The Critical Role of the Conserved Thr247 Residue in the Functioning of the Osmosensor EnvZ, a Histidine Kinase/Phosphatase, in Escherichia coli

Dutta, Rinku; Yoshida, Takeshi; Inouye, Masayori

doi:10.1074/jbc.m005872200

Cited by 77 publications

(78 citation statements)

References 54 publications

(63 reference statements)

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“…The activated regulator protein then triggers expression of specific target genes. Many histidine kinases also possess phosphatase activities that allow dephosphorylation and hence inactivation of the regulator protein (37). In the case of EnvZ/OmpR, the level of phosphorylation of OmpR is important for the tight control of osmoregulation (40).…”

Section: Discussionmentioning

confidence: 99%

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CzcR-CzcS, a Two-component System Involved in Heavy Metal and Carbapenem Resistance in Pseudomonas aeruginosa

Perron

Caille

Rossier

et al. 2004

Journal of Biological Chemistry

287

272

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Pseudomonas aeruginosa is an environmental bacterium involved in mineralization of organic matter. It is also an opportunistic pathogen able to cause serious infections in immunocompromised hosts. As such, it is exposed to xenobiotics including solvents, heavy metals, and antimicrobials. We studied the response of P. aeruginosa upon exposure to heavy metals or antibiotics to investigate whether common regulatory mechanisms govern resistance to both types of compounds. We showed that sublethal zinc concentrations induced resistance to zinc, cadmium, and cobalt, while lethal zinc concentrations selected mutants constitutively resistant to these heavy metals. Both zinc-induced and stable zinc-resistant strains were also resistant to the carbapenem antibiotic imipenem. On the other hand, only 20% of clones selected on imipenem were also resistant to zinc. Heavy metal resistance in the mutants could be correlated by quantitative real time PCR with increased expression of the heavy metal efflux pump CzcCBA and its cognate two-component regulator genes czcR-czcS. Western blot analysis revealed reduced expression of the basic amino acid and carbapenem-specific OprD porin in all imipenem-resistant mutants. Sequencing of the czcR-czcS DNA region in eight independent zincand imipenem-resistant mutants revealed the presence of the same V194L mutation in the CzcS sensor protein. Overexpression in a susceptible wild type strain of the mutated CzsS protein, but not of the wild type form, resulted in decreased oprD and increased czcC expression. We further show that zinc is released from latex urinary catheters into urine in amounts sufficient to induce carbapenem resistance in P. aeruginosa, possibly compromising treatment of urinary tract infections by this class of antibiotics.Pseudomonas aeruginosa is a Gram-negative bacterium thriving in environments polluted with organic matter. It is also an opportunistic pathogen frequently encountered in the hospital, causing morbidity and mortality in immunocompromised and cystic fibrosis patients (1). P. aeruginosa is characterized by an intrinsically high level of resistance to xenobiotics including antimicrobial agents, solvents, and heavy metals (2), which can be accounted for by a combination of its low outer membrane permeability and the presence of multiple efflux pumps (3). These pumps belong to the resistance, nodulation, cell division (RND) 1 transporter family, present in many Gramnegative bacteria (4). To extrude substrates from the cytoplasm across the two membranes, these systems are composed of a proton antiporter located in the cytoplasmic membrane, a membrane fusion protein spanning the periplasmic space, and an outer membrane protein (5). Members of the RND family, namely the Mex pumps, have recently gained increasing interest. In particular, the constitutively expressed MexAB-OprM (6, 7) and the inducible MexXY (8) efflux pumps endow the PAO1 reference strain and other clinical isolates (9) with a natural resistance to a wide range of antimicrobial agents. Proton-dr...

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Section: Discussionmentioning

confidence: 99%

“…According to sequence analysis, CzcS, like EnvZ in Escherichia coli, belongs to class I histidine kinases. In this class the catalytic and ATP-binding domain is separated in two regions (37). The A domain is involved in the dimerization, phosphotransfer, and phosphatase activity, while the B domain binds ATP (38,39).…”

Section: Discussionmentioning

confidence: 99%

CzcR-CzcS, a Two-component System Involved in Heavy Metal and Carbapenem Resistance in Pseudomonas aeruginosa

