虾青素具有多种生理功能,应用广泛。酵母红法夫酵母是微生物虾青素的理想来源。然而,有利于虾青素合成的应激条件通常会抑制细胞生长,导致该酵母中虾青素的生产率低。在这项研究中,1mg/L褪黑素(MT)可以增加生物量,虾青素含量,P.rhodozyma的产量分别为21.9、93.9和139.1%,达到6.9g/L,0.3毫克/克DCW,和2.2mg/L,分别。基于RNA-seq的转录组分析显示,MT可以干扰P.rhodozyma细胞的转录组谱。此外,差异表达基因(DEG)分析表明,MT显著诱导或抑制的基因主要参与虾青素的合成,代谢物代谢,衬底运输,抗压力,信号转导,和转录因子。提出了MT调节虾青素合成的机理。其机制是MT进入细胞与各种信号通路的组分相互作用或直接调节其转录水平。然后将改变的信号传递给转录因子,可以调控DEGs等一系列下游基因的表达。锌指转录因子基因(ZFTF),最上调的DEG之一,选择MT诱导的P.rhodozyma中过表达。发现与MT处理条件相比,转化体的生物量和虾青素合成进一步增加。结合MT治疗和ZFTF在P.rhodozyma中的过表达,生物量,虾青素含量,产量为8.6g/L,0.6mg/gDCW,和4.8mg/L,比无MT条件下的WT菌株增加了52.1、233.3和399.7%。在这项研究中,对虾青素的合成与调控理论进行了深化,提出了一种高效的微生物虾青素工业化生产的双重策略。
Astaxanthin has multiple physiological functions and is applied widely. The yeast Phaffia rhodozyma is an ideal source of microbial astaxanthin. However, the stress conditions beneficial for astaxanthin synthesis often inhibit cell growth, leading to low productivity of astaxanthin in this yeast. In this study, 1 mg/L melatonin (MT) could increase the biomass, astaxanthin content, and yield in P. rhodozyma by 21.9, 93.9, and 139.1%, reaching 6.9 g/L, 0.3 mg/g DCW, and 2.2 mg/L, respectively. An RNA-seq-based transcriptomic analysis showed that MT could disturb the transcriptomic profile of P. rhodozyma cell. Furthermore, differentially expressed gene (DEG) analysis show that the genes induced or inhibited significantly by MT were mainly involved in astaxanthin synthesis, metabolite metabolism, substrate transportation, anti-stress, signal transduction, and transcription factor. A mechanism of MT regulating astaxanthin synthesis was proposed in this study. The mechanism is that MT entering the cell interacts with components of various signaling pathways or directly regulates their transcription levels. The altered signals are then transmitted to the transcription factors, which can regulate the expressions of a series of downstream genes as the DEGs. A zinc finger transcription factor gene (ZFTF), one of the most upregulated DEGs, induced by MT was selected to be overexpressed in P. rhodozyma. It was found that the biomass and astaxanthin synthesis of the transformant were further increased compared with those in MT-treatment condition. Combining MT-treatment and ZFTF overexpression in P. rhodozyma, the biomass, astaxanthin content, and yield were 8.6 g/L, 0.6 mg/g DCW, and 4.8 mg/L and increased by 52.1, 233.3, and 399.7% than those in the WT strain under MT-free condition. In this study, the synthesis and regulation theory of astaxanthin is deepened, and an efficient dual strategy for industrial production of microbial astaxanthin is proposed.