scholarly journals The BarA-UvrY Two-Component System Regulates Virulence in Avian Pathogenic Escherichia coli O78:K80:H9

2006 ◽  
Vol 74 (8) ◽  
pp. 4900-4909 ◽  
Author(s):  
Christopher D. Herren ◽  
Arindam Mitra ◽  
Senthil Kumar Palaniyandi ◽  
Adam Coleman ◽  
Subbiah Elankumaran ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Chicken Embryo ◽  
Phenotypic Traits ◽  
Virulence Determinants ◽  
Two Component System ◽  
Component System ◽  
Two Component ◽  
Embryo Lethality

ABSTRACT The BarA-UvrY two-component system (TCS) in Escherichia coli is known to regulate a number of phenotypic traits. Both in vitro and in vivo assays, including the chicken embryo lethality assay, showed that this TCS regulates virulence in avian pathogenic E. coli (APEC) serotype O78:K80:H9. A number of virulence determinants, such as the abilities to adhere, invade, persist within tissues, survive within macrophages, and resist bactericidal effects of serum complement, were compromised in mutants lacking either the barA or uvrY gene. The reduced virulence was attributed to down regulation of type 1 and Pap fimbriae, reduced exopolysaccharide production, and increased susceptibility to oxidative stress. Our results indicate that BarA-UvrY regulates virulence properties in APEC and that the chicken embryo lethality assay can be used as a surrogate model to determine virulence determinants and their regulation in APEC strains.

scholarly journals Quantitative Kinetic Analyses of Shutting Off a Two-Component System

mBio ◽  
2017 ◽  
Vol 8 (3) ◽  
Author(s):  
Rong Gao ◽  
Ann M. Stock
Keyword(s):  
Phosphatase Activity ◽  
Response Regulator ◽  
Two Component System ◽  
Component System ◽  
Signaling Systems ◽  
Cellular Environment ◽  
Two Component ◽  
The Impact

ABSTRACT Cells rely on accurate control of signaling systems to adapt to environmental perturbations. System deactivation upon stimulus removal is as important as activation of signaling pathways. The two-component system (TCS) is one of the major bacterial signaling schemes. In many TCSs, phosphatase activity of the histidine kinase (HK) is believed to play an essential role in shutting off the pathway and resetting the system to the prestimulus state. Two basic challenges are to understand the dynamic behavior of system deactivation and to quantitatively evaluate the role of phosphatase activity under natural cellular conditions. Here we report a kinetic analysis of the response to shutting off the archetype Escherichia coli PhoR-PhoB TCS pathway using both transcription reporter assays and in vivo phosphorylation analyses. Upon removal of the stimulus, the pathway is shut off by rapid dephosphorylation of the PhoB response regulator (RR) while PhoB-regulated gene products gradually reset to prestimulus levels through growth dilution. We developed an approach combining experimentation and modeling to assess in vivo kinetic parameters of the phosphatase activity with kinetic data from multiple phosphatase-diminished mutants. This enabled an estimation of the PhoR phosphatase activity in vivo , which is much stronger than the phosphatase activity of PhoR cytoplasmic domains analyzed in vitro . We quantitatively modeled how strong the phosphatase activity needs to be to suppress nonspecific phosphorylation in TCSs and discovered that strong phosphatase activity of PhoR is required for cross-phosphorylation suppression. IMPORTANCE Activation of TCSs has been extensively studied; however, the kinetics of shutting off TCS pathways is not well characterized. We present comprehensive analyses of the shutoff response for the PhoR-PhoB system that reveal the impact of phosphatase activity on shutoff kinetics. This allows development of a quantitative framework not only to characterize the phosphatase activity in the natural cellular environment but also to understand the requirement for specific strengths of phosphatase activity to suppress nonspecific phosphorylation. Our model suggests that the ratio of the phosphatase rate to the nonspecific phosphorylation rate correlates with TCS expression levels and the ratio of the RR to HK, which may contribute to the great diversity of enzyme levels and activities observed in different TCSs.

scholarly journals Manipulation of the Anoxic Metabolism in Escherichia coli by ArcB Deletion Variants in the ArcBA Two-Component System

2012 ◽  
Vol 78 (24) ◽  
pp. 8784-8794 ◽  
Author(s):  
Gonzalo N. Bidart ◽  
Jimena A. Ruiz ◽  
Alejandra de Almeida ◽  
Beatriz S. Méndez ◽  
Pablo I. Nikel
Keyword(s):  
Escherichia Coli ◽  
Metabolic Flux ◽  
Expression Patterns ◽  
Phenotypic Traits ◽  
Wild Type ◽  
Two Component System ◽  
Component System ◽  
Content Type ◽  
Anoxic Conditions ◽  
Two Component

