Two-Component System That Regulates Methanol and Formaldehyde Oxidation in
            <i>Paracoccus denitrificans</i>

Harms, N.; Reijnders, W. N. M.; Koning, Sonja M.; Spanning, Rob J.M. van

doi:10.1128/jb.183.2.664-670.2001

Cited by 28 publications

(42 citation statements)

References 36 publications

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“…No genes with high homologies to the regulatory genes involved in methanol or methylamine oxidation functions that have been characterized in M. extorquens (9) or Paracoccus denitrificans (14,29,30) are identifiable in the M. flagellatus chromosome, pointing to either a lack of regulatory systems or to the existence of nonhomologous regulatory systems.…”

Section: Resultsmentioning

confidence: 99%

Genome of Methylobacillus flagellatus , Molecular Basis for Obligate Methylotrophy, and Polyphyletic Origin of Methylotrophy

et al. 2007

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Along with methane, methanol and methylated amines represent important biogenic atmospheric constituents; thus, not only methanotrophs but also nonmethanotrophic methylotrophs play a significant role in global carbon cycling. The complete genome of a model obligate methanol and methylamine utilizer, Methylobacillus flagellatus (strain KT) was sequenced. The genome is represented by a single circular chromosome of approximately 3 Mbp, potentially encoding a total of 2,766 proteins. Based on genome analysis as well as the results from previous genetic and mutational analyses, methylotrophy is enabled by methanol and methylamine dehydrogenases and their specific electron transport chain components, the tetrahydromethanopterin-linked formaldehyde oxidation pathway and the assimilatory and dissimilatory ribulose monophosphate cycles, and by a formate dehydrogenase. Some of the methylotrophy genes are present in more than one (identical or nonidentical) copy. The obligate dependence on single-carbon compounds appears to be due to the incomplete tricarboxylic acid cycle, as no genes potentially encoding alpha-ketoglutarate, malate, or succinate dehydrogenases are identifiable. The genome of M. flagellatus was compared in terms of methylotrophy functions to the previously sequenced genomes of three methylotrophs, Methylobacterium extorquens (an alphaproteobacterium, 7 Mbp), Methylibium petroleiphilum (a betaproteobacterium, 4 Mbp), and Methylococcus capsulatus (a gammaproteobacterium, 3.3 Mbp). Strikingly, metabolically and/or phylogenetically, the methylotrophy functions in M. flagellatus were more similar to those in M. capsulatus and M. extorquens than to the ones in the more closely related M. petroleiphilum species, providing the first genomic evidence for the polyphyletic origin of methylotrophy in Betaproteobacteria.Methylotrophy is the metabolic capacity to grow on reduced carbon compounds containing no COC bonds, such as methane, methanol, methylated amines, etc. (1, 39). While the role of methanotrophs in the reduction of global emissions of methane has been well recognized (27), less attention has been paid to nonmethanotrophic methylotrophs as participants in the global carbon cycle. However, recent models estimate methanol emissions into the atmosphere at 82 to 273 teragrams (Tg) year Ϫ1 (with living plants as the major source [19,25]), putting methanol emission on a scale similar to that of methane emissions (approximately 600 Tg year Ϫ1 [36]) and pointing toward the global role of nonmethanotrophic methanol utilizers. While no global modeling has been attempted to characterize the production of methylated amines, they are known to be abundant in marine and freshwater environments and represent dynamic constituents of not only carbon but also nitrogen global cycles (44). So far, only nonmethanotrophic methylotrophs have been implicated in utilizing methylated amines (1, 39).Methylobacillus flagellatus strain KT utilizes methanol and methylated amines as the sole sources of carbon and energy and is classifi...

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Section: Resultsmentioning

confidence: 99%

Genome of Methylobacillus flagellatus , Molecular Basis for Obligate Methylotrophy, and Polyphyletic Origin of Methylotrophy

et al. 2007

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“…We called these latter genes The predicted sensor histidine kinase, AfdS, has significant amino acid sequence similarity to RfdS (39% identity, 52% similarity) and to a P. denitrificans protein, FlhS (51% identity, 64% similarity), that is required to increase GSH-FDH expression (see Fig. 6) (17). The potential response regulator, AfdR, is predicted to contain a C-terminal DNA-binding domain in the NarL response regulator family with significant amino acid sequence similarity to RfdR (44% identity, 59% similarity) and P. denitrificans FlhR (61% identity, 76% similarity).…”

