Transcription factor Reb1 is required for proper transcriptional start site usage at the divergently transcribed TFC6-ESC2 locus in Saccharomyces cerevisiae

Abstract

Eukaryotic promoters generally contain nucleosome depleted regions near their transcription start sites. In the model organism Saccharomyces cerevisiae, these regions are adjacent to binding sites for general regulatory transcription factors, and the Reb1 protein is commonly bound to promoter DNA near such regions. The yeast TFC6 promoter is a unique RNA polymerase II promoter in that it is autoregulated by its own gene product Tfc6p, which is part of the RNA polymerase III transcription factor complex TFIIIC. We previously demonstrated that mutation of a potential Reb1 binding site adjacent to the TFIIIC binding site in the TFC6 promoter modestly reduces transcript levels, but leads to a severe decrease in Tfc6 protein levels due to an upstream shift in the TFC6 transcription start site. Here we confirm that Reb1p indeed binds to the TFC6 promoter, and is important for proper transcription start site selection and protein expression. Interestingly, loss of Reb1p association at this site has a similar effect on the adjacent divergently transcribed ESC2 promoter, resulting in a significant increase of 5′-extended ESC2 transcripts and reduction of Esc2 protein levels. This altered divergent transcription may be the result of changes in nucleosome positioning at this locus in the absence of Reb1p binding. We speculate that an important function of general regulatory factors such as Reb1p is to establish and maintain proper transcription start sites at promoters, and that when binding of such factors is compromised, resulting effects on mRNA translation may be an underappreciated aspect of gene regulation studies.