The mid- to late- 1980's were witness to a dramatic increase in the incidence of rheumatic fever and other severe and invasive diseases caused by the gram positive bacterium Streptococcus pyogenes. Reports in the popular media referring to S. pyogenes as the """"""""flesh eating bacterium"""""""" have served to focus attention to this problem, and to highlight our ignorance of the reasons for this increase. In particular, we need to better understand the molecular interactions that occur between S. pyogenes and host cells during infection. To address this issue, the proposed experiments will exploit the complementary interests and expertise of two groups: 1) Dr. M. Caparon (USA, PI) in molecular microbial genetics and pathogenesis, especially in relation to Streptococcus pyogenes and streptococcal diseases; and 2) Dr. E.D. Sverdlov (Russia) in eukaryotic gene expression, and nucleic acid chemistry. Specifically, Dr. Caparon's group will conduct infections of human skin cells with wild type and specific mutant S. pyogenes strains and prepare polyA+ mRNA from these infected cells. Dr. Sverdlov's group will then use this material to identify known and novel genes whose expression is modulated by streptococcal infection. To accomplish this they will use a new and innovative technique which they have pioneered called suppression subtractive hybridization (SSH). This SSH method overcomes the limitations of previous methods for subtractive hybridization and provides a greater than 1000-fold enrichment of low abundance transcripts, and also the enrichment of transcripts whose expression is regulated quantitatively, regardless of the basal abundance of the message. Studies following identification of such infection-regulated genes may contribute to the development of new therapies against S. pyogenes, and to curing or preventing diseases that it causes.
