scholarly journals Escherichia coliLrp regulates one-third of the genome via direct, cooperative, and indirect routes

2018 ◽  
Author(s):  
Grace M. Kroner ◽  
Michael B. Wolfe ◽  
Peter L. Freddolino
Keyword(s):  
Escherichia Coli ◽  
Crucial Role ◽  
Regulatory Activity ◽  
Global Regulator ◽  
Full Spectrum ◽  
Global Trends ◽  
Cooperative Action ◽  
E Coli ◽  
Physiological Conditions ◽  
Regulatory Behavior

AbstractThe global regulator Lrp plays a crucial role in regulating metabolism, virulence and motility in response to environmental conditions. Lrp has previously been shown to activate or repress approximately 10% of genes inEscherichia coli. However, the full spectrum of targets, and how Lrp acts to regulate them, has stymied earlier study. We have combined matched ChIP-seq and RNA sequencing under nine physiological conditions to map the binding and regulatory activity of Lrp as it directs responses to nutrient abundance. In addition to identifying hundreds of novel Lrp targets, we observe two new global trends: first, that Lrp will often bind to promoters in a poised position under conditions when it has no regulatory activity, and second, that nutrient levels induce a global shift in the equilibrium between non-specific and sequence-specific DNA binding. The overall regulatory behavior of Lrp, which as we now show regulates 35% ofE. coligenes directly or indirectly under at least one condition, thus arises from the interaction between changes in Lrp binding specificity and cooperative action with other regulators.

scholarly journals Escherichia coliLrp Regulates One-Third of the Genome via Direct, Cooperative, and Indirect Routes

2018 ◽  
Vol 201 (3) ◽  
Author(s):  
Grace M. Kroner ◽  
Michael B. Wolfe ◽  
Peter L. Freddolino
Keyword(s):  
Dna Binding ◽  
Environmental Conditions ◽  
Regulatory Protein ◽  
Growth Stages ◽  
Regulatory Activity ◽  
Full Spectrum ◽  
Content Type ◽  
Global Trends ◽  
E Coli ◽  
Nutrient Levels

ABSTRACTThe global regulator Lrp plays a crucial role in regulating metabolism, virulence, and motility in response to environmental conditions. Lrp has previously been shown to activate or repress approximately 10% of the genes inEscherichia coli. However, the full spectrum of targets, and how Lrp acts to regulate them, have stymied earlier study. We have combined matched chromatin-immunoprecipitation sequencing (ChIP-seq) and RNA sequencing (RNA-seq) under nine physiological conditions to comprehensively map the binding and regulatory activity of Lrp as it directs responses to nutrient abundance. In addition to identifying hundreds of novel Lrp targets, we observe two new global trends, as follows: first, that Lrp will often bind to promoters in a poised position under conditions when it has no regulatory activity to enable combinatorial interactions with other regulators, and second, that nutrient levels induce a global shift in the equilibrium between less-sequence-specific and more-sequence-specific DNA binding. The overall regulatory behavior of Lrp, which as we now show extends to 38% ofE. coligenes directly or indirectly under at least one condition, thus arises from the interaction between changes in Lrp binding specificity and cooperative action with other regulators.IMPORTANCETo survive, bacteria such asE. colimust rapidly respond to changing environmental conditions, including nutrient levels. A decrease in nutrient availability causes bacteria to stop rapid replication and enter stationary phase, where they perform limited to no cell division. TheE. coliglobal regulatory protein Lrp has been previously implicated in modulating the expression of genes particularly important at this transition from rapid to slowed growth. Here, we monitor Lrp’s DNA binding locations and effect on gene expression under three different nutrient conditions across three growth stages. We find that Lrp’s role is even broader than previously suspected and that it appears to interact with many other bacterial regulators to perform its function in a condition-specific manner.

scholarly journals Crucial Role of ppGpp in the Resilience of Escherichia coli to Growth Disruption

mSphere ◽  
2020 ◽  
Vol 5 (6) ◽  
pp. e01132-20
Author(s):  
Clément Patacq ◽  
Nicolas Chaudet ◽  
Fabien Létisse
Keyword(s):  
Escherichia Coli ◽  
Bacterial Growth ◽  
Crucial Role ◽  
Stringent Response ◽  
Content Type ◽  
E Coli ◽  
Changing Environments ◽  
Growth Recovery ◽  
Growth Disruption

