Transcriptomic Profiles of Zymomonas mobilis 8b to Furfural Acute and Long-Term Stress in Both Glucose and Xylose Conditions.

Shihui Yang,Philip T Pienkos,Yat-Chen Chou,Yun Hu,Xia Wang,Min Zhang,Mary Ann Franden,Steven D Brown

doi:10.3389/fmicb.2020.00013

Abstract

Zymomonas mobilis 8b is an ethanologenic bacterium engineered to utilize both glucose and xylose. The impacts of lignocellulosic hydrolyzate inhibitors on the growth of Zymomonas mobilis 8b have been investigated. However, the molecular responses of these inhibitors have not been completely elucidated yet. In this study, molecular responses to furfural were investigated using transcriptomic approaches of both chip-based microarray and a directional mRNA-Seq. Furfural acute shock time-course experiment with 3 g/L furfural supplemented when cells reached exponential phase and stress response experiment in the presence of 2 g/L furfural from the beginning of fermentation were carried out to study the physiological and transcriptional profiles of short-term and long-term effects of furfural on 8b. Furfural negatively affected 8b growth in terms of final biomass and the fermentation time. Transcriptomic studies indicated that the response of 8b to furfural was dynamic and complex, and differences existed between short-term shock and long-term stress responses. However, the gene function categories were similar with most down-regulated genes related to translation and biosynthesis, while the furfural up-regulated genes were mostly related to general stress responses. Several gene candidates have been identified and genetic studies indicated that expression of ZMO0465 and cysteine synthase operon ZMO0003-0006 driven by its native promoter in a shuttle vector enhanced the furfural tolerance of 8b. In addition, the relationship between microarray and mRNA-Seq was compared with good correlations. The directional mRNA-Seq data not only provided the gene expression profiling, but also can be applied for transcriptional architecture improvement to identify and confirm operons, novel transcripts, hypothetical gene functions, transcriptional start sites, and promoters with different strength.

Highlights

Lignocellulosic biomass is an abundant renewable and sustainable resource, which is considered as an excellent source of sugars for microbial conversion into liquid fuels and valuable biochemicals
Since most biomass hydrolyzate contains a large amount of both glucose and xylose, and xylose has been identified as a stressor for Z. mobilis (Mohagheghi et al, 2014), we examined the transcriptional profiles of Z. mobilis 8b with xylose-fermenting ability to both glucose and xylose in short-term and long-term furfural stress conditions in this study
Our results showed that the growth rates were dramatically affected by the addition of exogenous furfural when either glucose or xylose was used as the carbon source (Supplementary Figure S1)

Summary

Introduction

Lignocellulosic biomass is an abundant renewable and sustainable resource, which is considered as an excellent source of sugars for microbial conversion into liquid fuels and valuable biochemicals. Various inhibitory compounds with strong inhibition on hydrolytic enzymes and fermenting strains are generated due to the partial overdegradation of lignocellulose These inhibitors include weak acids (e.g., acetic acid), furan aldehydes such as furfural, and lignin degradation products (e.g., vanillin) (Jönsson et al, 2013). Among these compounds, furfural, derived from dehydration of pentose during biomass deconstruction, is one of the most prevalent inhibitors due to its high concentration in the hydrolyzates and strong toxicity to microorganisms (Heer and Sauer, 2008; Wierckx et al, 2011; Yang et al, 2018a). To make economical lignocellulosic biofuels, it is very important to develop robust strains with high titer, yield and productivity in the presence of furfural and other inhibitors, and numerous efforts have been devoted to meeting this goal (Mills et al, 2009)

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Conclusion