Section: Resultsmentioning

confidence: 99%

“…2b). In the region surrounding adhI-cycI are genes that could encode enzymes to oxidize methanol to formaldehyde (XoxF), periplasmic c-type cytochromes (CycI and CycB), a protein to facilitate the formation of S-hydroxymethylglutathione from formaldehyde and GSH (GfaA), and open reading frames which encode homologs of proteins of unknown function that are derived from genes typically linked to those encoding GSH-FDH family members in other eubacteria (14,15,17). This region also contains genes (RSP2591 to RSP2593) predicted to encode a sensor histidine kinase, a response regulator, and a predicted integral membrane protein of unknown function (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…In Saccharomyces cerevisiae, GSH-FDH levels are increased by the presence of methylated compounds like methyl methanesulphonate or formaldehyde (36). In many eubacteria, including R. sphaeroides, expression of the GSH-FDH gene (adhI) is increased when either formaldehyde or metabolic sources of this compound, such as methanol or choline, are present (6,15,17). These and other results led to the proposal that a pathway intermediate, possibly formaldehyde itself, increases adhI transcription.…”

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confidence: 86%

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Positive and Negative Transcriptional Regulators of Glutathione-Dependent Formaldehyde Metabolism

Hickman¹,

Witthuhn²,

Dominguez

et al. 2004

J Bacteriol

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A glutathione (GSH)-dependent pathway is used for formaldehyde metabolism by a wide variety of prokaryotes and eukaryotes. In this pathway, S-hydroxymethylglutathione, produced by the reaction of formaldehyde with the thiolate moiety of glutathione, is the substrate for a GSH-dependent formaldehyde dehydrogenase (GSH-FDH). While expression of GSH-FDH often increases in the presence of metabolic or exogenous sources of formaldehyde, little is known about the factors that regulate this response. Here, we identify two signal transduction pathways that regulate expression of adhI, the gene encoding GSH-FDH, in Rhodobacter sphaeroides. The loss of the histidine kinase response regulator pair RfdRS or the histidine kinase RfdS increases adhI transcription in the absence of metabolic sources of formaldehyde. Cells lacking RfdRS further increase adhI expression in the presence of metabolic sources of formaldehyde (methanol), suggesting that this negative regulator of GSH-FDH expression does not respond to this compound. In contrast, mutants lacking the histidine kinase response regulator pair AfdRS or the histidine kinase AfdS cannot induce adhI expression in the presence of either formaldehyde or metabolic sources of this compound. AfdR stimulates activity of the adhI promoter in vitro, indicating that this protein is a direct activator of GSH-FDH expression. Activation by AfdR is detectable only after incubation of the protein with acetyl phosphate, suggesting that phosphorylation is necessary for transcription activation. Activation of adhI transcription by acetyl-phosphate-treated AfdR in vitro is inhibited by a truncated RfdR protein, suggesting that this protein is a direct repressor of GSH-FDH expression. Together, the data indicate that AfdRS and RfdRS positively and negatively regulate adhI transcription in response to different signals.Formaldehyde is a cytogenic compound that is produced by environmental, industrial, and metabolic processes including the oxidative demethylation of amino acids and osmoprotectants or oxidation of one-carbon compounds like methanol, methyl halides, and methane (10, 11, 16, 20-22, 24, 35, 37, 38). The toxicity of formaldehyde stems from its reactivity with amino and sulfhydryl groups of biological molecules, causing alkylation, mutations, and cross-links that destroy the function of membranes, proteins, and nucleic acids (18,25). When the sources of formaldehyde, its toxicity, and the carbon skeletons and reducing power derived from its oxidation are considered, it is clear that cells benefit from metabolism of this compound.We are studying formaldehyde metabolism by the facultative bacterium Rhodobacter sphaeroides. This ␣-proteobacterium uses a glutathione (GSH)-dependent pathway for formaldehyde metabolism that is similar to those present in several prokaryotic and eukaryotic cells (5, 7). In this pathway, formaldehyde reacts with the thiolate moiety of GSH to form Shydroxymethylglutathione, a substrate for a GSH-dependent formaldehyde dehydrogenase (GSH-FDH). GSH-FDH oxidizes...