ABSTRACTBacteria grow in constantly changing environments that can suddenly become completely depleted of essential nutrients. The stringent response, a rewiring of the cellular metabolism mediated by the alarmone (p)ppGpp, plays a crucial role in adjusting bacterial growth to the severity of the nutritional stress. The ability of (p)ppGpp to trigger a slowdown of cell growth or induce bacterial dormancy has been widely investigated. However, little is known about the role of (p)ppGpp in promoting growth recovery after severe growth inhibition. In this study, we performed a time-resolved analysis of (p)ppGpp metabolism in Escherichia coli as it recovered from a sudden slowdown in growth. The results show that E. coli recovers by itself from the growth disruption provoked by the addition of serine hydroxamate, the serine analogue that we used to induce the stringent response. Growth inhibition was accompanied by a severe disturbance of metabolic activity and, more surprisingly, a transient overflow of valine and alanine. Our data also show that ppGpp is crucial for growth recovery since in the absence of ppGpp, E. coli’s growth recovery was slower. In contrast, an increased concentration of pppGpp was found to have no significant effect on growth recovery. Interestingly, the observed decrease in intracellular ppGpp levels in the recovery phase correlated with bacterial growth, and the main effect involved in the return to the basal level was identified by flux calculation as growth dilution. This report thus significantly expands our knowledge of (p)ppGpp metabolism in E. coli physiology.IMPORTANCE The capacity of microbes to resist and overcome environmental insults, known as resilience, allows them to survive in changing environments but also to resist antibiotic and biocide treatments and immune system responses. Although the role of the stringent response in bacterial resilience to nutritional stresses has been well studied, little is known about its importance in the ability of the bacteria to not just resist but also recover from these disturbances. To address this important question, we investigated growth disruption resilience in the model bacterium Escherichia coli and its dependence on the stringent response alarmone (p)ppGpp by quantifying ppGpp and pppGpp levels as growth was disrupted and then recovered. Our findings may thus contribute to understanding how ppGpp improves E. coli’s resilience to nutritional stress and other environmental insults.

scholarly journals Crucial role of ppGpp in the resilience of Escherichia coli to growth disruption

2020 ◽  
Author(s):  
Clément Patacq ◽  
Nicolas Chaudet ◽  
Fabien Letisse
Keyword(s):  
Escherichia Coli ◽  
Growth Inhibition ◽  
Bacterial Growth ◽  
Crucial Role ◽  
Stringent Response ◽  
E Coli ◽  
Changing Environments ◽  
Growth Recovery ◽  
Growth Disruption

ABSTRACTBacteria grow in constantly changing environments that can suddenly become completely deleted in essential nutrients. The stringent response, a rewiring of the cellular metabolism mediated by the alarmone (p)ppGpp, plays a crucial role in adjusting bacterial growth to the severity of the nutritional insult. The ability of (p)ppGpp to trigger a slowdown of cell growth or induce bacterial dormancy has been widely investigated. However, little is known about the role of (p)ppGpp in promoting growth recovery after severe growth inhibition. In this study, we performed a time-resolved analysis of (p)ppGpp metabolism in Escherichia coli as it recovered from a sudden slowdown in growth. Results show that E. coli recovers by itself from the growth disruption provoked by the addition of serine hydroxamate, the serine analogue that we used to induce the stringent response. Growth inhibition was accompanied by a severe disturbance of metabolic activity and more surprisingly, by a transient overflow of valine and alanine. Our data also show that ppGpp is crucial for growth recovery since in the absence of ppGpp, E. coli’s growth recovery was slower. In contrast, an increased concentration of pppGpp was found to have no significant effect on growth recovery. Interestingly, the observed decrease in intracellular ppGpp levels in the recovery phase correlated with bacterial growth and the main effect involved was identified as growth dilution rather than active degradative process. This report thus significantly expands our knowledge of (p)ppGpp metabolism in E. coli physiology.IMPORTANCEThe capacity of microbes to resist and overcome environmental insults, know as resilience, allows them to survive in changing environments but also to resist antibiotic and biocide treatments, immune system responses. Although the role of the stringent response in bacterial resilience to nutritional insults has been well studied, little is known about its importance in the ability of the bacteria to not just resist but also recover from these disturbances. To address this important question, we investigated growth disruption resilience in the model bacterium Escherichia coli and its dependency on the stringent response alarmone (p)ppGpp by quantifying ppGpp and pppGpp levels as growth was disrupted and then recovered. Our findings may thus contribute to understanding how ppGpp improves E. coli’s resilience to nutritional stress and other environmental insults.

scholarly journals Inactivation of Transcriptional Regulators during Within-Household Evolution of Escherichia coli