ABSTRACTBioprocesses conducted under conditions with restricted O2supply are increasingly exploited for the synthesis of reduced biochemicals using different biocatalysts. The model facultative anaerobeEscherichia colihas elaborate sensing and signal transduction mechanisms for redox control in response to the availability of O2and other electron acceptors. The ArcBA two-component system consists of ArcB, a membrane-associated sensor kinase, and ArcA, the cognate response regulator. The tripartite hybrid kinase ArcB possesses a transmembrane, a PAS, a primary transmitter (H1), a receiver (D1), and a phosphotransfer (H2) domain. Metabolic fluxes were compared under anoxic conditions in a wild-typeE. colistrain, its ΔarcBderivative, and two partialarcBdeletion mutants in which ArcB lacked either the H1 domain or the PAS-H1-D1 domains. These analyses revealed that elimination of different segments in ArcB determines a distinctive distribution ofd-glucose catabolic fluxes, different from that observed in the ΔarcBbackground. Metabolite profiles, enzyme activity levels, and gene expression patterns were also investigated in these strains. Relevant alterations were observed at the P-enol-pyruvate/pyruvate and acetyl coenzyme A metabolic nodes, and the formation of reduced fermentation metabolites, such as succinate,d-lactate, and ethanol, was favored in the mutant strains to different extents compared to the wild-type strain. These phenotypic traits were associated with altered levels of the enzymatic activities operating at these nodes, as well as with elevated NADH/NAD+ratios. Thus, targeted modification of global regulators to obtain different metabolic flux distributions under anoxic conditions is emerging as an attractive tool for metabolic engineering purposes.

scholarly journals Negative Regulation of DNA Repair Gene (ung) Expression by the CpxR/CpxA Two-Component System in Escherichia coli K-12 and Induction of Mutations by Increased Expression of CpxR

2004 ◽  
Vol 186 (24) ◽  
pp. 8317-8325 ◽  
Author(s):  
Hiroshi Ogasawara ◽  
Jun Teramoto ◽  
Kiyo Hirao ◽  
Kaneyoshi Yamamoto ◽  
Akira Ishihama ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
In Vitro Transcription ◽  
Negative Regulation ◽  
Two Component System ◽  
Repair Gene ◽  
Component System ◽  
Dna Repair Gene ◽  
Two Component ◽  
K 12

ABSTRACT In Escherichia coli K-12 overexpressing CpxR, transcription of the ung gene for uracil-DNA glycosylase was repressed, ultimately leading to the induction of mutation. Gel shift, DNase I footprinting, and in vitro transcription assays all indicated negative regulation of ung transcription by phosphorylated CpxR. Based on the accumulated results, we conclude that ung gene expression is negatively regulated by the two-component system of CpxR/CpxA signal transduction.

Transcriptional regulation of drug efflux genes by EvgAS, a two-component system in Escherichia coli

Microbiology ◽  
2003 ◽  
Vol 149 (10) ◽  
pp. 2819-2828 ◽  
Author(s):  
Yoko Eguchi ◽  
Taku Oshima ◽  
Hirotada Mori ◽  
Rikizo Aono ◽  
Kaneyoshi Yamamoto ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multi Drug Resistance ◽  
Drug Efflux ◽  
Two Component System ◽  
Component System ◽  
Two Component ◽  
Outer Membrane Channel ◽  
The Relationship ◽  
Mdr Genes

A constitutively active mutant of histidine kinase sensor EvgS was found to confer multi-drug resistance (MDR) to an acrA-deficient Escherichia coli, indicating the relationship between the two-component system EvgAS and the expression of the MDR system. The observed MDR also depended on an outer-membrane channel, TolC. Microarray and S1 mapping assays indicated that, in the presence of this constitutive mutant EvgS, the level of transcription increased for some MDR genes, including the drug efflux genes emrKY, yhiUV, acrAB, mdfA and tolC. Transcription in vitro of emrK increased by the addition of phosphorylated EvgA. Transcription activation of tolC by the activated EvgS was, however, dependent on both EvgAS and PhoPQ (Mg2+-responsive two-component system), in agreement with the presence of the binding site (PhoP box) for the regulator PhoP in the tolC promoter region. Transcription in vitro of yhiUV also appears to require an as-yet-unidentified additional transcriptional factor besides EvgA. Taken together we propose that the expression of the MDR system is under a complex regulatory network, including the phosphorylated EvgA serving as the master regulator.

