Controlling Multiple Gene Expression in Saccharomyces Cerevisiae By Optogenetic Switches

Our group is interested in the regulated production of small to large cohorts of proteins and peptides and the establishment of a toolkit for the modular assembly of circular expression yeast artificial chromosomes (xYAC) in S. cerevisiae. To simplify induction of protein production we will provide a reliable and easy to use induction system: Light-induced TALE-transcription factors based on two different light-sensing systems from Arabidopsis thaliana. While the CRY2/CIB1 system is blue light-sensitive and has no need for an exogenous cofactor, the red light-sensing PhyB/PIF3 interaction depends on a chromophore, which is not available in yeast. To avoid the exogenous supply of the chromophore, yeast will be engineered to express the necessary cofactor biosynthesis genes along with the red light-sensing proteins. In order to engineer two autonomous and non-interfering light-dependent gene switches, we will combine the red and the blue light-sensing systems on one single xYAC. With this approach it will be possible to easily regulate gene expression of a large number of proteins in a reversible, specific, dose-dependent and orthogonal manner totally independent of each other just by applying light of different wavelength. Results on the optimization of chromophore biosynthesis, evaluation of the individual light-induction systems, and the parallel use of red and blue light-controlled gene expression within our xYAC platform will be presented.