2017 ◽  
Vol 199 (13) ◽  
Author(s):  
Dagmara I. Kisiela ◽  
Matthew Radey ◽  
Sandip Paul ◽  
Stephen Porter ◽  
Kseniya Polukhina ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Adaptive Dynamics ◽  
Transcriptional Regulators ◽  
Index Patient ◽  
Transcriptional Analysis ◽  
Global Regulator ◽  
Blood Isolate ◽  
Content Type ◽  
E Coli ◽  
Sepsis Model

ABSTRACT We analyzed the within-household evolution of two household-associated Escherichia coli strains from pandemic clonal group ST131-H30, using isolates recovered from five individuals within two families, each of which had a distinct strain. Family 1's strain was represented by a urine isolate from the index patient (older sister) with recurrent cystitis and a blood isolate from her younger sister with fatal urosepsis. Family 2's strain was represented by a urine isolate from the index patient (father) with pyelonephritis and renal abscesses, blood and kidney drainage isolates from the daughter with emphysematous pyelonephritis, and urine and fecal isolates from the mother with cystitis. Collectively, the several variants of each family's strain had accumulated a total of 8 (family 1) and 39 (family 2) point mutations; no two isolates were identical. Of the 47 total mutations, 36 resulted in amino acid changes or truncation of coded proteins. Fourteen such mutations (39%) targeted genes encoding transcriptional regulators, and 9 (25%) involved DNA-binding transcription factors (TFs), which significantly exceeded the relative contribution of TF genes to the isolates' genomes (∼6%). At least one-half of the transcriptional regulator mutations were inactivating, based on phenotypic and/or transcriptional analysis. In particular, inactivating mutations in the global regulator LrhA (repressor of type 1 fimbriae and flagella) occurred in the blood isolates from both households and increased the virulence of E. coli strains in a murine sepsis model. The results indicate that E. coli undergoes adaptive evolution between and/or within hosts, generating subpopulations with distinctive phenotypes and virulence potential. IMPORTANCE The clonal evolution of bacterial strains associated with interhost transmission is poorly understood. We characterized the genome sequences of clonal descendants of two Escherichia coli strains, recovered at different time points from multiple individuals within two households who had different types of urinary tract infection. We found evidence that the E. coli strains underwent extensive mutational diversification between and within these individuals, driven disproportionately by inactivation of transcriptional regulators. In urosepsis isolates, the mutations observed in the global regulator LrhA increased bacterial virulence in a murine sepsis model. Our findings help in understanding the adaptive dynamics and strategies of E. coli during short-term natural evolution.

scholarly journals Control and regulation of acetate overflow in Escherichia coli

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Pierre Millard ◽  
Brice Enjalbert ◽  
Sandrine Uttenweiler-Joseph ◽  
Jean-Charles Portais ◽  
Fabien Létisse
Keyword(s):  
Escherichia Coli ◽  
Glucose Metabolism ◽  
Kinetic Model ◽  
Local Control ◽  
Overflow Metabolism ◽  
Central Metabolism ◽  
Global Regulator ◽  
E Coli ◽  
By Products ◽  
Acetate Overflow

Overflow metabolism refers to the production of seemingly wasteful by-products by cells during growth on glucose even when oxygen is abundant. Two theories have been proposed to explain acetate overflow in Escherichia coli – global control of the central metabolism and local control of the acetate pathway – but neither accounts for all observations. Here, we develop a kinetic model of E. coli metabolism that quantitatively accounts for observed behaviours and successfully predicts the response of E. coli to new perturbations. We reconcile these theories and clarify the origin, control, and regulation of the acetate flux. We also find that, in turns, acetate regulates glucose metabolism by coordinating the expression of glycolytic and TCA genes. Acetate should not be considered a wasteful end-product since it is also a co-substrate and a global regulator of glucose metabolism in E. coli. This has broad implications for our understanding of overflow metabolism.

scholarly journals Multiple functions of l0036 in the regulation of the pathogenicity island of enterohaemorrhagic Escherichia coli O157:H7

2005 ◽  
Vol 393 (2) ◽  
pp. 591-599 ◽  
Author(s):  
Nien-Pei Tsai ◽  
Yi-Chih Wu ◽  
Jenn-Wei Chen ◽  
Chih-Feng Wu ◽  
Chi-Meng Tzeng ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Current Knowledge ◽  
Regulatory Gene ◽  
Pathogenicity Island ◽  
Negative Regulator ◽  
Multiple Point ◽  
Global Regulator ◽  
Multiple Functions ◽  
E Coli ◽  
Enterohaemorrhagic Escherichia Coli

