The acid-inducible asr gene in Escherichia coli: Transcriptional control by the phoBR operon

Escherichia coli responds to external acidification (pH 4.0 to 5.0) by synt hesizing a newly identified, similar to 450-nucleotide RNA component. At ma ximal levels of induction it is one of the most abundant small RNAs in the cell and is relatively stable bacterial RNA. The acid-inducible RNA was pur ified, and the gene encoding it, designated asr (for acid shock RNA), mappe d at 35.98 min on the E. coli chromosome, Analysis of the asr DNA sequence revealed an open reading frame coding for a Ill-amino-acid polypeptide with a deduced molecular mass of approximately 11.6 kDa, According to computer- assisted analysis, the predicted polypeptide contains a typical signal sequ ence of 30 amino acids and might represent either a periplasmic or an outer membrane protein, The asr gene cloned downstream from a T7 promoter was tr anslated in vivo after transcription using a T7 RNA polymerase transcriptio n system. Expression of a plasmid-encoded asr::lacZ fusion under a native a sr promoter was reduced similar to 15-fold in a complex medium, such as Lur ia-Bertani medium, versus the minimal medium. Transcription of the chromoso mal asr was abolished in the presence of a phoB-phoR (a two component regul atory system, controlling the pho regulon inducible by phosphate starvation ) deletion mutant. Acid-mediated induction of the asr gene in the Delta(pho B-phoR) mutant strain was restored by introduction of the plasmid with clon ed phoB-phoR genes. Primer extension analysis of the asr transcript reveale d a region similar to the Pho box (the consensus sequence found in promoter s transcriptionally activated by the PhoB protein) upstream from the determ ined transcription start. The asr promoter DNA region was demonstrated to b ind PhoB protein in vitro. We discuss our results in terms of how bacteria might employ the phoB-phoR regulatory system to sense an external acidity a nd regulate transcription of the asr gene.