under its own promoter as a reporter gene in Escherichia coli; and (iii) molecular in vitro interactions between the PadR, Usp1, and the padA promoter were studied. Although the usp1 mutant strain retained phenolic acid-dependent PAD activity, it displayed a greater sensitivity to strong acidic conditions compared to that of the wild-type strain. PadR cannot be inactivated directly by phenolic acid in E. coli recombinant cultures but is inactivated by Usp1 when the two proteins are coexpressed in E. coli. The PadR inactivation observed in recombinant E. coli cells was supported by electrophoretic mobility shift assays. Although Usp1 seems not to be absolutely required for the PASR, its capacity to inactivate PadR indicates that it could serve as an important mediator in acid stress response mechanisms through its capacity to interact with transcriptional regulators.

scholarly journals Structure of hibernating ribosomes studied by cryoelectron tomography in vitro and in situ

2010 ◽  
Vol 190 (4) ◽  
pp. 613-621 ◽  
Author(s):  
Julio O. Ortiz ◽  
Florian Brandt ◽  
Valério R.F. Matias ◽  
Lau Sennels ◽  
Juri Rappsilber ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
Spatial Organization ◽  
Three Dimensional ◽  
E Coli ◽  
Escherichia Coli Cells ◽  
Three Dimensional Configuration

Ribosomes arranged in pairs (100S) have been related with nutritional stress response and are believed to represent a “hibernation state.” Several proteins have been identified that are associated with 100S ribosomes but their spatial organization has hitherto not been characterized. We have used cryoelectron tomography to reveal the three-dimensional configuration of 100S ribosomes isolated from starved Escherichia coli cells and we have described their mode of interaction. In situ studies with intact E. coli cells allowed us to demonstrate that 100S ribosomes do exist in vivo and represent an easily reversible state of quiescence; they readily vanish when the growth medium is replenished.

scholarly journals Collaborative Regulation of Escherichia coli Glutamate-Dependent Acid Resistance by Two AraC-Like Regulators, GadX and GadW (YhiW)

2002 ◽  
Vol 184 (24) ◽  
pp. 7001-7012 ◽  
Author(s):  
Zhuo Ma ◽  
Hope Richard ◽  
Don L. Tucker ◽  
Tyrrell Conway ◽  
John W. Foster
Keyword(s):  
Escherichia Coli ◽  
Acid Stress ◽  
Acid Resistance ◽  
Negative Regulator ◽  
Receptor Protein ◽  
Growth Media ◽  
Infectious Dose ◽  
E Coli ◽  
Control Circuits

ABSTRACT An important feature of Escherichia coli pathogenesis is an ability to withstand extremely acidic environments of pH 2 or lower. This acid resistance property contributes to the low infectious dose of pathogenic E. coli species. One very efficient E. coli acid resistance system encompasses two isoforms of glutamate decarboxylase (gadA and gadB) and a putative glutamate:γ-amino butyric acid (GABA) antiporter (gadC). The system is subject to complex controls that vary with growth media, growth phase, and growth pH. Previous work has revealed that the system is controlled by two sigma factors, two negative regulators (cyclic AMP receptor protein [CRP] and H-NS), and an AraC-like regulator called GadX. Earlier evidence suggested that the GadX protein acts both as a positive and negative regulator of the gadA and gadBC genes depending on environmental conditions. New data clarify this finding, revealing a collaborative regulation between GadX and another AraC-like regulator called GadW (previously YhiW). GadX and GadW are DNA binding proteins that form homodimers in vivo and are 42% homologous to each other. GadX activates expression of gadA and gadBC at any pH, while GadW inhibits GadX-dependent activation. Regulation of gadA and gadBC by either regulator requires an upstream, 20-bp GAD box sequence. Northern blot analysis further indicates that GadW represses expression of gadX. The results suggest a control circuit whereby GadW interacts with both the gadA and gadX promoters. GadW clearly represses gadX and, in situations where GadX is missing, activates gadA and gadBC. GadX, however, activates only gadA and gadBC expression. CRP also represses gadX expression. It does this primarily by repressing production of sigma S, the sigma factor responsible for gadX expression. In fact, the acid induction of gadA and gadBC observed when rich-medium cultures enter stationary phase corresponds to the acid induction of sigma S production. These complex control circuits impose tight rein over expression of the gadA and gadBC system yet provide flexibility for inducing acid resistance under many conditions that presage acid stress.

