Investigation of Combinatorial Gene Regulation in Saccharomyces Species

Transcriptional control of gene expression is a result of complex interactions between the cis-regulatory elements (CRE) at gene promoters. To understand the regulatory logic of a cell, we need to identify the CRE combinations that regulate gene expression. This dissertation describes a sensitive computational method to identify phylogenetically conserved CRE combinations for any species of interest. In contrast to previous methods, I do not need to align genomes to identify these combinations. I applied the method in 7 sensu stricto and sensu lato Saccharomyces species. 80% of the predictions displayed some evidence of combinatorial transcriptional behavior in several existing datasets including 1) ChIP-chip data for co-localization of transcription factors, 2) gene expression data for co-expression of predicted regulatory targets, and 3) gene ontology databases for common pathway membership of predicted regulatory targets. To establish definitive evidence that these CRE interactions influence TF occupancy, I performed ChIP-Seq experiments on transcription factors in a wild-type strain and strains in which a predicted cofactor was deleted. These experiments showed that TF occupancy at the promoters of the CRE combination target genes depends on the predicted cofactor while occupancy of other promoters is independent of the predicted cofactor. In addition to identifying phylogenetically conserved CRE combinations, the method can annotate potential regulatory differences between species. Previous studies of cis-regulatory rewiring between species assume that CREs act independently and have found that promoter divergence does not necessarily explain expression divergence. By analyzing the S. cerevisiae and S. bayanus genomes, I identified differences in combinatorial cis-regulation between the species and showed that the predicted changes in gene regulation explain several of the species-specific differences seen in gene expression datasets. In some instances, the same CRE combinations appear to regulate genes involved in distinct biological processes in the two different species. The results of this research demonstrate 1) that combinatorial cis-regulation can be inferred from similarities between species and 2) that combinatorial cis-regulation can explain differences between species