The cis requirements for transcriptional activation by HilA, a virulence determinant encoded on SPI-1

In several models of pathogenesis, Salmonella requires genes encoded on Salmonella pathogenicity island 1 (SPI-1) for virulence. In Salmonella enterica serovar Typhimurium (S. typhimurium), most SPI-1 genes are arranged in operons and are co-ordinately regulated in response to environmental signals via the SPI-1-encoded protein HilA. In order to understand how HilA controls the transcription of SPI-1 genes, we have analysed the invF and prgH promoters. We have reconstituted HilA-dependent activation of both promoters in Escherichia coli by supplying hilA on a plasmid, strongly suggesting that HilA acts directly on the promoters. By analysing the HilA-dependent activity of deletions and mutations in P-invF, we identified cis elements necessary for HilA-dependent activation. Through biochemical studies, we have defined a probable HilA-binding sequence in the invF promoter. This 'HilA box' is intact in the minimal promoter identified through deletion analysis, and it is disrupted in one class of P-invF mutants that has reduced activation by HilA. The prgH promoter also contains a HilA box in the same position relative to the +1 of transcription. This work is the first to connect HilA-dependent environmental regulation with a specific sequence in a SPI-1 virulence gene promoter.