scholarly journals MprAB Is a Stress-Responsive Two-Component System That Directly Regulates Expression of Sigma Factors SigB and SigE in Mycobacterium tuberculosis

2006 ◽  
Vol 188 (6) ◽  
pp. 2134-2143 ◽  
Author(s):  
Hongjun He ◽  
Raymond Hovey ◽  
Jason Kane ◽  
Vineet Singh ◽  
Thomas C. Zahrt
Keyword(s):  
Signal Transduction ◽  
Mycobacterium Tuberculosis ◽  
Sigma Factors ◽  
Two Component System ◽  
Direct Role ◽  
Component System ◽  
Two Component ◽  
Two Component Signal Transduction

ABSTRACT The genetic mechanisms mediating the adaptation of Mycobacterium tuberculosis within the host are poorly understood. The best-characterized regulatory systems in this organism include sigma factors and two-component signal transduction systems. mprAB is a two-component system required by M. tuberculosis for growth in vivo during the persistent stage of infection. In this report, we demonstrate that MprAB is stress responsive and regulates the expression of numerous stress-responsive genes in M. tuberculosis. With DNA microarrays and quantitative real-time reverse transcription-PCR, genes regulated by MprA in M. tuberculosis that included two stress-responsive sigma factors were identified. Response regulator MprA bound to conserved motifs in the upstream regions of both sigB and sigE in vitro and regulated the in vivo expression of sigB and sigE in M. tuberculosis. In addition, mprA itself was induced following exposure to stress, establishing a direct role for this regulatory system in stress response pathways of M. tuberculosis. Induction of mprA and sigE by MprA in response to stress was mediated through the cognate sensor kinase MprB and required expression of the extracytoplasmic loop domain. These results provide the first evidence that recognition of and adaptation to specific stress in M. tuberculosis are mediated through activation of a two-component signal transduction system that directly regulates the expression of stress-responsive determinants.

The Campylobacter jejuni dccRS two-component system is required for optimal in vivo colonization but is dispensable for in vitro growth

2004 ◽  
Vol 54 (5) ◽  
pp. 1269-1286 ◽  
Author(s):  
Joanna K. MacKichan ◽  
Erin C. Gaynor ◽  
Christopher Chang ◽  
Shaun Cawthraw ◽  
Diane G. Newell ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
In Vitro Growth ◽  
Two Component System ◽  
Component System ◽  
Two Component

scholarly journals Evidence of Robustness in a Two-Component System Using a Synthetic Circuit

2019 ◽  
Vol 202 (4) ◽  
Author(s):  
Arkajyoti Dutta ◽  
Paulami Rudra ◽  
Suman Kumar Banik ◽  
Jayanta Mukhopadhyay
Keyword(s):  
Signaling Pathway ◽  
Regulatory Networks ◽  
Sensor Kinase ◽  
Two Component System ◽  
Component System ◽  
Content Type ◽  
Two Component ◽  
Synthetic Circuit

ABSTRACT Variation in the concentration of biological components is inescapable for any cell. Robustness in any biological circuit acts as a cushion against such variation and enables the cells to produce homogeneous output despite the fluctuation. The two-component system (TCS) with a bifunctional sensor kinase (that possesses both kinase and phosphatase activities) is proposed to be a robust circuit. Few theoretical models explain the robustness of a TCS, although the criteria and extent of robustness by these models differ. Here, we provide experimental evidence to validate the extent of the robustness of a TCS signaling pathway. We have designed a synthetic circuit in Escherichia coli using a representative TCS of Mycobacterium tuberculosis, MprAB, and monitored the in vivo output signal by systematically varying the concentration of either of the components or both. We observed that the output of the TCS is robust if the concentration of MprA is above a threshold value. This observation is further substantiated by two in vitro assays, in which we estimated the phosphorylated MprA pool or MprA-dependent transcription yield by varying either of the components of the TCS. This synthetic circuit could be used as a model system to analyze the relationship among different components of gene regulatory networks. IMPORTANCE Robustness in essential biological circuits is an important feature of the living organism. A few pieces of evidence support the existence of robustness in vivo in the two-component system (TCS) with a bifunctional sensor kinase (SK). The assays were done under physiological conditions in which the SK was much lower than the response regulator (RR). Here, using a synthetic circuit, we varied the concentrations of the SK and RR of a representative TCS to monitor output robustness in vivo. In vitro assays were also performed under conditions where the concentration of the SK was greater than that of the RR. Our results demonstrate the extent of output robustness in the TCS signaling pathway with respect to the concentrations of the two components.

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