Diarrhoeagenic enterohaemorrhagic Escherichia coli and enteropathogenic E. coli attach to human intestinal epithelium and efface brush-border microvilli, forming an A/E (attaching and effacing) lesion. These human pathogens are phenotypically similar to the mouse pathogen Citrobacter rodentium. Genetically, they all have a homologous set of virulent genes involved in the A/E lesion, and these genes are organized on a LEE (locus of enterocyte effacement), a pathogenicity island. This island comprises 41 specific open reading frames, of which most are organized at five operons, LEE1, LEE2, LEE3, LEE4 and tir (LEE5). The expression of the LEE genes is regulated in a complicated manner, and current knowledge is that there are at least two positive regulators, Ler (LEE-encoded regulator) and GrlA (global regulator of LEE activator), and one negative regulator, called GrlR (global regulator of LEE repressor). In enterohaemorrhagic E. coli, GrlA is encoded by l0043, whereas GrlR is encoded by l0044. Here we report a fourth regulatory gene located in LEE3, namely l0036. Its expression is tightly controlled. When overexpressed, this factor, named Mpc (multiple point controller), interacts with Ler and suppresses the expression of the LEE proteins. When the translation is not initiated or terminated before maturation, the type III secretion of effectors is completely abolished. Therefore, together with the fact that several cis elements reside in the region that l0036 spans, l0036 appeared to have multiple functions in the regulation of LEE expression.

scholarly journals pSM19035-encoded ζ toxin induces stasis followed by death in a subpopulation of cells

Microbiology ◽  
2006 ◽  
Vol 152 (8) ◽  
pp. 2365-2379 ◽  
Author(s):  
Virginia S. Lioy ◽  
M. Teresa Martín ◽  
Ana G. Camacho ◽  
Rudi Lurz ◽  
Haike Antelmann ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Proliferation ◽  
Minimal Medium ◽  
Time Window ◽  
Constitutive Expression ◽  
Protein Translation ◽  
Cellular Level ◽  
Global Regulator ◽  
E Coli ◽  
Toxin Accumulation

The toxin–antitoxin operon of pSM19035 encodes three proteins: the ω global regulator, the ε labile antitoxin and the stable ζ toxin. Accumulation of ζ toxin free of ε antitoxin induced loss of cell proliferation in both Bacillus subtilis and Escherichia coli cells. Induction of a ζ variant (ζY83C) triggered stasis, in which B. subtilis cells were viable but unable to proliferate, without selectively affecting protein translation. In E. coli cells, accumulation of free ζ toxin induced stasis, but this was fully reversed by expression of the ε antitoxin within a defined time window. The time window for reversion of ζ toxicity by expression of ε antitoxin was dependent on the initial cellular level of ζ. After 240 min of constitutive expression, or inducible expression of high levels of ζ toxin for 30 min, expression of ε failed to reverse the toxic effect exerted by ζ in cells growing in minimal medium. Under the latter conditions, ζ inhibited replication, transcription and translation and finally induced death in a fraction (∼50 %) of the cell population. These results support the view that ζ interacts with its specific target and reversibly inhibits cell proliferation, but accumulation of ζ might lead to cell death due to pleiotropic effects.

scholarly journals Characterization of DNA-binding specificity and analysis of binding sites of the Pseudomonas aeruginosa global regulator, Vfr, a homologue of the Escherichia coli cAMP receptor protein

Microbiology ◽  
2006 ◽  
Vol 152 (12) ◽  
pp. 3485-3496 ◽  
Author(s):  
Kristen J. Kanack ◽  
Laura J. Runyen-Janecky ◽  
Evan P. Ferrell ◽  
Sang-Jin Suh ◽  
Susan E. H. West
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Receptor Protein ◽  
Camp Receptor Protein ◽  
Global Regulator ◽  
E Coli ◽  
Consensus Binding Sequence ◽  
Target Sequences ◽  
Camp Receptor ◽  
Binding Sequence

