| The evolution of sequences that control gene expression contributes significantly to the phenotypic diversification of organisms. Variation in the type, number, affinity and arrangement of transcription factor binding sites within a regulatory sequence determines when, where and how much of a transcript is generated. Based on the biochemical properties of a protein, its cellular role, and environmental context, there is a range of expression levels that the cell must have to survive and an optimal expression level that maximizes fitness. This output range constrains variation in regulatory sequences and pushes a promoter sequence to produce the optimal output. Here I discuss three projects where I examined how variability is constrained in functional transcriptional initiation complexes to better understand how transcriptional regulatory information is integrated and evolves. In the first, I developed a computational method to infer the physical constraints on the cooperative binding of Cbf1 and Met31 in yeast by quantifying how the arrangement of their sites are constrained in functional promoters. In the second, I experimentally determined the amount of functional variability within sigma70 --35 binding sites in different promoter and environmental contexts to understand how context constrains the evolution of binding sites. Finally in the third, I experimentally determined the amount of functional variability in the DNA-binding domain of the transcription factor MarA, to understand how both informational and energetic constraints affect transcription factor evolution. |
