Pseudomonas aeruginosa Ethanol Oxidation by AdhA in Low-Oxygen Environments

Abstract

Pseudomonas aeruginosa has a broad metabolic repertoire that facilitates its coexistence with different microbes. Many microbes secrete products that P. aeruginosa can then catabolize, including ethanol, a common fermentation product. Here, we show that under oxygen-limiting conditions P. aeruginosa utilizes AdhA, an NAD-linked alcohol dehydrogenase, as a previously undescribed means for ethanol catabolism. In a rich medium containing ethanol, AdhA, but not the previously described PQQ-linked alcohol dehydrogenase, ExaA, oxidizes ethanol and leads to the accumulation of acetate in culture supernatants. AdhA-dependent acetate accumulation and the accompanying decrease in pH promote P. aeruginosa survival in LB-grown stationary-phase cultures. The transcription of adhA is elevated by hypoxia and under anoxic conditions, and we show that it is regulated by the Anr transcription factor. We have shown that lasR mutants, which lack an important quorum sensing regulator, have higher levels of Anr-regulated transcripts under low-oxygen conditions than their wild-type counterparts. Here, we show that a lasR mutant, when grown with ethanol, has an even larger decrease in pH than the wild type (WT) that is dependent on both anr and adhA. The large increase in AdhA activity is similar to that of a strain expressing a hyperactive Anr-D149A variant. Ethanol catabolism in P. aeruginosa by AdhA supports growth on ethanol as a sole carbon source and electron donor in oxygen-limited settings and in cells growing by denitrification under anoxic conditions. This is the first demonstration of a physiological role for AdhA in ethanol oxidation in P. aeruginosa. IMPORTANCE Ethanol is a common product of microbial fermentation, and the Pseudomonas aeruginosa response to and utilization of ethanol are relevant to our understanding of its role in microbial communities. Here, we report that the putative alcohol dehydrogenase AdhA is responsible for ethanol catabolism and acetate accumulation under low-oxygen conditions and that it is regulated by Anr.

Pseudomonas aeruginosa lasR mutant fitness in microoxia is supported by an Anr-regulated oxygen-binding hemerythrin

“…P. aeruginosa is especially damaging to the lungs of individuals with the genetic disease cystic fibrosis (CF), where it establishes chronic infections of the airway and is a major predictor of morbidity and mortality (1,2). The success of P. aeruginosa in disease is due to a confluence of factors, including intrinsic and acquired antibiotic resistance (3), production of a battery of secreted, virulence-associated molecules (4), the ability to form antibiotic-and immune cell-recalcitrant biofilms on biotic surfaces and implanted devices (5), and versatile metabolic capabilities (6,7), such as a pronounced ability to grow in the hypoxic or anoxic conditions engendered by biofilms and chronic infections (8,9). P. aeruginosa utilizes quorum sensing (QS) to coordinate the expression of a broad set of genes involved in virulence and nutrient acquisition (10).…”

“…Anr and its homologs have been well-characterized as oxygen-sensitive transcription factors (21)(22)(23). The P. aeruginosa Anr regulon includes genes encoding enzymes involved in microoxic and anoxic respiration, fermentation, microoxic ethanol oxidation, CupA fimbriae synthesis, and a number of hypothetical proteins including a putative hemerythrin PA14_42860 (PA1673) (7,(24)(25)(26).…”

Conditional antagonism in co-cultures of Pseudomonas aeruginosa and Candida albicans: An intersection of ethanol and phosphate signaling distilled from dual-seq transcriptomics

“…While we did not elucidate the mechanism by which ethanol influences PhoB activity, we identified ExaA-based ethanol catabolism as a potential regulator of the PhoB-mediated ethanol response including 5-MPCA production [94,95]. Ethanol induction of PhoB did not require either acetyl phosphate or ppGpp (Table 1), two pathways previously linked to ethanol catabolism [96,31], and thus we speculate that the effects could be multifactorial and may involve conditions such as changes in NAD(P)H/NAD(P)+ ratios, the generation of acetaldehyde or acetate as metabolic intermediates, or other roles played by enzymes involved in ethanol oxidation [97].…”

Conditional antagonism in co-cultures ofPseudomonas aeruginosaandCandida albicans: an intersection of ethanol and phosphate signaling distilled from dual-seq transcriptomics

“…By analyzing eADAGE node activity, we were able to identify subtle signals for which genes did not individually meet the cutoffs for DEGs and whose pathways were not detected through over-representation analysis. Subtle changes in gene expression could still have dramatic phenotypic effects; for example, although the ethanol catabolism genes were not included in any enriched pathway, they were present in eADAGE differentially active signatures, and we identified ExaA-based ethanol catabolism as a potential regulator of the PhoB-mediated ethanol response including 5-MPCA production [91, 92]. Similarly, we observed enrichment of genes involved in isoprenoid metabolism which may be indicative of P. aeruginosa response to C. albicans -produced farnesol [18, 28, 93].…”