Transcriptional regulation of NAD metabolism in bacteria: genomic reconstruction of NiaR (YrxA) regulon

Abstract

A comparative genomic approach was used to reconstruct transcriptional regulation of NAD biosynthesis in bacteria containing orthologs of Bacillus subtilis gene yrxA, a previously identified niacin-responsive repressor of NAD de novo synthesis. Members of YrxA family (re-named here NiaR) are broadly conserved in the Bacillus/Clostridium group and in the deeply branching Fusobacteria and Thermotogales lineages. We analyzed upstream regions of genes associated with NAD biosynthesis to identify candidate NiaR-binding DNA motifs and assess the NiaR regulon content in these species. Representatives of the two distinct types of candidate NiaR-binding sites, characteristic of the Firmicutes and Thermotogales, were verified by an electrophoretic mobility shift assay. In addition to transcriptional control of the nadABC genes, the NiaR regulon in some species extends to niacin salvage (the pncAB genes) and includes uncharacterized membrane proteins possibly involved in niacin transport. The involvement in niacin uptake proposed for one of these proteins (re-named NiaP), encoded by the B. subtilis gene yceI, was experimentally verified. In addition to bacteria, members of the NiaP family are conserved in multicellular eukaryotes, including human, pointing to possible NaiP involvement in niacin utilization in these organisms. Overall, the analysis of the NiaR and NrtR regulons (described in the accompanying paper) revealed mechanisms of transcriptional regulation of NAD metabolism in nearly a hundred diverse bacteria.

Genomics-driven Reconstruction of Acinetobacter NAD Metabolism

“…Gene abNiaP was tentatively assigned a role of Nm and/or nicotinic acid (Na) transporter based on homology with the previously characterized distant member of this family from Bacillus subtilis (39). Gene deletion and phenotype analysis experiments were designed to verify this functional assignment and to test whether abNiaP is essential for Nm and/or Na salvage or (as in B. subtilis) there are additional mechanisms of their uptake.…”

“…S3 for dose dependence data). As in case of B. subtilis (39), an alternative (NiaP-independent) uptake mechanism is operational at higher niacin concentration (Ͼ100 M).…”

“…Uptake of Nm and Na-Two complementary approaches were used to assess the physiological role of the predicted abNiaP transporter similar to how it was previously described for NiaP from B. subtilis (39). In the first "knock-in" approach the overexpression of the niaP gene from A. baylyi in E. coli BW25113 ⌬nadA strain led to a notable increase in its growth rate on minimal medium supplemented with a limiting concentration of either Nm or Na (0.16 M) (Fig.…”

Characterization of NAD salvage pathways and their role in virulence in Streptococcus pneumoniae

“…There are two major ways that bacteria obtain the essential cofactor NAD: through de novo synthesis, which occurs in Escherichia coli and multiple members of the genera Bacillus and Clostridium, and through the salvage pathway, which occurs in the order Lactobacillales, specifically in members of the family Streptococcaceae. Some bacteria, such as Salmonella enterica serovar Typhimurium, have both systems (Chandler & Gholson, 1972;Rodionov et al, 2008;Sorci et al, 2013;Spector et al, 1985;Zhu et al, 1988). While the de novo pathway is characterized by synthesizing NAD from the amino acid aspartic acid, the salvage pathway imports intermediates several steps downstream in the NAD de novo synthesis pathway (Rodionov et al, 2008).…”

“…While the de novo pathway is characterized by synthesizing NAD from the amino acid aspartic acid, the salvage pathway imports intermediates several steps downstream in the NAD de novo synthesis pathway (Rodionov et al, 2008). The two major importers in the NAD salvage pathway are NiaX, responsible for niacin uptake, and PnuC, responsible for nicotinamide riboside (Herbert et al, 2003;Rodionov et al, 2008Rodionov et al, , 2009Rodionov et al, 2008Rodionov et al, , 2009Sauer et al, 2004).…”