Vfr, a global regulator of Pseudomonas aeruginosa virulence factors, is a homologue of the Escherichia coli cAMP receptor protein, CRP. Vfr is 91 % similar to CRP and maintains many residues important for CRP to bind cAMP, bind DNA, and interact with RNA polymerase at target promoters. While vfr can complement an E. coli crp mutant in β-galactosidase production, tryptophanase production and catabolite repression, crp can only complement a subset of Vfr-dependent phenotypes in P. aeruginosa. Using specific CRP binding site mutations, it is shown that Vfr requires the same nucleotides as CRP for optimal transcriptional activity from the E. coli lac promoter. In contrast, CRP did not bind Vfr target sequences in the promoters of the toxA and regA genes. Footprinting analysis revealed Vfr protected sequences upstream of toxA, regA, and the quorum sensing regulator lasR, that are similar to but significantly divergent from the CRP consensus binding sequence, and Vfr causes similar DNA bending to CRP in bound target sequences. Using a preliminary Vfr consensus binding sequence deduced from the Vfr-protected sites, Vfr target sequences were identified upstream of the virulence-associated genes plcN, plcHR, pbpG, prpL and algD, and in the vfr/orfX, argH/fimS, pilM/ponA intergenic regions. From these sequences the Vfr consensus binding sequence, 5′-ANWWTGNGAWNY : AGWTCACAT-3′, was formulated. This study suggests that Vfr shares many of the same functions as CRP, but has specialized functions, at least in terms of DNA target sequence binding, required for regulation of a subset of genes in its regulon.

scholarly journals Molecular Characterization of Human Atypical Sorbitol-Fermenting Enteropathogenic Escherichia coli O157 Reveals High Diversity

2016 ◽  
Vol 54 (5) ◽  
pp. 1357-1363 ◽  
Author(s):  
Annelene Kossow ◽  
Wenlan Zhang ◽  
Martina Bielaszewska ◽  
Sophie Rhode ◽  
Kevin Hansen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Clinical Symptoms ◽  
Diverse Group ◽  
Full Spectrum ◽  
Content Type ◽  
E Coli ◽  
Uremic Syndrome ◽  
Encoding Gene ◽  
Encoding Genes

Alongside the well-characterized enterohemorrhagicEscherichia coli(EHEC) O157:H7, serogroup O157 comprises sorbitol-fermenting typical and atypical enteropathogenicE. coli(EPEC/aEPEC) strains that carry the intimin-encoding geneeaebut not Shiga toxin-encoding genes (stx). Since little is known about these pathogens, we characterized 30 clinical isolates from patients with hemolytic uremic syndrome (HUS) or uncomplicated diarrhea with respect to their flagellin gene (fliC) type and multilocus sequence type (MLST). Moreover, we applied whole-genome sequencing (WGS) to determine the phylogenetic relationship with othereae-positive EHEC serotypes and the composition of therfbO157 region.fliCtyping resulted in fivefliCtypes (H7, H16, H34, H39, and H45). Isolates of eachfliCtype shared a unique ST. In comparison to the 42 HUS-associatedE. coli(HUSEC) strains, only thestx-negative isolates withfliCH7 shared their ST with EHEC O157:H7/H−strains. With the exception of one O157:H−fliCH16isolate, HUS was exclusively associated withfliCH7. WGS corroborated the separation of thefliCH7 isolates, which were closely related to the EHEC O157:H7/H−isolates, and the diverse group of isolates exhibiting differentfliCtypes, indicating independent evolution of the different serotypes. This was also supported by the heterogeneity within therfbO157 region that exhibited extensive recombinations. The genotypic subtypes and distribution of clinical symptoms suggested that thestx-negative O157 strains withfliCH7 were originally EHEC strains that loststx. The remaining isolates form a distinct and diverse group of atypical EPEC isolates that do not possess the full spectrum of virulence genes, underlining the importance of identifying the H antigen for clinical risk assessment.

scholarly journals Plasmidic qnr Genes Confer Clinical Resistance to Ciprofloxacin under Urinary Tract Physiological Conditions

2017 ◽  
Vol 61 (4) ◽  
Author(s):  
Guillermo Martín-Gutiérrez ◽  
José Manuel Rodríguez-Martínez ◽  
Álvaro Pascual ◽  
Jerónimo Rodríguez-Beltrán ◽  
Jesús Blázquez
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract Infection ◽  
Urinary Tract ◽  
Susceptibility Testing ◽  
Urine Ph ◽  
Content Type ◽  
E Coli ◽  
Physiological Conditions ◽  
Ph Values ◽  
Clinical Resistance

ABSTRACT Escherichia coli variants expressing plasmid-mediated qnr genes are usually susceptible to fluoroquinolones by standard susceptibility testing. Here we show that, under specific urinary tract physiological conditions, susceptible laboratory and clinical strains harboring qnr determinants become fully resistant to ciprofloxacin (CIP). Therefore, physiological conditions, mainly urine pH values, should be considered when performing susceptibility testing of CIP activity against E. coli in treating urinary tract infection (UTI) and for selecting appropriate antibiotics for UTI treatment.

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