Perron

Caille

Rossier

et al. 2004

Journal of Biological Chemistry

287

272

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“…Point mutations within a related bacterial HK, EnvZ, have been identified that affect the kinase activity, the phosphatase activity, or both enzymatic activities of this bifunctional enzyme. Based on current work characterizing the effects of point mutations on the enzymatic activity of EnvZ (Dutta and Inouye, 1996;Hsing et al, 1998;Dutta et al, 2000), however, it is not possible to predict how an amino acid substitution of a Phe residue for Leu 453 would affect CRE1 expression or function in vivo. Examination of the sequencing chromatograms revealed that the single base change detected in the CRE1 coding sequence was found in a homozygous rather than heterozygous state (data not shown).…”

Section: Discussionmentioning

confidence: 99%

A Transcriptome-Based Characterization of Habituation in Plant Tissue Culture

Pischke¹,

Huttlin²,

Hegeman³

et al. 2006

Plant Physiology

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For the last 50 years, scientists have recognized that varying ratios of the plant hormones cytokinin and auxin induce plant cells to form particular tissues: undifferentiated calli, shoot structures, root structures, or a whole plant. Proliferation of undifferentiated callus tissue, greening, and the formation of shoot structures are all cytokinin-dependent processes. Habituation refers to a naturally occurring phenomenon whereby callus cultures, upon continued passage, lose their requirement for cytokinin. Earlier studies of calli with a higher-than-normal cytokinin content indicate that overproduction of cytokinin by the culture tissues is a possible explanation for this acquired cytokinin independence. A transcriptome-based analysis of a well established habituated Arabidopsis (Arabidopsis thaliana) cell culture line was undertaken, to explore genome-wide expression changes underlying the phenomenon of habituation. Increased levels of expression of the cytokinin receptor CRE1, as well as altered levels of expression of several other genes involved in cytokinin signaling, indicated that naturally acquired deregulation of cytokinin-signaling components could play a previously unrecognized role in habituation. Up-regulation of several cytokinin oxidases, downregulation of several known cytokinin-inducible genes, and a lack of regulation of the cytokinin synthases indicated that increases in hormone concentration may not be required for habituation. In addition, up-regulation of the homeodomain transcription factor FWA, transposon-related elements, and several DNA-and chromatin-modifying enzymes indicated that epigenetic changes contribute to the acquisition of cytokinin habituation.

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“…(lanes [25][26][27][28] with all the other experiments], the reason for which is not known at present.…”

Section: Resultsmentioning

confidence: 99%

“…To confirm the above conclusions further, we took advantage of an EnvZc mutant, T247Y, which has kinase activity but lacks phosphatase activity (26). As shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

The critical role of DNA in the equilibrium between OmpR and phosphorylated OmpR mediated by EnvZ in Escherichiacoli

Qin¹

2001

Proceedings of the National Academy of Sciences

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Escherichia coli modulates its porin expression through a histidine kinase, EnvZ, and its cognate response regulator, OmpR. EnvZ is a bifunctional enzyme that possesses both OmpR kinase and phosphorylated OmpR (OmpR-P) phosphatase activities and thus controls the cellular level of OmpR-P. In an in vitro-assay system, the addition of OmpR to the reaction mixture consisting of the cytoplasmic domain of EnvZ (EnvZc) and ATP produces a barely detectable amount of OmpR-P because of the dual activities of EnvZ. Here we report that DNA fragments containing the upstream promoter regions of the porin genes (ompF and ompC) can shift the equilibrium between OmpR and OmpR-P dramatically toward OmpR-P. Among the four reactions occurring in the mixture, only the EnvZ phosphatase activity was inhibited severely by the specific DNA, in contrast to the previous report by Kenney and her associates that The autophosphorylation of EnvZc and the phosphotransfer from phosphorylated EnvZc to OmpR were not affected by DNA, whereas the autodephosphorylation of OmpR-P was inhibited slightly. We propose that the apparent inhibitory effect of DNA on the EnvZ phosphatase function is caused by sequestrating OmpR-P from the reaction as a result of OmpR-P binding to DNA.

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The Critical Role of the Conserved Thr247 Residue in the Functioning of the Osmosensor EnvZ, a Histidine Kinase/Phosphatase, in Escherichia coli

Cited by 77 publications

References 54 publications

CzcR-CzcS, a Two-component System Involved in Heavy Metal and Carbapenem Resistance in Pseudomonas aeruginosa

CzcR-CzcS, a Two-component System Involved in Heavy Metal and Carbapenem Resistance in Pseudomonas aeruginosa

A Transcriptome-Based Characterization of Habituation in Plant Tissue Culture

The critical role of DNA in the equilibrium between OmpR and phosphorylated OmpR mediated by EnvZ in Escherichiacoli

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