Excited about UP elements
Now that our "Regulation of Sxy" manuscript is submitted, I can shift gear and get back to working on the A/T rich sequences in CRP-S promoters that I hypothesize work as UP elements. First some background about UP elements: bacterial RNA polymerases make two specific contacts with promoter DNA, one is the well-characterized recognition of the -10 and -35 sequences by the sigma subunit, and the second is the less-familiar recognition of upstream A/T rich sequences (UP elements) by RNAP's two alpha subunits.
All of H. influenzae's 13 CRP-S promoters have A/T rich sequences resembling UP elements, and I have shown that these sequences are required for transcription of the pilABCD operon. My working hypothesis is that CRP and Sxy act at CRP-S sites to bend DNA and bring the upstream UP elements close to RNAP so that the alpha subunits can make contact with the UP elements. To test this I am going to remove the CRP-S site and some flanking sequence from the promoter, thus bringing the putative UP elements closer to the -10,-35 RNAP binding site. I predict that the pil promoter will then be active in the absence of either CRP or Sxy. In other words, because I have cut up the DNA and positioned the UP elements physically adjacent to the -10,-35 site, CRP and Sxy are no no longer needed to bring the UP elements close to RNAP by bending the intervening DNA.