scholarly journals Chromatinization of Escherichia coli with archaeal histones

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Maria Rojec ◽  
Antoine Hocher ◽  
Kathryn M Stevens ◽  
Matthias Merkenschlager ◽  
Tobias Warnecke
Keyword(s):  
Escherichia Coli ◽  
Nucleosome Occupancy ◽  
Micrococcal Nuclease ◽  
E Coli ◽  
Genome Wide ◽  
Micrococcal Nuclease Digestion ◽  
Nuclease Digestion ◽  
Methanothermus Fervidus ◽  
Chromatin Proteins

Nucleosomes restrict DNA accessibility throughout eukaryotic genomes, with repercussions for replication, transcription, and other DNA-templated processes. How this globally restrictive organization emerged during evolution remains poorly understood. Here, to better understand the challenges associated with establishing globally restrictive chromatin, we express histones in a naive system that has not evolved to deal with nucleosomal structures: Escherichia coli. We find that histone proteins from the archaeon Methanothermus fervidus assemble on the E. coli chromosome in vivo and protect DNA from micrococcal nuclease digestion, allowing us to map binding footprints genome-wide. We show that higher nucleosome occupancy at promoters is associated with lower transcript levels, consistent with local repressive effects. Surprisingly, however, this sudden enforced chromatinization has only mild repercussions for growth unless cells experience topological stress. Our results suggest that histones can become established as ubiquitous chromatin proteins without interfering critically with key DNA-templated processes.

scholarly journals Chitin, Chitin Oligosaccharide, and Chitin Disaccharide Metabolism of Escherichia coli Revisited: Reassignment of the Roles of ChiA, ChbR, ChbF, and ChbG

2021 ◽  
pp. 1-17
Author(s):  
Axel Walter ◽  
Simon Friz ◽  
Christoph Mayer
Keyword(s):  
Escherichia Coli ◽  
Catalytic Mechanism ◽  
Phosphotransferase System ◽  
E Coli ◽  
The Family ◽  
Acetyl Group ◽  
Bona Fide ◽  
Modeling Data

<i>Escherichia coli</i> is unable to grow on polymeric and oligomeric chitin, but grows on chitin disaccharide (GlcNAc-GlcNAc; <i>N,N</i>′-diacetylchitobiose) and chitin trisaccharide (GlcNAc-GlcNAc-GlcNAc; <i>N,N</i>′<i>,N</i>′′-triacetylchitotriose) via expression of the <i>chb</i> operon (<i>chbBCARFG</i>). The phosphotransferase system (PTS) transporter ChbBCA facilitates transport of both saccharides across the inner membrane and their concomitant phosphorylation at the non-reducing end, intracellularly yielding GlcNAc 6-phosphate-GlcNAc (GlcNAc6P-GlcNAc) and GlcNAc6P-GlcNAc-GlcNAc, respectively. We revisited the intracellular catabolism of the PTS products, thereby correcting the reported functions of the 6-phospho-glycosidase ChbF, the monodeacetylase ChbG, and the transcriptional regulator ChbR. Intracellular accumulation of glucosamine 6P-GlcNAc (GlcN6P-GlcNAc) and GlcN6P-GlcNAc-GlcNAc in a <i>chbF</i> mutant unraveled a role for ChbG as a monodeacetylase that removes the <i>N-</i>acetyl group at the non-reducing end. Consequently, GlcN6P- but not GlcNAc6P-containing saccharides likely function as coactivators of ChbR. Furthermore, ChbF removed the GlcN6P from the non-reducing terminus of the former saccharides, thereby degrading the inducers of the <i>chb</i> operon and facilitating growth on the saccharides. Consequently, ChbF was unable to hydrolyze GlcNAc6P-residues from the non-reducing end, contrary to previous assumptions but in agreement with structural modeling data and with the unusual catalytic mechanism of the family 4 of glycosidases, to which ChbF belongs. We also refuted the assumption that ChiA is a bifunctional endochitinase/lysozyme ChiA, and show that it is unable to degrade peptidoglycans but acts as a bona fide chitinase in vitro and in vivo, enabling growth of <i>E. coli</i> on chitin oligosaccharides when ectopically expressed. Overall, this study revises our understanding of the chitin, chitin oligosaccharide, and chitin disaccharide metabolism of <i>E. coli</i>.

scholarly journals Loss of RpoS results in attenuated Escherichia coli colonization of human intestinal organoids and a competitive disadvantage within the germ-free mouse intestine

2020 ◽  
Author(s):  
Madeline R. Barron ◽  
Roberto J. Cieza ◽  
David R. Hill ◽  
Sha Huang ◽  
Veda K. Yadagiri ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stress Response ◽  
General Stress Response ◽  
E Coli ◽  
Germ Free ◽  
General Stress ◽  
Intestinal Organoids ◽  
Mouse Intestine

