Anaerobic regulation of transcription initiation in the arcDABC operon of Pseudomonas aeruginosa

Abstract

The arcDABC operon of Pseudomonas aeruginosa encodes the enzymes of the arginine deiminase pathway, which is inducible under conditions of oxygen limitation and serves to generate ATP from arginine. The 5' end of arc mRNA extracted from anaerobically grown cells was determined by Si and primer extension mapping. The transcription initiation site was located upstream of the arcD gene and 41.5 bp downstream of the center of the sequence TTGAC....ATCAG. This sequence, termed the ANR box, is similar to the consensus FNR recognition site of Escherichia coli. Transcription of the arc operon in P. aeruginosa was strongly decreased by a deletion of the TTGAC half site or by a mutation in the anr gene, which is known to code for the FNR-like regulatory protein ANR. During a transition from aerobic to anaerobic growth conditions, the concentrations of arc mRNAs and the levels of the ArcD and ArcA proteins rose in a parallel fashion. Mutational analysis of the arc promoter region led to the conclusion that the distance between the ANR box and the -10 promoter region is important for promoter strength, whereas the -35 region does not appear to be critical for arc promoter function. These findings and previous results indicate that anaerobic induction of the arc operon occurs at the level of transcription and requires the ANR box in cis and the ANR protein in trans.To obtain metabolic energy, Pseudomonas aeruginosa preferentially uses aerobic respiration. Under oxygen-limiting conditions, in the presence of nitrate or nitrite, P. aeruginosa switches to anaerobic respiration (9,37). In the absence of terminal electron acceptors, arginine can be used as the sole energy source (52). Anaerobic degradation of arginine to ornithine produces 1 mol of ATP per mol of arginine and depends on the three enzymes of the arginine deiminase pathway: arginine deiminase (ADI), catabolic omithine carbamoyltransferase, and carbamate kinase (Fig.

Transcriptome Analysis of the ArgR Regulon inPseudomonas aeruginosa

“…However, the presence of additional arginine transport systems was evidenced by the reduced, but still inducible, arginine uptake of the aot mutant (35). The arcD gene, which encodes an arginine:ornithine antiporter, is induced under anaerobic conditions from the arcDABC operon (8). Many new members of the ArgR regulon that encode polypeptides for solute transport are inducible by arginine: the PA5152-5155 operon, PA3934, PA2042, PA1971, and AdcB for a putative arginine:agmatine antiporter.…”

“…While Anr, the anaerobic regulatory protein (8), is essential for induction of the arcDABC operon, which encodes an arginine:ornithine antiporter and enzymes of the ADI pathway, exogenous arginine can further induce its expression level through the interactions of Anr and ArgR (28,38).…”

Transcriptional Control of the Hydrogen Cyanide Biosynthetic Genes hcnABC by the Anaerobic Regulator ANR and the Quorum-Sensing Regulators LasR and RhlR in Pseudomonas aeruginosa

“…Severe oxygen limitation was achieved by growing the cells in tightly closed 125-ml bottles containing 60 ml of MMC, with gentle shaking; at the end of growth, the oxygen present initially was consumed by the cells. Antibiotics were used at the concentrations (in micrograms per milliliter) indicated in parentheses: for P. aeruginosa strains, carbenicillin (250), chloramphenicol (250), gentamicin (10), tetracycline (125), and mercuric chloride (10); for P. fluorescens, gentamicin (10) and tetracycline (125); and for E. coli, ampicillin (100), chloramphenicol (25), gentamicin (10), and tetracycline (25). 5-Bromo-4-chloro-3-indolyl-␤-D-galactoside (X-Gal) was incorporated into solid media at 0.02% to monitor ␤-galactosidase expression (49).…”

arcD, the first gene of the arc operon for anaerobic arginine catabolism in Pseudomonas aeruginosa, encodes an arginine-ornithine exchanger

“…The arcABC genes have been cloned and sequenced (2, 3) and are organized in an operon (24,30). The arc operon is under control of a positive regulatory protein, ANR, an FNR-like protein of P. aenrginosa (16,18,37) …”

“…The arcABC genes have been cloned and sequenced (2,3) and are organized in an operon (24,30). The arc operon is under control of a positive regulatory protein, ANR, an FNR-like protein of P. aenrginosa (16,18,37), which is thought to activate transcription of the arc genes during oxygen limitation (26). The proximal part of the arc operon harbors the arcD gene (23).…”