转录激活因子Yap1调控酿酒酵母对萜烯类物质胁迫应答分子机制研究

Terpenes are large class of aromatic compounds with great medicinal and energy values used widely in the synthesis of fragrance, pharmaceuticals, food additives and aviation fuels. There is a huge demand in the market every year at home and abroad. Microbial fermentation is a green and sustainable way to obtain the terpenoids. However, terpenoids are very toxic to the cells because of the strong lipophilic characteristic which can cause serious oxidative stress. Therefore, it’s crucial to improve the terpene stress tolerance of the microorganisms for the both increase of the production quantity and quality. It was found that in the previous study the transcription factor Yap1 with the effect of "multi-gene regulation" was involved in the oxidative stress response in the yeast cells. Therefore, this project will focus on the global regulation of terpene stress tolerance in yeast strains through the directed evolution of the transcription factor Yap1 that could make different expression patterns of multiple genes; moreover, combining the different -omics techniques including metabolomics and transcriptomics to identify the key metabolites and genes that are involved in the terpene stress responses. The gene functions and the metabolic pathways and/or signal transduction pathways will be also studied by employing the bio-information analysis. It will uncover the response mechanism of terpene stress from both the metabolic and molecular level in our study and be helpful to broaden the application scope of Saccharomyces cerevisiae as the excellent chassis cells for industrial biochemical production and to provide necessary theoretical basis for the genetic modification of yeast strains.

萜烯类物质具有重大医药和能源价值，广泛用于合成香料、药物、食品添加剂、航空燃料等，市场需求量巨大。微生物发酵获取萜类物质是一条绿色可持续的途径，但由于萜类物质极强的亲脂性，对微生物细胞具有破坏作用，引发严重的氧化胁迫。因此提高菌株对萜烯物质的耐受性对于发酵产品的产量与品质至关重要。前期研究发现，具有“多基因调控”效应的转录因子Yap1参与细胞的氧化胁迫应答反应。因此，本项目将通过对转录因子Yap1的定向进化，改变细胞内多个基因的表达模式，达到对细胞代谢活动的全局调控，提高菌株的萜烯耐受性；结合代谢组学、转录组学研究方法识别细胞参与萜烯胁迫的关键性代谢物和基因，对基因功能及其参与的代谢途径和信号转导途径进行生物信息学分析，进而从代谢和分子两个水平探究酵母细胞对萜烯胁迫的应答机制。本研究将有助于拓宽酿酒酵母作为优良底盘细胞的应用范围，对抗逆性酵母菌株的分子改良提供必要的理论依据。

萜烯类物质具有重大医药和能源价值，广泛用于合成香料、药物、食品添加剂、航空燃料等，市场需求量巨大。微生物发酵获取萜类物质是一条绿色可持续的途径，但由于萜类物质极强的亲脂性，对微生物细胞具有破坏作用，引发严重的氧化胁迫。因此提高菌株对萜烯物质的耐受性对于发酵产品的产量与品质至关重要。本项目通过对转录因子Yap1的定向进化，改变细胞内多个基因的表达模式，达到对细胞代谢活动的全局调控，提高了菌株的萜烯耐受性；同时采用驯化培养技术获得了具有稳定遗传特性的萜烯耐受性高的突变菌株；结合代谢组学、转录组学研究方法识别了细胞参与萜烯胁迫的关键性代谢物和基因，通过对关键基因进行KEGG代谢途径富集分析，发现主要的代谢途径包括：Carbohydrate transport and metabolism、Translation, ribosomal structure and biogenesis、Posttranslational modification, protein turnover、Intracellular trafficking, secretion, and vesicular transport。本研究将有助于拓宽酿酒酵母作为优良底盘细胞的应用范围，对抗逆性酵母菌株的分子改良提供必要的理论依据。