Identification and Molecular Characterization of the Mg2+ Stimulon of Escherichia coli

ABSTRACT Transcription profile microarray analysis in Escherichia coli was performed to identify the member genes of the Mg2+ stimulon that respond to the availability of external Mg2+ in a PhoP/PhoQ two-component system-dependent manner. The mRNA levels of W3110 in the presence of 30 mM MgCl2, WP3022 (phoP defective), and WQ3007 (phoQ defective) were compared with those of W3110 in the absence of MgCl2. The expression ratios of a total of 232 genes were <0.75 in all three strains (the supplemental data are shown at http://www.nara.kindai.ac.jp/nogei/seiken/array.html ), suggesting that the PhoP/PhoQ system is involved directly or indirectly in the transcription of these genes. Of those, 26 contained the PhoP box-like sequences with the direct repeats of (T/G)GTTTA within 500 bp upstream of the initiation codon. Furthermore, S1 nuclease assays of 26 promoters were performed to verify six new Mg2+ stimulon genes, hemL, nagA, rstAB, slyB, vboR, and yrbL, in addition to the phoPQ, mgrB, and mgtA genes reported previously. In gel shift and DNase I footprinting assays, all of these genes were found to be regulated directly by PhoP. Thus, we concluded that the phoPQ, mgrB, mgtA, hemL, nagA, rstAB, slyB, vboR, and yrbL genes make up the Mg2+ stimulon in E. coli.

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Extracellular Ca2+ transients affect poly-(R)-3-hydroxybutyrate regulation by the AtoS-AtoC system in Escherichia coli

Biochemical Journal ◽

10.1042/bj20081169 ◽

2009 ◽

Vol 417 (3) ◽

pp. 667-672 ◽

Cited By ~ 17

Author(s):

Marina C. Theodorou ◽

Ekaterini Tiligada ◽

Dimitrios A. Kyriakidis

Keyword(s):

Escherichia Coli ◽

Channel Blocker ◽

Inhibitory Effect ◽

Dependent Manner ◽

Signal Transduction System ◽

Two Component System ◽

Component System ◽

E Coli ◽

Two Component ◽

Membrane Bound

Escherichia coli is exposed to wide extracellular concentrations of Ca2+, whereas the cytosolic levels of the ion are subject to stringent control and are implicated in many physiological functions. The present study shows that extracellular Ca2+ controls cPHB [complexed poly-(R)-3-hydroxybutyrate] biosynthesis through the AtoS-AtoC two-component system. Maximal cPHB accumulation was observed at higher [Ca2+]e (extracellular Ca2+ concentration) in AtoS-AtoC-expressing E. coli compared with their ΔatoSC counterparts, in both cytosolic and membrane fractions. The reversal of EGTA-mediated down-regulation of cPHB biosynthesis by the addition of Ca2+ and Mg2+ was under the control of the AtoS-AtoC system. Moreover, the Ca2+-channel blocker verapamil reduced total and membrane-bound cPHB levels, the inhibitory effect being circumvented by Ca2+ addition only in atoSC+ bacteria. Histamine and compound 48/80 affected cPHB accumulation in a [Ca2+]e-dependent manner directed by the AtoS-AtoC system. In conclusion, these data provide evidence for the involvement of external Ca2+ on cPHB synthesis regulated by the AtoS-AtoC two-component system, thus linking Ca2+ with a signal transduction system, most probably through a transporter.

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Repression of Escherichia coli PhoP-PhoQ Signaling by Acetate Reveals a Regulatory Role for Acetyl Coenzyme A

Journal of Bacteriology ◽

10.1128/jb.185.8.2563-2570.2003 ◽

2003 ◽

Vol 185 (8) ◽

pp. 2563-2570 ◽

Cited By ~ 21

Author(s):

Joseph A. Lesley ◽

Carey D. Waldburger

Keyword(s):

Escherichia Coli ◽

Coenzyme A ◽

Inhibition Mechanism ◽

Acetate Metabolism ◽

Dependent Manner ◽

Two Component System ◽

Acetyl Coenzyme A ◽

Component System ◽

Acetyl Coenzyme ◽

Two Component

ABSTRACT The PhoP-PhoQ two-component system regulates the transcription of numerous genes in response to changes in extracellular divalent cation concentration and pH. Here we demonstrate that the Escherichia coli PhoP-PhoQ two-component system also responds to acetate. Signaling by the E. coli PhoP-PhoQ system was repressed during growth in acetate (≥25 mM) in a PhoQ-dependent manner. The periplasmic sensor domain of PhoQ was not required for acetate to repress signaling. Acetate-mediated repression of the PhoP-PhoQ system was not related to changes in the intracellular concentration of acetate metabolites such as acetyl-phosphate or acetyladenylate. Genetic analysis of acetate metabolism pathways suggested that a perturbation of acetyl coenzyme A turnover was the cause of decreased PhoP-PhoQ signaling during growth in acetate. Consistent with this hypothesis, intracellular acetyl coenzyme A levels rose during growth in the presence of exogenous acetate. Acetyl coenzyme A inhibited the autokinase activity of PhoQ in vitro, suggesting that the in vivo repressing effect may be due to a direct inhibition mechanism.

