In the Salmonella typhi genome, there are three “pathogenicity” islands. One of them, the 118kb main pathogenicity island, contains a Type IV pil operon. Because of high homology between the S. typhi pil operon and that of the Escherichia coli K-12 R64 plasmid, functions of the pil genes may be predicted. In the S. typhi pil gene cluster, there are four noncoding regions: the region upstream of pilL, intragenic regions between pilM and pilN and between pilN and pil0, and the region upstream of rci. Previous studies showed that the region upstream of pilL should contain a promoter to drive pil operon expression....[ Read more ]

In the Salmonella typhi genome, there are three “pathogenicity” islands. One of them, the 118kb main pathogenicity island, contains a Type IV pil operon. Because of high homology between the S. typhi pil operon and that of the Escherichia coli K-12 R64 plasmid, functions of the pil genes may be predicted. In the S. typhi pil gene cluster, there are four noncoding regions: the region upstream of pilL, intragenic regions between pilM and pilN and between pilN and pil0, and the region upstream of rci. Previous studies showed that the region upstream of pilL should contain a promoter to drive pil operon expression.

Two chromosomal insertion cassettes were generated with catechol 2,3-dioxygenase as the reporter gene. In these cassettes, either the region upstream of pilL or the region upstream of rci was cloned in front of the promoterless reporter gene.

It was found first that both the pilL and rci promoters have different expression levels during log phase, late-log phase and stationary phase. Initial expression occurs at late-log phase, and maximum expression during the transition to stationary phase. Activities in log phase were extremely low. Next this study shows that environmental factors, such as osmolarity and pH, affect the expression of the pilL and rci promoters. It was found that the optimum concentration of NaCl for pilL promoter expression was at l00mM, whereas that for rci promoter expression was at 200mM. Both promoters showed decreasing activities with increasing salt concentration. In an investigation of pH effects, pilL promoter expression was shown to be pH-dependent with highest activities at pH 7, while the rci promoter was found to be "pH-independent”, i.e. no obvious activity change with different pH conditions.

Further work was conducted to investigate possible influences of some two-component regulators upon the activity of the two promoters through the generation of some insertion mutant strains: envZ/ompR^-, phoPQ^-, pmrAB^-, rcsCB^-, rpoS^-, sirA^- and ssrAB^-. Among these regulators, rcsCB^- and envZ/ompR^- mutants showed significant drops in activities of both the pilL and rci promoters. In addition, stationary-phase dependent expression was shown to be rpoS-independent due to negligible differences in reporter gene activity measured in wild type and rpoS^- strains.