Regulation of Denitrification Genes in
            <i>Neisseria meningitidis</i>
            by Nitric Oxide and the Repressor NsrR

Rock, Jonathan D.; Thomson, Melanie J.; Read, Robert C.; Moir, James

doi:10.1128/jb.01368-06

Cited by 51 publications

(86 citation statements)

References 31 publications

(40 reference statements)

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“…NO has a half life of a few seconds, so the study of its effects on P. gingivalis required the use of a NO donor (12) to maintain exposure of the bacterial cells to the compound. Among many methods (30,31,43,44,48,56), DEA NONOate had been successfully used in studies involving bacteria (42) and had a short enough half life to avoid unnecessary prolonged bacterial exposure to NO and allow us to determine easily the dose needed to replicate NO levels found in the periodontal pocket (47). Our studies revealed that, to mimic the average NO physiological range of periodontal pockets, the ideal DEA NONOate dose in hypoxic BHI was 10 l of 36 mM, or 0.36 mol.…”

Section: Resultsmentioning

confidence: 99%

Nitric Oxide Stress Resistance in Porphyromonas gingivalis Is Mediated by a Putative Hydroxylamine Reductase

et al. 2012

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bPorphyromonas gingivalis, the causative agent of adult periodontitis, must maintain nitric oxide (NO) homeostasis and surmount nitric oxide stress from host immune responses or other oral bacteria to survive in the periodontal pocket. To determine the involvement of a putative hydroxylamine reductase (PG0893) and a putative nitrite reductase-related protein (PG2213) in P. gingivalis W83 NO stress resistance, genes encoding those proteins were inactivated by allelic exchange mutagenesis. The isogenic mutants P. gingivalis FLL455 (PG0893::ermF) and FLL456 (PG2213::ermF) were black pigmented and showed growth rates and gingipain and hemolytic activities similar to those of the wild-type strain. P. gingivalis FLL455 was more sensitive to NO than the wild type. Complementation of P. gingivalis FLL455 with the wild-type gene restored the level of NO sensitivity to a level similar to that of the parent strain. P. gingivalis FLL455 and FLL456 showed sensitivity to oxidative stress similar to that of the wild-type strain. DNA microarray analysis showed that PG0893 and PG2213 were upregulated 1.4-and 2-fold, respectively, in cells exposed to NO. In addition, 178 genes were upregulated and 201 genes downregulated more than 2-fold. The majority of these modulated genes were hypothetical or of unknown function. PG1181, predicted to encode a transcriptional regulator, was upregulated 76-fold. Transcriptome in silico analysis of the microarray data showed major metabolomic variations in key pathways. Collectively, these findings indicate that PG0893 and several other genes may play an important role in P. gingivalis NO stress resistance. Porphyromonas gingivalis is a Gram-negative anaerobic bacterium that is a primary etiologic agent of periodontal disease. This infection-induced disease is characterized by inflammation, which can result in host-mediated destruction of tooth-supporting tissues and structures (18). Nitric oxide (NO) is a key component of the host immune response. Oral neutrophils can constitutively secrete low concentrations of NO via the involvement of multiple NO synthases. However, with activation, these neutrophils have increased secretion of NO with antimicrobial effects. As in other host-pathogen interactions (22), P. gingivalis has been shown to trigger the production of NO in immune and nonimmune host cells by activating the expression of inducible nitric oxide synthases (9, 54) and can survive in NO concentrations ranging from 4.9 M to 19.2 M (47). Elevated NO concentrations are reported to cause vasodilatation and a decrease in platelet aggregation, which may contribute to gingival bleeding (18), and to have cytotoxic effects on surrounding host tissue that can lead to alveolar bone loss (7). NO is present in the saliva and gingival fluid of periodontitis patients (7,18,57). Further, saliva NO concentrations have been shown to increase with the severity of periodontitis (47).P. gingivalis, as an asaccharolytic microorganism, metabolizes nitrogenous compounds as a source of energy and generates a micr...

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Section: Resultsmentioning

confidence: 99%

Nitric Oxide Stress Resistance in Porphyromonas gingivalis Is Mediated by a Putative Hydroxylamine Reductase

et al. 2012

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“…Therefore, bacteria have developed elaborate regulatory mechanisms to sense and detoxify NO. The NsrR transcriptional control plays a role in the NO stress response as a repressor in 24,29,49,61) and Gram-positive (38, 59) bacteria. Flavohemoglobin Hmp is often specified by a gene controlled by NsrR among diverse bacteria (50).…”

