New transcriptional circuit evolved by coding sequence changes in a master regulator followed by cis-regulatory changes in its target genes

While changes in both the coding-sequence of transcriptional regulators and in the cis-regulatory sequences recognized by them have been implicated in the evolution of transcriptional circuits, little is known of how they evolve in concert. We describe an evolutionary pathway in fungi where a new transcriptional circuit (a-specific gene repression by Mat2) evolved by coding changes in an ancient master regulator, followed millions of years later by cis-regulatory sequence changes in the genes of its future regulon. We discerned this order of events by analyzing a group of species in which the coding changes in the regulator are present, but the cis-regulatory changes in the target genes are not. In this group we show that the coding changes became necessary for the regulators deeply conserved function and were therefore preserved. We propose that the changes first arose without altering the overall function of the regulator (although changing the details of its mechanism) and were later co-opted to "jump start" the formation of the new circuit.