Yeast zinc cluster proteins paralogs involved in the switch from fermentation to respiration show interdependency for DNA binding revealing a novel type of DNA recognition

In budding yeast, fermentation is the most important pathway for energy production. Under low glucose, ethanol is used for synthesis of this sugar requiring a shift to respiration. This process is controlled by the transcriptional regulators Cat8, Sip4, Rds2, and Ert1. We characterized Gsm1, a paralog of Rds2 and Ert1. Genome-wide analysis showed that Gsm1 has a DNA binding profile highly similar to Rds2, suggesting that these factors bind to DNA as heterodimers. Indeed, binding of Gsm1 and Rds2 is interdependent at the gluconeogenic gene FBP1. However, Rds2 is required for Gsm1 to bind at other promoters but not the reverse. Gsm1 and Rds2 also bind to DNA independently of each other. We showed that the DNA binding domains of Gsm1 and Rds2 bind cooperatively in vitro to the FBP1 promoter. In contrast, at the HAP4 gene, Ert1 cooperates with Rds2 for DNA binding. Mutational analysis suggests that Gsm/Rds2 and Ert1/Rds2 bind to short common DNA stretches revealing a novel mode of binding for this class of factors. Two-point mutations in a HAP4 site convert it to a Gsm1 binding site. In summary, our observations add another layer of transcriptional regulation with the formation of different heterodimers at specific promoters. We profiled Rds2 and Gsm1 proteins, both tagged with HA, in yeast Saccharomyces cerevisiae cells by ChIP-chip on tiling arrays. The Rds2 protein was profiled in WT and gsm1Δ cells, whereas the Gsm1 protein was profiled in WT and rds1Δ cells. All HA-tagged ChIP samples were labeled with Cy5 and hybridized against no tag ChIP samples (labeled with Cy3). The microarrays used were described before (Jeronimo and Robert, 2014). The data were normalized using the Limma Loess method, and replicates were combined as described previously (Ren et al. 2000). The data were subjected to one round of smoothing using a Gaussian sliding window with a standard deviation of 100 bp to generate data points in 10-bp intervals (Guillemette et al. 2005). Each experiment was done in duplicate. Please note that each bed file generated from both replicates is linked to the corresponding *rep1 sample records.