Design and characterization of a dual-mode promoter with activation and repression capability for tuning gene expression in yeast

Modularity in controlling gene expression artificially is becoming an essential aspect of synthetic biology. Artificial transcriptional control of gene expression is one of the most well-developed methods for the design of novel synthetic regulatory networks. Such networksareintendedtohelpunderstandnaturalcellular phenomena and to enable new biotechnological applications. Promoter sequence manipulation with cis-regulatory elements is a key approach to control gene expression transcriptionally. Here, we have designed a promoter that can be both activated and repressed, as a contribution to the library of synthetic biological ‘parts’. Starting with the minimal cytochrome C (minCYC) promoter in yeast, we incorporated five steroid hormone responsive elements (SHREs) and one lac operator site, respectively, upstream and downstream of the TATA box. This allows activation through the testosterone-responsive androgen receptor, and repression through the LacI repressor. Exposure to varying concentrations of testosterone (to vary activation) and IPTG (to vary repression) demonstrated the ability to tune the promoter’s output curve over a wide range. By integrating activating and repressing signals, the promoter permits a useful form of signal integration, and we are optimistic that it will serve as a component in future regulatory networks, including feedback controllers.