Multi-omics analysis of fatty alcohol production in engineered yeasts Saccharomyces cerevisiae and Yarrowia lipolytica

Fatty alcohols are widely used in various applications within a diverse set of industries, such as the soap and detergent industry, the personal care and cosmetics industry, as well as the food industry. The total world production of fatty alcohols is over 2 million tons with approximately equal parts derived from fossil oil and from plant oils or animal fats. Due to the environmental impact of these production methods, there is an interest in alternative methods for fatty alcohol production via microbial fermentation using cheap renewable feedstocks. In this study, we aimed to obtain a better understanding of how fatty alcohol biosynthesis impacts the host organism, baker's yeast Saccharomyces cerevisiae or oleaginous yeast Yarrowia lipolytica. Producing and non-producing strains were compared in growth and nitrogen-depletion cultivation phases. The multi-omics analysis included physiological characterization, transcriptome analysis by RNAseq, 13C metabolic flux analysis, and intracellular metabolomics. Both species accumulated fatty alcohols under nitrogen-depletion conditions but not during growth. The fatty alcohol-producing Y. lipolytica strain had a higher fatty alcohol production rate than an analogous S. cerevisiae strain. Nitrogen-depletion phase was associated with lower glucose uptake rates and a decrease in the intracellular concentration of acetyl-CoA in both yeast species, as well as increased organic acid secretion rates in Y. lipolytica. Expression of the fatty-alcohol producing enzyme fatty acyl-CoA reductase alleviated the growth defect caused by deletion of hexadecenal dehydrogenase encoding genes (HFD1 and HFD4) in Y. lipolytica. RNAseq analysis showed that fatty alcohol production triggered a cell wall stress response in S. cerevisiae. RNAseq analysis also showed that both nitrogen-depletion and fatty alcohol production have substantial effects on the expression of transporter encoding genes in Y. lipolytica. In conclusion, through this multi-omics study, we uncovered some effects of fatty alcohol production on the host metabolism. This knowledge can be used as guidance for further strain improvement towards the production of fatty alcohols.

脂肪醇（fatty alcohols）广泛应用于多个行业的多元场景，涵盖肥皂与洗涤剂工业、个人护理及化妆品工业，以及食品工业。当前全球脂肪醇总产量超200万吨，其中约一半来自化石油料，另一半则源自植物油或动物油脂。鉴于此类生产方式对环境造成的影响，学界与工业界均关注利用廉价可再生原料通过微生物发酵（microbial fermentation）生产脂肪醇的替代工艺。本研究旨在深入解析脂肪醇生物合成对宿主菌株的影响，所选宿主为酿酒酵母（Saccharomyces cerevisiae）与解脂耶氏酵母（Yarrowia lipolytica）。我们对比了产脂肪醇菌株与非产菌株在生长阶段及氮耗竭培养阶段的生长情况。多组学分析涵盖生理特性表征、基于RNA测序（RNA-seq）的转录组分析、13C代谢流分析以及细胞内代谢组学分析。两种酵母均在氮耗竭条件下积累脂肪醇，而在生长阶段无此现象。解脂耶氏酵母的产醇菌株相较同源的酿酒酵母产醇菌株，展现出更高的脂肪醇生产速率。在两种酵母中，氮耗竭阶段均伴随葡萄糖摄取速率降低以及细胞内乙酰辅酶A浓度下降；解脂耶氏酵母在此阶段的有机酸分泌速率则有所提升。脂酰辅酶A还原酶（fatty acyl-CoA reductase）的表达可缓解解脂耶氏酵母中敲除十六烯醛脱氢酶编码基因（HFD1与HFD4）所导致的生长缺陷。转录组分析显示，在酿酒酵母中，脂肪醇生产会触发细胞壁应激反应；而在解脂耶氏酵母中，氮耗竭与脂肪醇生产均对转运蛋白编码基因的表达产生显著影响。综上，本项多组学研究揭示了脂肪醇生产对宿主代谢的若干影响，相关结论可为后续菌株改良以提升脂肪醇产量提供理论指导。