Investigating the Physiological Effects of Mutations in the Proposed Backtrack Sit of Yeast RNA Polymerase II

Single page posterTranscription, the first step of gene expression, is a process fundamental to all known forms of life. In eukaryotic cells, the enzyme RNA polymerase II (Pol II) executes transcription by moving forward along the DNA and transferring the encoded genetic information to messenger RNA. However, Pol II also backtracks on the DNA, causing transcription to become arrested. When backtracking occurs, Pol II slides backwards on the DNA, displacing nascent RNA from the active site into a proposed “backtrack site,” comprising residues in the Pol II subunits Rpb1 and Rpb2 that interact with the RNA. The resulting stable “arrested complex” must be reactivated for elongation to continue. While backtracking has been implicated in numerous processes essential for regulating gene transcription, its physiological relevance is not yet certain. Using Saccharomyces cerevisiae (Baker’s yeast), we have engineered individual and combinations of mutations in the Rpb1 region of the backtrack site to disrupt the protein-RNA interactions that arise from backtracking. Through phenotypic and growth comparisons between wildtype and mutant strains, we examine how impairing the binding of RNA to the backtrack site affects yeast fitness and various Pol II functions in vivo, providing further insight into the possible functions of Pol II during backtracking and arrest.Research reported in this poster was supported by the Eunice Kennedy Shriver National Institute of Child Health & Human Development of the National Institutes of Health under award number R25HD0708