AbstractPluripotent stem-cell-derived human intestinal organoids (HIOs) are three-dimensional, multicellular structures that model a previously uncolonized, naïve intestinal epithelium in an in vitro system. We recently demonstrated that microinjection of the non-pathogenic Escherichia coli strain, ECOR2, into HIOs induced morphological and functional maturation of the HIO epithelium, including increased secretion of mucins and cationic antimicrobial peptides. In the current work, we use ECOR2 as a biological probe to investigate the bacterial response to colonization of the HIO lumen. In E. coli and other Gram-negative bacteria, adaptation to environmental stress is regulated by the general stress response sigma factor, RpoS. We generated an isogenic ∆rpoS ECOR2 mutant to compare challenges faced by a bacterium during colonization of the HIO lumen relative to the germ-free mouse intestine, which is currently the best available system for studying the initial establishment of bacterial populations within the gut. We demonstrate that loss of RpoS significantly decreases the ability of ECOR2 to colonize HIOs, though it does not prevent colonization of germ-free mice. Rather, the ∆rpoS ECOR2 exhibits a fitness defect in the germ-free mouse intestine only in the context of microbial competition. These results indicate that HIOs pose a differentially restrictive luminal environment to E. coli during colonization, thus increasing our understanding of the HIO model system as it pertains to studying the establishment of intestinal host-microbe symbioses.ImportanceTechnological advancements have and will continue to drive the adoption of organoid-based systems for investigating host-microbe interactions within the human intestinal ecosystem. Using E. coli deficient in the RpoS-mediated general stress response, we demonstrate that the type or severity of microbial stressors within the HIO lumen differ from those of the in vivo environment of the germ-free mouse gut. This study provides important insight into the nature of the HIO microenvironment from a microbiological standpoint.

scholarly journals Genetic Dissection of Specificity Determinants in the Interaction of HPr with Enzymes II of the Bacterial Phosphoenolpyruvate:Sugar Phosphotransferase System in Escherichia coli

2007 ◽  
Vol 189 (13) ◽  
pp. 4603-4613 ◽  
Author(s):  
Birte Reichenbach ◽  
Daniel A. Breustedt ◽  
Jörg Stülke ◽  
Bodo Rak ◽  
Boris Görke
Keyword(s):  
Escherichia Coli ◽  
Hydrophobic Interactions ◽  
Regulatory Protein ◽  
Sugar Transport ◽  
Salt Bridges ◽  
Phosphotransferase System ◽  
Homologous Proteins ◽  
Gram Positive ◽  
E Coli

ABSTRACT The histidine protein (HPr) is the energy-coupling protein of the phosphoenolpyruvate (PEP)-dependent carbohydrate:phosphotransferase system (PTS), which catalyzes sugar transport in many bacteria. In its functions, HPr interacts with a number of evolutionarily unrelated proteins. Mainly, it delivers phosphoryl groups from enzyme I (EI) to the sugar-specific transporters (EIIs). HPr proteins of different bacteria exhibit almost identical structures, and, where known, they use similar surfaces to interact with their target proteins. Here we studied the in vivo effects of the replacement of HPr and EI of Escherichia coli with the homologous proteins from Bacillus subtilis, a gram-positive bacterium. This replacement resulted in severe growth defects on PTS sugars, suggesting that HPr of B. subtilis cannot efficiently phosphorylate the EIIs of E. coli. In contrast, activation of the E. coli BglG regulatory protein by HPr-catalyzed phosphorylation works well with the B. subtilis HPr protein. Random mutations were introduced into B. subtilis HPr, and a screen for improved growth on PTS sugars yielded amino acid changes in positions 12, 16, 17, 20, 24, 27, 47, and 51, located in the interaction surface of HPr. Most of the changes restore intermolecular hydrophobic interactions and salt bridges normally formed by the corresponding residues in E. coli HPr. The residues present at the targeted positions differ between HPrs of gram-positive and -negative bacteria, but within each group they are highly conserved. Therefore, they may constitute a signature motif that determines the specificity of HPr for either gram-negative or -positive EIIs.

scholarly journals Examination of the Genome-Wide Transcriptional Response of Escherichia coli O157:H7 to Cinnamaldehyde Exposure

2012 ◽  
Vol 79 (3) ◽  
pp. 942-950 ◽  
Author(s):  
Jeyachchandran Visvalingam ◽  
Juan David Hernandez-Doria ◽  
Richard A. Holley
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Hplc Analysis ◽  
Expression Profiles ◽  
Transcriptional Analysis ◽  
Content Type ◽  
E Coli ◽  
Rp Hplc ◽  
Genome Wide ◽  
Cinnamic Alcohol

