Characterization of the GbdR Regulon in Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa displays tremendous metabolic diversity, controlled in part by the abundance of transcription regulators in the genome. We have been investigating P. aeruginosa's response to the host, particularly changes regulated by the hostderived quaternary amines choline and glycine betaine (GB). We previously identified GbdR as an AraC family transcription factor that directly regulates choline acquisition from host phospholipids (via binding to plcH and pchP promoters), is required for catabolism of the choline metabolite GB, and is an activator that induces transcription in response to GB or dimethylglycine. Our goal was to characterize the GbdR regulon in P. aeruginosa by using genetics and chemical biology in combination with transcriptomics and in vitro DNA-binding assays. Here we show that GbdR activation regulates transcription of 26 genes from 12 promoters, 11 of which have measureable binding to GbdR in vitro. The GbdR regulon includes the genes encoding GB, dimethylglycine, sarcosine, glycine, and serine catabolic enzymes and the BetX and CbcXWV quaternary amine transport proteins. We characterized the GbdR consensus binding site and used it to identify that the recently characterized acetylcholine esterase gene, choE (PA4921), is also regulated by GbdR. The regulon member not directly controlled by GbdR is the secreted lipase gene lipA, which was also the only regulon member repressed under GbdR-activating conditions. Determination of the GbdR regulon provides deeper understanding of how GbdR links bacterial metabolism and virulence. Additionally, identification of two uncharacterized regulon members suggests roles for these proteins in response to choline metabolites.

Self-regulating genomic island encoding tandem regulators confers chromatic acclimation to marine Synechococcus

“…3D) appears to be quite different from other AraC-class family members. Recent RNA-Seq studies of araC mutants in Escherichia coli and Salmonella demonstrate that AraC has a much broader regulon than previously known (27), and other AraC family members also control large regulons in a diverse set of bacteria (28,29). The small and highly specific FciA/FciB regulon is also different from the widespread responses in gene expression that have been observed for other forms of chromatic acclimation (13,30).…”

Liberate and Grab It, Ingest and Digest It: the GbdR Regulon of the Pathogen Pseudomonas aeruginosa

“…1 in reference 22.) Since some members of the GbdR regulon have previously been identified as virulence factors (24)(25)(26), the data reported by Hampel et al (22) provide a link between metabolism and the ability of P. aeruginosa to colonize its hosts and persist at infection sites. The harvest, transport, and catabolic genes in the GbdR regulon are evolutionarily conserved in a number of microorganisms that can metabolize choline under aerobic conditions, in particular in several plant-associated bacteria (e.g., Pseudomonas syringae [20,27]), thereby extending the physiological relevance of the reported findings well beyond the particular microorganism studied by Hampel et al (22).…”

“…In the current issue of the Journal of Bacteriology, Hampel et al (22) present a comprehensive analysis of the acquisition, import, and catabolism of choline and glycine betaine by P. aeruginosa, a process that is genetically controlled by GbdR, a glycine betaineresponsive activator protein and member of the AraC family of transcriptional regulators (23). Through a clever combination of chemical biology, genome-wide transcriptional profiling, bacterial genetics, DNA binding studies, and bioinformatics, Hampel et al (22) were able to define the GbdR regulon and identify those members that allow P. aeruginosa to liberate choline from host cells through hydrolytic enzymes, mediate its import, and afford its subsequent catabolism, first to glycine betaine and then further to the central metabolite pyruvate.…”

“…Through a clever combination of chemical biology, genome-wide transcriptional profiling, bacterial genetics, DNA binding studies, and bioinformatics, Hampel et al (22) were able to define the GbdR regulon and identify those members that allow P. aeruginosa to liberate choline from host cells through hydrolytic enzymes, mediate its import, and afford its subsequent catabolism, first to glycine betaine and then further to the central metabolite pyruvate. (For details of the catabolic pathway, see Fig.…”

“…To define the GbdR regulon of P. aeruginosa more comprehensively, Hampel et al (22) now used genome-wide transcriptional profiling experiments, but they used a very clever approach to avoid possible pitfalls that potentially could result from comparing the transcriptional profiles of a wild-type P. aeruginosa strain and its isogenic gbdR mutant. Since GbdR was already known to regulate genes for the import of choline and the inducers glycine betaine and dimethylglycine, gbdR mutant strains could potentially exhibit undesirable downstream metabolic and regulatory effects.…”