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“…AdhI and CycI play a central role in GSHdependent formaldehyde detoxification, and a function of class III alcohol dehydrogenases like AdhI in oxidative stress is established (13,54). Moreover, it is known that the FlhR homolog in Paracoccus denitrificans is involved in the regulation of the cycB-xoxJ operon and fdhA (55). The loss of regulation through a triple-base mutation in the predicted binding site within the flhR mRNA and the observed interaction between CcsR1 and the respective region of the flhR mRNA in vitro support the predictions of CcsR1-4 binding to this specific region.…”

Section: Discussionmentioning

confidence: 99%

A Cluster of Four Homologous Small RNAs Modulates C ₁ Metabolism and the Pyruvate Dehydrogenase Complex in Rhodobacter sphaeroides under Various Stress Conditions

et al. 2015

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In bacteria, regulatory RNAs play an important role in the regulation and balancing of many cellular processes and stress responses. Among these regulatory RNAs, trans-encoded small RNAs (sRNAs) are of particular interest since one sRNA can lead to the regulation of multiple target mRNAs. In the purple bacterium Rhodobacter sphaeroides, several sRNAs are induced by oxidative stress. In this study, we focused on the functional characterization of four homologous sRNAs that are cotranscribed with the gene for the conserved hypothetical protein RSP_6037, a genetic arrangement described for only a few sRNAs until now. Each of the four sRNAs is characterized by two stem-loops that carry CCUCCUCCC motifs in their loops. They are induced under oxidative stress, as well as by various other stress conditions, and were therefore renamed here sRNAs CcsR1 to CcsR4 (CcsR1-4) for conserved CCUCCUCCC motif stress-induced RNAs 1 to 4. Increased CcsR1-4 expression decreases the expression of genes involved in C 1 metabolism or encoding components of the pyruvate dehydrogenase complex either directly by binding to their target mRNAs or indirectly. One of the CcsR1-4 target mRNAs encodes the transcriptional regulator FlhR, an activator of glutathione-dependent methanol/formaldehyde metabolism. Downregulation of this glutathione-dependent pathway increases the pool of glutathione, which helps to counteract oxidative stress. The FlhR-dependent downregulation of the pyruvate dehydrogenase complex reduces a primary target of reactive oxygen species and reduces aerobic electron transport, a main source of reactive oxygen species. Our findings reveal a previously unknown strategy used by bacteria to counteract oxidative stress. IMPORTANCEPhototrophic organisms have to cope with photo-oxidative stress due to the function of chlorophylls as photosensitizers for the formation of singlet oxygen. Our study assigns an important role in photo-oxidative stress resistance to a cluster of four homologous sRNAs in the anoxygenic phototrophic bacterium Rhodobacter sphaeroides. We reveal a function of these regulatory RNAs in the fine-tuning of C 1 metabolism. A model that relates oxidative stress defense to C 1 metabolism is presented. The purple bacterium Rhodobacter sphaeroides is able to adapt its life-style to changing environmental conditions by making use of many different metabolic pathways like aerobic and anaerobic respiration, fermentation, and anaerobic anoxygenic photosynthesis. Major determinants of the life-style of R. sphaeroides are light quality and quantity, as well as oxygen tension. Under microaerobic conditions, R. sphaeroides produces photosynthetic complexes in the dark, while light inhibits their formation (1, 2). In addition to the intricate transcriptional regulation of photosynthesis genes by diverse protein regulators, photosynthetic complex formation is also based on oxygen-dependent mRNA stability for photosynthetically relevant genes and control by sRNAs (3, 4). Light-dependent inhibition of photosynthetic com...

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Two-Component System That Regulates Methanol and Formaldehyde Oxidation in Paracoccus denitrificans

Cited by 28 publications

References 36 publications

Genome of Methylobacillus flagellatus , Molecular Basis for Obligate Methylotrophy, and Polyphyletic Origin of Methylotrophy

Genome of Methylobacillus flagellatus , Molecular Basis for Obligate Methylotrophy, and Polyphyletic Origin of Methylotrophy

Positive and Negative Transcriptional Regulators of Glutathione-Dependent Formaldehyde Metabolism

A Cluster of Four Homologous Small RNAs Modulates C ₁ Metabolism and the Pyruvate Dehydrogenase Complex in Rhodobacter sphaeroides under Various Stress Conditions

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Two-Component System That Regulates Methanol and Formaldehyde Oxidation in Paracoccus denitrificans

Cited by 28 publications

References 36 publications

Genome of Methylobacillus flagellatus , Molecular Basis for Obligate Methylotrophy, and Polyphyletic Origin of Methylotrophy

Genome of Methylobacillus flagellatus , Molecular Basis for Obligate Methylotrophy, and Polyphyletic Origin of Methylotrophy

Positive and Negative Transcriptional Regulators of Glutathione-Dependent Formaldehyde Metabolism

A Cluster of Four Homologous Small RNAs Modulates C 1 Metabolism and the Pyruvate Dehydrogenase Complex in Rhodobacter sphaeroides under Various Stress Conditions

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Product

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A Cluster of Four Homologous Small RNAs Modulates C ₁ Metabolism and the Pyruvate Dehydrogenase Complex in Rhodobacter sphaeroides under Various Stress Conditions