Article: article from journal or magazin.
RNA processing modulates the expression of the arcDABC operon in Pseudomonas aeruginosa.
Journal of Molecular Biology
Anaerobic growth of Pseudomonas aeruginosa on arginine depends on the arcDABC operon encoding the enzymes of the arginine deiminase pathway. The co-ordinate, anaerobic induction of these enzymes requires the FNR-like regulatory protein ANR, which activates the arc promoter lying upstream from arcD. By Northern hybridization experiments, three abundant arcA, arcAB and arcABC transcripts and three minor arcDA, arcDAB and arcDABC transcripts could be detected. The 5' ends of the arcA, arcAB and arcABC mRNAs were determined by S1 and primer extension mapping. These 5' ends appear to be generated by endonucleolytic cleavage (processing) in arcD mRNA rather than by a second promoter; this was concluded from the effects of insertion and deletion mutations in arcD. Intergenic inverted repeats between arcA and arcB as well as between arcB and arcC were shown to be involved in the formation of 3' ends of arc transcripts. Deletion of either intergenic region in the P. aeruginosa chromosome led to the loss of the arcA or arcAB transcript, respectively. Dot blot experiments revealed that arc mRNAs extracted from the wild-type strain had similar chemical half-lives in the arcA, arcB and arcC regions, ranging from 16 to 13 minutes. The half-life of arcD mRNA, by contrast, was significantly shorter, suggesting that this mRNA segment may be destabilized by the processing cuts within arcD. Deletion of the putative intergenic stem-loop structures did not result in a dramatic loss of arc mRNA stability. Thus, the intergenic hairpin structures do not contribute importantly to the overall mRNA stability; they might act primarily as partial transcription terminators and locally protect the 3' ends from exonuclease action. The expression levels of the four Arc proteins correlated approximately with the relative abundance of the corresponding mRNA segments. In conclusion, mRNA processing and, presumably, partial termination of transcription contribute to differential gene expression within the arc operon.
Amino Acid Transport Systems, Antiporters, Arginine/metabolism, Bacterial Proteins/genetics, Bacterial Proteins/metabolism, Base Sequence, DNA Mutational Analysis, Gene Expression Regulation, Bacterial, Genes, Bacterial, Hydrolases/genetics, Hydrolases/metabolism, Membrane Proteins/genetics, Membrane Proteins/metabolism, Molecular Sequence Data, Operon, Ornithine Carbamoyltransferase/genetics, Ornithine Carbamoyltransferase/metabolism, Phosphotransferases/genetics, Phosphotransferases/metabolism, Phosphotransferases (Carboxyl Group Acceptor), Promoter Regions, Genetic, Pseudomonas aeruginosa/genetics, Pseudomonas aeruginosa/metabolism, RNA Precursors/metabolism, RNA Processing, Post-Transcriptional, Repetitive Sequences, Nucleic Acid/genetics
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