Project Summary/Abstract The rewiring of transcriptional circuits over evolutionary time is a major source of biological novelty. This proposal seeks to determine the detailed molecular mechanisms that underlie transcriptional rewiring using unicellular yeasts as a model system. The strategy is based on direct experimentation in many different yeast species in the Saccharomycetaceae lineage, and utilizes genome-wide transcriptional profiling, chromatic immunoprecipitation, phylogenetic comparisons, and ancestral protein reconstructions. Circuit comparisons among these yeasts uncover specific examples of transcriptional rewiring, and deeper analyses reveal the molecular mechanisms by which the wiring changes occurred. Although much rewiring is probably neutral, some of it appears adaptive: indeed a major mechanism for evolutionary novelty involves rewiring transcriptional circuitry to allow new expression patterns of existing gene products. Thus, to truly understand the structures of transcription circuits in modern species, we need to know the mechanisms by which they rapidly evolve and how these mechanisms lead to and, thereby can account for, modern structures.