Essential role of Bordetella NadC in a quinolinate salvage pathway for NAD biosynthesis

Abstract

Nicotinamide adenine dinucleotide (NAD) is produced via de novo biosynthesis pathways and by salvage or recycling routes. The classical Bordetella bacterial species are known to be auxotrophic for nicotinamide or nicotinic acid. This study confirmed that Bordetella bronchiseptica, Bordetella pertussis and Bordetella parapertussis have the recycling/salvage pathway genes pncA and pncB, for use of nicotinamide or nicotinic acid, respectively, for NAD synthesis. Although these Bordetellae lack the nadA and nadB genes needed for de novo NAD biosynthesis, remarkably, they have one de novo pathway gene, nadC, encoding quinolinate phosphoribosyltransferase. Genomic analyses of taxonomically related Bordetella and Achromobacter species also indicated the presence of an 'orphan' nadC and the absence of nadA and nadB. When supplied as the sole NAD precursor, quinolinate promoted B. bronchiseptica growth, and the ability to use it required nadC. Co-expression of Bordetella nadC with the nadB and nadA genes of Paraburkholderia phytofirmans allowed B. bronchiseptica to grow in the absence of supplied pyridines, indicative of de novo NAD synthesis and functional confirmation of Bordetella NadC activity. Expression of nadC in B. bronchiseptica was influenced by nicotinic acid and by a NadQ family transcriptional repressor, indicating that these organisms prioritize their use of pyridines for NAD biosynthesis.

The Bordetella bronchiseptica nic locus encodes a nicotinic acid degradation pathway and the 6‐hydroxynicotinate‐responsive regulator BpsR

“…). The WT strain carrying the control plasmid vector failed to grow in the absence of pyridines, but NAD prototrophy conferred by rescued its growth, as previously observed (Brickman et al ., ). Δ bpsR mutant RBB27 (vector control) similarly did not grow in the absence of pyridines, and also showed its typical poor growth on 30 μM Na.…”

“…In standard Bordetella laboratory SS culture, Na is used at micromolar concentrations required for sufficient NAD biosynthesis; however, phenotypic modulation of Bvg‐regulated genes occurs at millimolar Na concentrations that would also potentially be sufficient to serve as a carbon source. To determine whether Na catabolic pathway activity influences modulation, expression of a low‐copy transcriptional lacZ fusion to the Bvg‐activated adenylate cyclase toxin gene promoter ( cyaA‐lacZ ) (Brickman et al ., ) was used to monitor the response (Fig. ).…”

“…Furthermore, fluctuations in Nic pathway activity appear to alter intracellular Na concentrations, impacting Bordetella virulence gene expression. We have found no obvious correlations between the presence or absence of the nic gene clusters and the complement of NAD biosynthesis genes (Brickman et al ., ) among the Bordetella species.…”

“…A. Proposed Bordetella pathways for NAD biosynthesis based on previous studies (Brickman et al ., ) and nicotinic acid catabolism, modeled on the pathway determined by Jiménez and colleagues (). NAD pathway enzymes: PncA, nicotinamide deamidase; PncB, nicotinic acid phosphoribosyltransferase; PncC, nicotinamide mononucleotide deamidase; NadC, quinolinic acid phosphoribosyltransferase; NadD, nicotinic acid mononucleotide adenylyltransferase; NadE, NAD synthetase; NadK, NAD kinase.…”

“…In vitro, these organisms are nonfermentative-they apparently do not catabolize sugars, but can use glutamate and aketoglutarate as sole carbon sources, and depending on the species, will also catabolize lactate, acetate and A. Proposed Bordetella pathways for NAD biosynthesis based on previous studies (Brickman et al, 2017) and nicotinic acid catabolism, modeled on the pathway determined by Jim enez and colleagues (2008). NAD pathway enzymes: PncA, nicotinamide deamidase; PncB, nicotinic acid phosphoribosyltransferase; PncC, nicotinamide mononucleotide deamidase; NadC, quinolinic acid phosphoribosyltransferase; NadD, nicotinic acid mononucleotide adenylyltransferase; NadE, NAD synthetase; NadK, NAD kinase.…”