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Molecular Characterization of the PhoP-PhoQ Two-Component System in Escherichia coli K-12: Identification of Extracellular Mg2+-Responsive Promoters

Journal of Bacteriology ◽

10.1128/jb.181.17.5516-5520.1999 ◽

1999 ◽

Vol 181 (17) ◽

pp. 5516-5520 ◽

Cited By ~ 117

Author(s):

Akinori Kato ◽

Hiroyuki Tanabe ◽

Ryutaro Utsumi

Keyword(s):

Escherichia Coli ◽

Direct Repeat ◽

Two Component System ◽

Transcription Start ◽

Promoter Regions ◽

S1 Nuclease ◽

Transcription Start Sites ◽

Two Component ◽

K 12

ABSTRACT We identified Mg2+-responsive promoters of thephoPQ, mgtA, and mgrB genes ofEscherichia coli K-12 by S1 nuclease analysis. Expression of these genes was induced by magnesium limitation and depended on PhoP and PhoQ. The transcription start sites were also determined, which allowed us to find a (T/G)GTTTA direct repeat in their corresponding promoter regions.

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Kinetic Characterization of the WalRKSpn (VicRK) Two-Component System of Streptococcus pneumoniae: Dependence of WalKSpn (VicK) Phosphatase Activity on Its PAS Domain

Journal of Bacteriology ◽

10.1128/jb.01690-09 ◽

2010 ◽

Vol 192 (9) ◽

pp. 2346-2358 ◽

Cited By ~ 56

Author(s):

Alina D. Gutu ◽

Kyle J. Wayne ◽

Lok-To Sham ◽

Malcolm E. Winkler

Keyword(s):

Amino Acids ◽

Streptococcus Pneumoniae ◽

Phosphatase Activity ◽

Pas Domain ◽

Kinetic Characterization ◽

Two Component System ◽

Histidine Kinases ◽

Component System ◽

Two Component

ABSTRACT The WalRK two-component system plays important roles in maintaining cell wall homeostasis and responding to antibiotic stress in low-GC Gram-positive bacteria. In the major human pathogen, Streptococcus pneumoniae, phosphorylated WalR Spn (VicR) response regulator positively controls the transcription of genes encoding the essential PcsB division protein and surface virulence factors. WalR Spn is phosphorylated by the WalK Spn (VicK) histidine kinase. Little is known about the signals sensed by WalK histidine kinases. To gain information about WalK Spn signal transduction, we performed a kinetic characterization of the WalRK Spn autophosphorylation, phosphoryltransferase, and phosphatase reactions. We were unable to purify soluble full-length WalK Spn . Consequently, these analyses were performed using two truncated versions of WalK Spn lacking its single transmembrane domain. The longer version (Δ35 amino acids) contained most of the HAMP domain and the PAS, DHp, and CA domains, whereas the shorter version (Δ195 amino acids) contained only the DHp and CA domains. The autophosphorylation kinetic parameters of Δ35 and Δ195 WalK Spn were similar [Km (ATP) ≈ 37 μM; k cat ≈ 0.10 min−1] and typical of those of other histidine kinases. The catalytic efficiency of the two versions of WalK Spn ∼P were also similar in the phosphoryltransfer reaction to full-length WalR Spn . In contrast, absence of the HAMP-PAS domains significantly diminished the phosphatase activity of WalK Spn for WalR Spn ∼P. Deletion and point mutations confirmed that optimal WalK Spn phosphatase activity depended on the PAS domain as well as residues in the DHp domain. In addition, these WalK Spn DHp domain and ΔPAS mutations led to attenuation of virulence in a murine pneumonia model.

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HprSR is a Reactive Chlorine Species-Sensing, Two-Component System in Escherichia coli

Journal of Bacteriology ◽

10.1128/jb.00449-21 ◽

2021 ◽

Author(s):

Sara El Hajj ◽

Camille Henry ◽

Camille Andrieu ◽

Alexandra Vergnes ◽

Laurent Loiseau ◽

...