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confidence: 99%

Global Transcriptional Control by NsrR in Bacillus subtilis

et al. 2012

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The NO-sensitive NsrR repressor of Bacillus subtilis, which carries a [4Fe-4S] cluster, controls transcription of nasD and hmp (class I regulation) under anaerobic conditions. Here, we describe another class of NsrR regulation (class II regulation) that controls a more diverse collection of genes. Base substitution analysis showed that [4Fe-4S]-NsrR recognizes a partial dyad symmetry within the class I cis-acting sites, whereas NO-insensitive interaction of NsrR with an A؉T-rich class II regulatory site showed relaxed sequence specificity. Genome-wide transcriptome studies identified genes that are under the control of the class II NsrR regulation. The class II NsrR regulon includes genes controlled by both AbrB and Rok repressors, which also recognize A؉T-rich sequences, and by the Fur repressor. Transcription of class II genes was elevated in an nsrR mutant during anaerobic fermentative growth with pyruvate. Although NsrR binding to the class II regulatory sites was NO insensitive in vitro, transcription of class II genes was moderately induced by NO, which involved reversal of NsrR-dependent repression, suggesting that class II repression is also NO sensitive. In all NsrR-repressed genes tested, the loss of NsrR repressor activity was not sufficient to induce transcription as induction required the ResD response regulator. The ResD-ResE signal transduction system is essential for activation of genes involved in aerobic and anaerobic respiration. This study indicated coordinated regulation between ResD and NsrR and uncovered a new role of ResD and NsrR in transcriptional regulation during anaerobiosis of B. subtilis. N itric oxide (NO) has been shown to function as a signal molecule in bacterial physiology, but it can be toxic at high concentrations (17). Therefore, bacteria have developed elaborate regulatory mechanisms to sense and detoxify NO. The NsrR transcriptional control plays a role in the NO stress response as a repressor in 24,29,49,61) and Gram-positive (38, 59) bacteria. Flavohemoglobin Hmp is often specified by a gene controlled by NsrR among diverse bacteria (50). Hmp is required for Salmonella virulence in mice (6) and for its survival in a macrophage cell line (24). Hmp detoxifies NO by converting it to nitrate under aerobic conditions (21, 26). Other genes controlled by NsrR include those involved in NO metabolism and detoxification (summarized in reference 60), as predicted in a comparative genomics study (50). Furthermore, recent studies showed that NsrR controls NO-induced expression of NO-resistant alternative oxidase (Aox) in Vibrio fischeri, thus supporting bacterial growth during NO stress (14).NsrR is an [Fe-S] cluster-bearing protein, and binding of NsrR to its target genes is relieved in the presence of NO, leading to an upregulation (reviewed in reference 60). The mechanism by which NsrR controls transcription of genes in response to NO has been investigated in vitro. NsrR from Neisseria gonorrhoeae (29) and Streptomyces coelicolor (59) was shown to contain a [2Fe-2S] cluster when ...

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“…To allow construction of the nsrR aniA and the nsrR norB double mutants, the Sp resistance cassette interrupting the deleted nsrR gene (31) was replaced with a Tc cassette. An inverse PCR using primers NoregInv1 and NoregInv2 (31) and High Fidelity polymerase (Roche) allowed the removal of the Sp cassette inserted in nsrR (nsrR-Sp) on pJR113 and the introduction of artificial HindIII sites. The ϳ4-kb DNA product obtained was digested with HindIII and rendered blunt by treatment with a DNA polymerase I Klenow fragment prior to ligation with a 2.5-kb Tc cassette excised from pCMT18 with EcoRV.…”

Section: Methodsmentioning

confidence: 99%

“…The chief means of NO detoxification in N. meningitidis is the membrane-bound NO reductase NorB, with CycP having a secondary role (1). We have recently shown that the denitrification genes aniA and norB are both regulated by a repressor, NsrR, which is a sensor of NO (31).…”

mentioning

confidence: 99%

“…NsrR was first identified as a nitrite-sensing repressor in the nitrifying bacterium Nitrosomonas europaea (3) and has subsequently been identified in a number of bacteria including Escherichia coli (4), Bacillus subtilis (26), N. gonorrhoeae (28), and Salmonella enterica serovar Typhimurium (14), as well as N. meningitidis (31). Rodionov et al (32) predicted the role of NsrR in a number of alpha-, beta-, and gammaproteobacteria, in members of the order Bacillales, and in Streptomyces spp.…”

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confidence: 99%

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The Nitric Oxide (NO)-Sensing Repressor NsrR of Neisseria meningitidis Has a Compact Regulon of Genes Involved in NO Synthesis and Detoxification

et al. 2008

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We have analyzed the extent of regulation by the nitric oxide (NO)-sensitive repressor NsrR from Neisseria meningitidis MC58, using microarray analysis. Target genes that appeared to be regulated by NsrR, based on a comparison between an nsrR mutant and a wild-type strain, were further investigated by quantitative real-time PCR, revealing a very compact set of genes, as follows: norB (encoding NO reductase), dnrN (encoding a protein putatively involved in the repair of nitrosative damage to iron-sulfur clusters), aniA (encoding nitrite reductase), nirV (a putative nitrite reductase assembly protein), and mobA (a gene associated with molybdenum metabolism in other species but with a frame shift in N. meningitidis). In all cases, NsrR acts as a repressor. The NO protection systems norB and dnrN are regulated by NO in an NsrR-dependent manner, whereas the NO protection system cytochrome c (encoded by cycP) is not controlled by NO or NsrR, indicating that N. meningitidis expresses both constitutive and inducible NO protection systems. In addition, we present evidence to show that the anaerobic response regulator FNR is also sensitive to NO but less so than NsrR, resulting in complex regulation of promoters such as aniA, which is controlled by both FNR and NsrR: aniA was found to be maximally induced by intermediate NO concentrations, consistent with a regulatory system that allows expression during denitrification (in which NO accumulates) but is down-regulated as NO approaches toxic concentrations.

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Regulation of Denitrification Genes in Neisseria meningitidis by Nitric Oxide and the Repressor NsrR

Cited by 51 publications

References 31 publications

Nitric Oxide Stress Resistance in Porphyromonas gingivalis Is Mediated by a Putative Hydroxylamine Reductase

Nitric Oxide Stress Resistance in Porphyromonas gingivalis Is Mediated by a Putative Hydroxylamine Reductase

Global Transcriptional Control by NsrR in Bacillus subtilis

The Nitric Oxide (NO)-Sensing Repressor NsrR of Neisseria meningitidis Has a Compact Regulon of Genes Involved in NO Synthesis and Detoxification

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