Rex (Encoded by DVU_0916) in Desulfovibrio vulgaris Hildenborough Is a Repressor of Sulfate Adenylyl Transferase and Is Regulated by NADH

Abstract

e Although the enzymes for dissimilatory sulfate reduction by microbes have been studied, the mechanisms for transcriptional regulation of the encoding genes remain unknown. In a number of bacteria the transcriptional regulator Rex has been shown to play a key role as a repressor of genes producing proteins involved in energy conversion. In the model sulfate-reducing microbe Desulfovibrio vulgaris Hildenborough, the gene DVU_0916 was observed to resemble other known Rex proteins. Therefore, the DVU_0916 protein has been predicted to be a transcriptional repressor of genes encoding proteins that function in the process of sulfate reduction in D. vulgaris Hildenborough. Examination of the deduced DVU_0916 protein identified two domains, one a winged helix DNA-binding domain common for transcription factors, and the other a Rossman fold that could potentially interact with pyridine nucleotides. A deletion of the putative rex gene was made in D. vulgaris Hildenborough, and transcript expression studies of sat, encoding sulfate adenylyl transferase, showed increased levels in the D. vulgaris Hildenborough Rex (Rex DvH ) mutant relative to the parental strain. The Rex DvH -binding site upstream of sat was identified, confirming Rex DvH to be a repressor of sat. We established in vitro that the presence of elevated NADH disrupted the interaction between Rex DvH and DNA. Examination of the 5= transcriptional start site for the sat mRNA revealed two unique start sites, one for respiring cells that correlated with the Rex DvH -binding site and a second for fermenting cells. Collectively, these data support the role of Rex DvH as a transcription repressor for sat that senses the redox status of the cell.T he anaerobic process of microbially influenced corrosion is estimated to account for 15% of the total cost of ferrous metal and concrete/stonework corrosion. For the U.S. energy industry, this amounts to approximately $100 billion yearly (1). Most notably, the metabolic processes of the sulfate-reducing microbes (SRM) have been implicated in this observed corrosion (2, 3). Clearly, it would be beneficial to understand the mechanism and the regulation of genes involved in the key processes leading to metal dissolution.The SRM convert energy by dissimilatory sulfate reduction, where sulfate is used as a terminal electron acceptor in respiration. Although SRM are mainly found in sulfate-rich anoxic environments (4), they are also found in anoxic habitats depleted of sulfate because they are able to use electron acceptors other than sulfate (5, 6). Desulfovibrio vulgaris Hildenborough, a Gram-negative deltaproteobacterium, is a model organism for examining sulfate reduction as it is genetically accessible and is readily cultured in the lab (7,8). Also, the genome for D. vulgaris Hildenborough has been sequenced (9) and has recently been reannotated (10).The process of dissimilatory sulfate reduction is carried out by a well-conserved biochemical pathway in heterogeneous SRM (5,(11)(12)(13). In brief, sulfate (SO 4 2Ϫ ) is tak...

A Post-Genomic View of the Ecophysiology, Catabolism and Biotechnological Relevance of Sulphate-Reducing Prokaryotes

“…This response is probably linked to the redox status of the cell, as a reducing intracellular environment generated by fermentation is likely to prime cells for sulphate reduction, even if this is not available. Several of these proteins belong to the regulon of the redox-responsive regulator Rex that senses the intracellular NADH/NAD + ratio Ravcheev et al, 2012;Rodionov et al, 2004), is a repressor of the sat gene (Christensen et al, 2015) and is probably involved in this response. A screening of mutants deficient in pyruvate fermentation revealed that deletion of the genes for malic enzyme, fumarate reductase, a dicarboxylate transporter and the NfnAB transhydrogenase led to a strong growth inhibition, whereas mutants in the Rnf and Hdr-Flx proteins had somewhat slower growth than the wild type .…”

“…More intensive investigations of single transcriptional regulators, such as Fur (Bender et al, 2007), CooA , SahR , Rex (Christensen et al, 2015) and TunR (Kazakov, Rajeev, et al, 2013), have been reported, and a few insights from these studies will be presented below. CooA, the CO sensor (Youn, Kerby, Conrad, & Roberts, 2004) and activator for cooSC, encoding CO dehydrogenase, was shown by the Stahl laboratory to have regulatory functions in D. vulgaris even in the absence of externally added CO .…”

“…In studies to explore the regulation of the genes encoding the enzymes needed for sulphate reduction, sat (sulphate adenylyl transferase), apsBA (adenylyl phosphosulphate reductase) and dsrABD (dissimilatory sulphite reductase) in the SRP, attention turned to the ortholog of the redox sensor and repressor, rex (Christensen et al, 2015). This regulator was expected to sense the redox status of the cell by binding NAD + and/or NADH (Sickmier et al, 2005).…”

“…The Rnf complex plays an important role in linking the NADH and ferredoxin pools in the cell, and in the Desulfovibrionaceae, it is predicted to be part of the Rex regulon along with several genes involved in sulphate reduction Ravcheev et al, 2012;Rodionov, Dubchak, Arkin, Alm, & Gelfand, 2004). Rex is a transcriptional regulator that senses the intracellular NADH/NAD + ratio and in D. vulgaris was recently shown to act as a repressor of the sat gene, the first gene involved in the sulphate reduction pathway (Christensen et al, 2015) (see also Section 7.4).…”

“…Many additional marker-exchange mutants have been generated in D. vulgaris Hildenborough, for example, ΔhmcA encoding a 16-haem periplasmic cytochrome subunit of the Hmc membrane complex ; Δroo encoding rubredoxin:oxygen oxidoreductase (Rodrigues, Vicente, et al, 2006); Δfur, a transcriptional regulator of genes encoding iron uptake functions (Bender et al, 2007); ΔqmoABCD, a quinone-interacting membrane-bound oxidoreductase (Zane et al, 2010); Δrex, a transcriptional repressor of genes functioning energy metabolism responsive to NADH/ NAD + ratios (Christensen et al, 2015); ΔfdhAB and ΔfdhABC 3 , soluble periplasmic formate dehydrogenases (da ; and ΔflxA, a component of a novel NADH dehydrogenase needed for ethanol metabolism (Ramos et al, 2015). Each has added to our understanding of electron flow in the SRP.…”