The spv region of SalmoneUla virulence plasmids is essential for the development of a systemic infection in mice.
The sigma S-regulated gene spvR of Salmonella typhimurium encodes an autoregulatory protein required for transcriptional activation of the virulence operon spvABCD. A mutation in the histone-like protein H-NS, which negatively controls the sigma S level, has been reported to increase spv gene expression in S. typhimurium strain LT2. In agreement with this, we found that transcription of spvR and spvABCD was derepressed in hns strains of Escherichia coli and S. typhimurium. Moreover, levels of spv gene expression in hns rpoS double mutants were higher than expression levels in mutants deficient in rpoS alone, and were close to those measured in wild-type strains. This demonstrates that H-NS contributes to spv gene regulation independently of its function in controlling the sigma S level. Since the same start site was used for spvR gene transcription in wild-type as in hns and hns rpoS mutant strains, it is likely that the spvR promoter. spvRp1, can be recognized efficiently by an RNA polymerase containing sigma 70. The spvR promoter region shows an intrinsic DNA curvature that might be a determinant in H-NS- and/or sigma S-mediated control. A single amino acid substitution, Leu to Pro at position 265, abolished the regulatory function of SpvR in E. coli and Salmonella, implicating the C-terminal domain of SpvR in its structure and/or regulatory function. The spvR265 allele is not transcribed at detectable levels in hns or hns rpoS strains, suggesting that activation of spvRp1 in these strains remains dependent on SpvR. Thus, we propose a model for spvR gene regulation in which SpvR acts as a co-regulator of an RNA polymerase containing either sigma 70 (in the absence of H-NS) or sigma S, to induce transcriptional initiation at spvRp1. Moreover, growth-phase regulation of spv gene expression was maintained in hns and hns rpoS strains, indicating that an additional element, besides sigma S, is involved in the growth-phase regulation in rich medium.
The SpvR protein is a DNA-binding protein of the LysR family, required for the transcription of the spvABCD virulence operon of Salmonella typhimurium. An alternative sigma factor, sigma S (RpoS), in conjunction with SpvR, controls the transcription of the spvR gene. In this study, we used a combination of primer extension experiments and deletion/fusion analyses of the spvR gene to identify sequences involved in spvR transcription in S. typhimurium. When induced in the stationary phase of growth in rich medium or during carbon starvation, transcription of spvR in S. typhimurium is driven by a single promoter (spvRp1) and initiates 17 nucleotides upstream of the spvR start codon. The level of spvR transcription originating at spvRp1 was 20-fold higher in the wild-type strain than in the rpoS mutant. In both strains, however, transcription at spvRp1 requires the SpvR protein. 5' Deletions up to position -86, relative to the spvR start codon, did not inhibit inducibility by sigma S and/or SPVR. In contrast, 5' deletion up to -75 abolished the activation of spvRp1 by SpvR in both the wild-type strain and rpoS mutant. Within the 11-bp sequence lying between position -86 and position -75, a 10-bp consensus motif TNTNTGCANA, present in both the spvR and spvA promoter regions, was identified and may contain the DNA recognition site for SpvR. In addition, we detected initiation of transcription within the spvR coding region. This finding may have implications for comparative studies of regulation with spvR gene fusions.
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