Transcriptional profiling reveals molecular basis and novel genetic targets for improved resistance to multiple fermentation inhibitors in Saccharomyces cerevisiae.

Yingying Chen,Na Wei,Xueyang Feng,Joseph Stevens,Jiayuan Sheng,Tao Jiang

doi:10.1186/s13068-015-0418-5

Yingying Chen, Na Wei + Show 4 more

Open Access

https://doi.org/10.1186/s13068-015-0418-5

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Abstract

BackgroundLignocellulosic biomass is a promising source of renewable biofuels. However, pretreatment of lignocellulosic biomass generates fermentation inhibitors that adversely affect the growth of industrial microorganisms such as Saccharomyces cerevisiae and prevent economic production of lignocellulosic biofuels. A critical challenge on developing S. cerevisiae with improved inhibitor resistance lies in incomplete understanding of molecular basis for inhibitor stress response and limited information on effective genetic targets for increasing yeast resistance to mixed fermentation inhibitors. In this study, we applied comparative transcriptomic analysis to determine the molecular basis for acetic acid and/or furfural resistance in S. cerevisiae.ResultsWe recently developed a yeast strain YC1 with superior resistance to acetic acid, furfural, and their mixture through inverse metabolic engineering. In this study, we first determined transcriptional changes through RNA sequencing in YC1 versus the wild-type strain S-C1 under three different inhibitor conditions, including acetic acid alone, furfural alone, and mixture of acetic acid and furfural. The genes associated with stress responses of S. cerevisiae to single and mixed inhibitors were revealed. Specifically, we identified 184 consensus genes that were differentially regulated in response to the distinct inhibitor resistance between YC1 and S-C1. Bioinformatic analysis next revealed key transcription factors (TFs) that regulate these consensus genes. The top TFs identified, Sfp1p and Ace2p, were experimentally tested as overexpression targets for strain optimization. Overexpression of the SFP1 gene improved specific ethanol productivity by nearly four times, while overexpression of the ACE2 gene enhanced the rate by three times in the presence of acetic acid and furfural. Overexpression of SFP1 gene in the resistant strain YC1 further resulted in 42 % increase in ethanol productivity in the presence of acetic acid and furfural, suggesting the effect of Sfp1p in optimizing the yeast strain for improved tolerance to mixed fermentation inhibitor.ConclusionsTranscriptional regulation underlying yeast resistance to acetic acid and furfural was determined. Two transcription factors, Sfp1p and Ace2p, were uncovered for the first time for their functions in improving yeast resistance to mixed fermentation inhibitors. The study demonstrated an omics-guided metabolic engineering framework, which could be developed as a promising strategy to improve complex microbial phenotypes.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-015-0418-5) contains supplementary material, which is available to authorized users.

Highlights

IntroductionPretreatment of lignocellulosic biomass generates fermentation inhibitors that adversely affect the growth of industrial microorganisms such as Saccharomyces cerevisiae and prevent economic production of lignocellulosic biofuels
Lignocellulosic biomass is a promising source of renewable biofuels
29 genes involved in transmembrane transport were differentially expressed in a pool of totally 192 genes that were differentially expressed in S-C1, while 9 genes were differentially expressed in a pool of totally 46 genes that were differentially expressed in YC1. These results indicated that the resistant strain YC1 had altered genetic regulatory networks and applied distinct molecular mechanisms from the wildtype strain to achieve improved stress response to acetic acid and/or furfural

Summary

Introduction

Pretreatment of lignocellulosic biomass generates fermentation inhibitors that adversely affect the growth of industrial microorganisms such as Saccharomyces cerevisiae and prevent economic production of lignocellulosic biofuels. Lignocellulosic biomass materials need to undergo harsh (physico)chemical treatment designed to release sugar compounds [5, 6], but at the same time, the hydrolysis pretreatment generates toxic byproducts such as weak acids, furan aldehydes, and phenolic compounds (referred to as “fermentation inhibitors”) [7,8,9]. A robust inhibitor resistance fermenting microorganism is critically important for developing economically viable lignocellulosic biofuels, but this remains a major technical barrier [9]. Two major groups of fermentation inhibitors generated from pretreatment of lignocellulosic biomass are weak acids (e.g., acetic acid and formic acid) and furan aldehydes [e.g., furfural and 5-hydroxy methylfurfural (HMF)] [9, 10]. Furfural and HMF are major byproducts generated from hydrolysis and dehydration of pentose and hexose sugars [9, 10]

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