A synthetic hybrid promoter for xylose-regulated control of gene expression in Saccharomyces yeasts

Authors: Hector, Ronald - Ron; Mertens, Jeffrey

Submitted to: Molecular Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/15/2016
Publication Date: 1/30/2017
Publication URL: https://handle.nal.usda.gov/10113/5695424
Citation: Hector, R.E., Mertens, J.A. 2017. A synthetic hybrid promoter for xylose-regulated control of gene expression in Saccharomyces yeasts. Molecular Biotechnology. 59(1):24-33. doi: 10.1007/s12033-016-9991-5.

Interpretive Summary: Metabolic engineering of brewer’s yeast (Saccharomyces cerevisiae) to efficiently convert biomass-derived sugars such as xylose to fuels and chemicals will require the expression of multiple genes. Expressing genes at constant high levels can be detrimental to the cell, resulting in decreased cell growth and productivity. In the case of engineering cells to use xylose, it would not be beneficial to express all of the required genes constitutively at elevated levels. To address this issue, new synthetic promoters were developed for brewer’s yeast that are repressed in the absence of xylose and induced when xylose is available. When xylose was present, the new promoters had activity comparable to other yeast promoters for high-level expression. Furthermore, the promoters were not active in the absence xylose and up to a 25-fold increase in expression was seen when xylose was added. These new promoters provide improved control of gene expression for engineered brewer’s yeast and are a starting point for generating new promoters with additional types of regulation, increased repression, and more tunable expression levels.

Technical Abstract: Metabolism of non-glucose carbon sources is often highly regulated at the transcriptional and post-translational levels. This level of regulation is lacking in Saccharomyces cerevisiae strains engineered to metabolize xylose. To better control transcription in S. cerevisiae, the xylose-dependent, DNA-binding repressor (XylR) from Caulobacter crescentus was used to block transcription from synthetic promoters based on the constitutive Ashbya gossypii TEF promoter. The new hybrid promoters were repressed in the absence of xylose and showed up to a 25-fold increase in the presence of xylose. Activation of the promoter was also highly sensitive to xylose with activity seen at concentrations below 2 uM xylose. These new xylose inducible promoters allow improved control of gene expression for engineered strains of Saccharomyces yeasts.