IDENTIFICATION OF SALMONELLA-TYPHI PROMOTERS ACTIVATED BY INVASION OFEUKARYOTIC CELLS

The interaction of pathogenic microorganisms with host tissues, and th e underlying genetic events which regulate these interactions, are dif ficult to analyse where no suitable animal model exists, The approach described here, for obtaining information on the genes involved in the se interactions, employs an infection system based on the invasion of Henle cells by Salmonella typhi to select promoter-containing DNA sequ ences able to activate gene expression inside eukaryotic cells, Severa l DNA fragments exhibiting different promoter strengths and extent of selective activation within eukaryotic cells were identified. Three we re selected and characterized according to the expression level of the reporter gene, the polynucleotide sequence, the transcription start, and the dependence upon regulatory proteins, All fragments gave much s tronger expression of the reporter gene when the recombinant S. typhi carrier strains invaded cells compared with the expression measured in growth medium, One promoter-containing region exhibited sequence homo logy to sigma(54)-dependent promoters, whereas another appears to be d ependent on the stationary-phase RNA polymerase subunit sigma(s), S. t yphi containing the S1 subunit gene of pertussis toxin cloned under th e control of these promoters, selectively expressed the S1 subunit fol lowing infection of different phagocytic and non-phagocytic cell lines of human or murine origin, Deletion and point mutant derivatives of t wo promoters enabled the identification of the main motif required for promoter activity. This method may be helpful for the analysis of pat hogenesis in organisms previously difficult to study because of the la ck of a convenient animal model, and could provide insights into the c hronology and topology of gene expression during infection, including a possible genetic basis for tissue tropism.