Methods and approaches for regulating gene expression are central to the success of metabolic engineering. In many cases, an inducible transcription factor or repression system is used to control the expression of one or more target genes. In yeast, a range of such systems are available, but activation of these systems requires a change in culture media, carbon source, or the addition of a costly chemical ligand. Here, we present a set of orthogonal synthetic transcription factors that can be used in both gene activation and repression formats and that are inducible by low cost chemicals. These new systems are based on the plant abscisic acid (ABA) hormone signaling pathway where ABA binds to the PYR1 receptor, which in turn forms a tertiary complex and inhibits HAB1, a PP2C phosphatase. This chemical induced dimerization (CID) system is amenable to engineering, and through the fusion of DNA binding, activator, and repressor domains can be converted from a signaling pathway to a system for inducible gene activation or repression using ABA or the agrochemical mandipropamide. In this talk, we present the optimization of this new gene regulation system; demonstrate use in S. cerevisiae as well as the non-conventional yeasts Kluyveromyces marxianus, and Yarrowia lipolytica; and, use the system as a means to control central pyruvate metabolism in K. marxianus by simultaneously activating and repressing pyruvate decarboxylase and pyruvate dehydrogenase. This new synthetic transcription factor system opens a new approach to controlling gene expression that can be broadly used across various yeast species.

Designing novel transcription factors for inducible gene expression in yeast

ACS Spring 2022

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