ABSTRACTCinnamaldehyde is a natural antimicrobial that has been found to be effective against many food-borne pathogens, includingEscherichia coliO157:H7. Although its antimicrobial effects have been well investigated, limited information is available on its effects at the molecular level. Sublethal treatment at 200 mg/liter cinnamaldehyde inhibited growth ofE. coliO157:H7 at 37°C and for ≤2 h caused cell elongation, but from 2 to 4 h growth resumed and cells reverted to normal length. To understand this transient behavior, genome-wide transcriptional analysis ofE. coliO157:H7 was performed at 2 and 4 h of exposure to cinnamaldehyde in conjunction with reverse-phase high-performance liquid chromatography (RP-HPLC) analysis for cinnamaldehyde and other cinnamic compounds. Drastically different gene expression profiles were obtained at 2 and 4 h. RP-HPLC analysis showed that cinnamaldehyde was structurally stable for at least 2 h. At 2 h of exposure, cinnamaldehyde induced expression of many oxidative stress-related genes and repressed expression of DNA, protein, O-antigen, and fimbrial synthetic genes. At 4 h, many cinnamaldehyde-induced repressive effects onE. coliO157:H7 gene expression were reversed, and cells became more motile and grew at a slightly higher rate. Data indicated that by 4 h,E. coliO157:H7 was able to convert cinnamaldehyde into the less toxic cinnamic alcohol using dehydrogenase/reductase enzymes (YqhD and DkgA). This is the first study to characterize the ability ofE. coliO157:H7 to convert cinnamaldehyde into cinnamic alcohol which, in turn, showed that the antimicrobial activity of cinnamaldehyde is mainly attributable to its carbonyl aldehyde group.

scholarly journals Characterization and in Vivo Cloning of prlC, a Suppressor of Signal Sequence Mutations in Escherichia coli K12

Genetics ◽  
1987 ◽  
Vol 116 (4) ◽  
pp. 513-521
Author(s):  
Nancy J Trun ◽  
Thomas J Silhavy
Keyword(s):  
Escherichia Coli ◽  
Genetic Mapping ◽  
Signal Sequence ◽  
Illegitimate Recombination ◽  
Mutant Phenotype ◽  
E Coli ◽  
Physical Location ◽  
Sequence Deletion ◽  
New Alleles

ABSTRACT The prlC gene of E. coli was originally identified as an allele, prlC1, which suppresses certain signal sequence mutations in the genes for several exported proteins. We have isolated six new alleles of prlC that also confer this phenotype. These mutations can be placed into three classes based on the degree to which they suppress the lamBsignal sequence deletion, lamBs78. Genetic mapping reveals that the physical location of the mutations in prlC correlates with the strength of the suppression, suggesting that different regions of the gene can be altered to yield a suppressor phenotype. We also describe an in vivo cloning procedure using λplacMu9H. The procedure relies on transposition and illegitimate recombination to generate a specialized transducing phage that carries prlC1. This method should be applicable to any gene for which there is a mutant phenotype.

scholarly journals Mechanistic insights into synergy between nalidixic acid and tetracycline against clinical isolates of Acinetobacter baumannii and Escherichia coli

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Amit Gaurav ◽  
Varsha Gupta ◽  
Sandeep K. Shrivastava ◽  
Ranjana Pathania
Keyword(s):  
Escherichia Coli ◽  
Acinetobacter Baumannii ◽  
Nalidixic Acid ◽  
Bacterial Infections ◽  
Clinical Isolates ◽  
Hospital Environment ◽  
Infection Model ◽  
Drug Resistant ◽  
E Coli

AbstractThe increasing prevalence of antimicrobial resistance has become a global health problem. Acinetobacter baumannii is an important nosocomial pathogen due to its capacity to persist in the hospital environment. It has a high mortality rate and few treatment options. Antibiotic combinations can help to fight multi-drug resistant (MDR) bacterial infections, but they are rarely used in the clinics and mostly unexplored. The interaction between bacteriostatic and bactericidal antibiotics are mostly reported as antagonism based on the results obtained in the susceptible model laboratory strain Escherichia coli. However, in the present study, we report a synergistic interaction between nalidixic acid and tetracycline against clinical multi-drug resistant A. baumannii and E. coli. Here we provide mechanistic insight into this dichotomy. The synergistic combination was studied by checkerboard assay and time-kill curve analysis. We also elucidate the mechanism behind this synergy using several techniques such as fluorescence spectroscopy, flow cytometry, fluorescence microscopy, morphometric analysis, and real-time polymerase chain reaction. Nalidixic acid and tetracycline combination displayed synergy against most of the MDR clinical isolates of A. baumannii and E. coli but not against susceptible isolates. Finally, we demonstrate that this combination is also effective in vivo in an A. baumannii/Caenorhabditis elegans infection model (p < 0.001)

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