Promoter substitution and deletion analysis of upstream region required for rpoS translational regulation

The RpoS sigma factor of enteric bacteria is required for the increased expression of a number of genes that are induced during nutrient limitation and growth into stationary phase and in response to high osmolarity. RpoS is also a virulence factor for several pathogenic species, including Salmonella typhimurium. The activity of RpoS is regulated at both the level of synthesis and protein turnover. Here we investigate the posttranscriptional control of RpoS synthesis by using rpoS-lac protein and operon fusions. Substitution of the native rpoS promoters with the tac or lac UV5 promoters allowed essentially normal regulation after growth into stationary phase in rich medium or after osmotic challenge. Regulation of these fusions required the function of hfq, encoding the RNA-binding protein host factor I (HF-I). Short deletions from the 5' end of the rpoS transcript did not affect regulation very much; however, a larger deletion mutation that still retains 220 bp upstream of the rpoS ATG codon, including a proposed antisense element inhibitory for rpoS translation, was no longer regulated by HF-I. Several models for regulation of rpoS expression by HF-I are discussed.