The stationary phase expression of many conserved, adaptive bacterial proteins is dependent on RpoS, a second vegetative sigma factor. The regulation of RpoS itself, however, is complex and not fully understood, particularly at the level of transcription. In this report, we show that the observed hydrogen peroxide sensitivity of a mutant defective in expression of barA, a bacterial virulence factor, can be explained by a reduction in catalase activity, an RpoS-controlled function. Levels of katE mRNA, encoding the major catalase of Escherichia coli, were much lower in the barA mutant, suggesting that BarA is required for the expression of this RpoS-regulated gene. Expression of another RpoS-regulated gene, osmY, was also found to be severely reduced in the barA mutant. Employing Western analyses with anti-RpoS antisera and Northern analyses using probes specific for rpoS, we found that BarA is required for the exponential phase induction of RpoS itself. Operon lacZ fusion expression studies and Northern analyses indicate that BarA itself is maximally expressed in early exponential phase cultures immediately preceding the transcriptional induction of RpoS. Results of primer extension studies indicate that exponential phase expression from the rpoSp1 promoter is reduced by more than 85% in a barA mutant but could be efficiently complemented by a plasmid-borne copy of barA in trans. These results suggest that regulatory signals that are operant in exponentially growing cultures play an important role in effecting stationary phase gene expression.