Keyword(s):

Escherichia Coli ◽

Hypochlorous Acid ◽

Methionine Sulfoxide ◽

Methionine Sulfoxide Reductase ◽

Repair System ◽

Bacterial Cells ◽

Two Component System ◽

Component System ◽

Two Component ◽

Redox Switch

Two-component systems (TCS) are signalling pathways that allow bacterial cells to sense, respond and adapt to fluctuating environments. Among the classical TCS of Escherichia coli , HprSR has recently been shown to be involved in the regulation of msrPQ , which encodes the periplasmic methionine sulfoxide reductase system. In this study, we demonstrate that hypochlorous acid (HOCl) induces the expression of msrPQ in an HprSR-dependant manner, whereas H 2 O 2 , NO and paraquat (a superoxide generator) do not. Therefore, HprS appears to be an HOCl-sensing histidine kinase. Using a directed mutagenesis approach, we show that Met residues located in the periplasmic loop of HprS are important for its activity: as HOCl preferentially oxidizes Met residues, we provide evidence that HprS could be activated via the reversible oxidation of its methionine residues, meaning that MsrPQ plays a role in switching HprSR off. We propose that the activation of HprS by HOCl could occur through a Met redox switch. HprSR appears to be the first characterized TCS able to detect reactive chlorine species (RCS) in E. coli . This study represents an important step towards understanding the mechanisms of RCS resistance in prokaryotes. IMPORTANCE Understanding how bacteria respond to oxidative stress at the molecular level is crucial in the fight against pathogens. HOCl is one of the most potent industrial and physiological microbiocidal oxidants. Therefore bacteria have developed counterstrategies to survive HOCl-induced stress. Over the last decade, important insights into these bacterial protection factors have been obtained. Our work establishes HprSR as a reactive chlorine species-sensing, two-component system in Escherichia coli MG1655, which regulates the expression of MsrPQ, a repair system for HOCl-oxidized proteins. Moreover we provide evidence suggesting that HOCl could activate HprS through a methionine redox switch.

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Disruption of Glycolysis by Nutritional Immunity Activates a Two-Component System That Coordinates a Metabolic and Antihost Response by Staphylococcus aureus

mBio ◽

10.1128/mbio.01321-19 ◽

2019 ◽

Vol 10 (4) ◽

Cited By ~ 4

Author(s):

Paola K. Párraga Solórzano ◽

Jiangwei Yao ◽

Charles O. Rock ◽

Thomas E. Kehl-Fie

Keyword(s):

Staphylococcus Aureus ◽

Metabolic Flux ◽

Bacterial Species ◽

Critical Role ◽

Dependent Manner ◽

Two Component System ◽

Component System ◽

Content Type ◽

Nutritional Immunity ◽

Two Component

ABSTRACT During infection, bacteria use two-component signal transduction systems to sense and adapt to the dynamic host environment. Despite critically contributing to infection, the activating signals of most of these regulators remain unknown. This also applies to the Staphylococcus aureus ArlRS two-component system, which contributes to virulence by coordinating the production of toxins, adhesins, and a metabolic response that enables the bacterium to overcome host-imposed manganese starvation. Restricting the availability of essential transition metals, a strategy known as nutritional immunity, constitutes a critical defense against infection. In this work, expression analysis revealed that manganese starvation imposed by the immune effector calprotectin or by the absence of glycolytic substrates activates ArlRS. Manganese starvation imposed by calprotectin also activated the ArlRS system even when glycolytic substrates were present. A combination of metabolomics, mutational analysis, and metabolic feeding experiments revealed that ArlRS is activated by alterations in metabolic flux occurring in the latter half of the glycolytic pathway. Moreover, calprotectin was found to induce expression of staphylococcal leukocidins in an ArlRS-dependent manner. These studies indicated that ArlRS is a metabolic sensor that allows S. aureus to integrate multiple environmental stresses that alter glycolytic flux to coordinate an antihost response and to adapt to manganese starvation. They also established that the latter half of glycolysis represents a checkpoint to monitor metabolic state in S. aureus. Altogether, these findings contribute to understanding how invading pathogens, such as S. aureus, adapt to the host during infection and suggest the existence of similar mechanisms in other bacterial species. IMPORTANCE Two-component regulatory systems enable bacteria to adapt to changes in their environment during infection by altering gene expression and coordinating antihost responses. Despite the critical role of two-component systems in bacterial survival and pathogenesis, the activating signals for most of these regulators remain unidentified. This is exemplified by ArlRS, a Staphylococcus aureus global regulator that contributes to virulence and to resisting host-mediated restriction of essential nutrients, such as manganese. In this report, we demonstrate that manganese starvation and the absence of glycolytic substrates activate ArlRS. Further investigations revealed that ArlRS is activated when the latter half of glycolysis is disrupted, suggesting that S. aureus monitors flux through the second half of this pathway. Host-imposed manganese starvation also induced the expression of pore-forming toxins in an ArlRS-dependent manner. Cumulatively, this work reveals that ArlRS acts as a sensor that links nutritional status, cellular metabolism, and virulence regulation.

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