自养和兼养条件下三角褐指藻岩藻黄素生物合成过程及其调控机制比较分析

Fucoxanthin, one of the carotenoids, has potential health promoting effects on anti-obesity, anti-cancer, as well as anti-diabetic. Phaeodactylum tricornutum is an ideal species for fucoxanthin production due to its rapid growth rate and high fucoxanthin content. However, the complete biosynthesis route of fucoxanthin is yet unclear, which limited the development of controllable fucoxanthin production. Our previous studies showed that the growth rate of P. tricornutum was enhanced with an increase of fucoxanthin content under high concentration of CO2 autotrophic cultivation, while the growth rate of P. tricornutum was enhanced but fucoxanthin content decreased under glucose-mixotrophic condition. Therefore, this project will focus on elucidating the regulation mechanism between carbon metabolism and fucoxanthin biosynthesis. On the one hand, we attempt to clarify the mechanism of intracellular carbon flux distribution and reveal the major resource of carbon skeleton for fucoxanthin biosynthesis via metabolomics analysis. On the other, we try to screen the key gene(s) that regulate central carbon metabolism and fucoxanthin biosynthesis in autotrophic and mixotrophic P. tricornutum according to proteomics and transcriptome analysis, overexpressing this specific gene, and then verify the function and the regulation mechanism involved in fucoxanthin synthesis of this gene. Further, the regulation mechanisms of fucoxanthin biosynthesis in autotrophic and mixotrophic P. tricornutum will be comprehensively revealed. The completion of this project will provide a new theoretical basis for the study of fucoxanthin biosynthesis, and lay the theoretical foundation for the achievement of controlled production of fucoxanthin in P. tricornutum.

岩藻黄素是一种具有减肥、抗癌、降血糖等生物学活性的类胡萝卜素，市场潜力巨大。三角褐指藻生长速度快、岩藻黄素含量高，是生产岩藻黄素的理想物种，但其岩藻黄素合成路径尚不完全明确，阻碍了岩藻黄素可控化生产的发展。我们前期研究发现，高CO2自养下三角褐指藻生长速度与岩藻黄素含量同步提高，而在葡萄糖兼养下生长速度加快、岩藻黄素含量显著降低。为此，本项目将从碳代谢与岩藻黄素生物合成之间的联系入手，一方面通过代谢组学研究，阐明自养与兼养下细胞内碳流的分配机制，揭示岩藻黄素合成过程中碳骨架的主要来源；一方面进行蛋白质组及转录组学分析，挖掘自养和兼养下调控碳代谢和岩藻黄素合成的关键基因，构建基因过表达藻株，验证基因的功能及其调控岩藻黄素合成的机制；从而全面解析两种模式下三角褐指藻岩藻黄素生物合成的调控机制。本项目的完成将为岩藻黄素生物合成研究提供新的理论依据，同时为实现岩藻黄素人工可控化生产奠定理论基础。

岩藻黄素是一种具有重要应用价值的天然类胡萝卜素，具有减肥、抗肿瘤、降血糖、抗氧化等一系列食用及医药用价值，市场前景广阔。硅藻，是海洋中重要的初级生产力，光合效率高、生长速度快，同时兼具富含油脂及岩藻黄素的特点，是目前工业化生产岩藻黄素的理想原材料。然而，微藻岩藻黄素生物合成路径尚的不完全明确，为人工可控化生产微藻岩藻黄素带来了极大的困难。本项目通过追踪岩藻黄素生物合成过程中碳骨架的来源与分配，发现了高碳自养条件下三角褐指藻通过蛋白质及可溶性糖降解为岩藻黄素及油脂的生物合成提供碳骨架，葡萄糖兼养条件下三角褐指藻摄取葡萄糖后可能直接利用葡萄糖进入糖类（尤其是可储存性糖）合成而非进入糖酵解过程或磷酸戊糖途径，揭示了自养和兼养培养下三角褐指藻胞内碳流在生长、岩藻黄素和油脂三者之间的分配机制；基于稳定同位素标记技术与组学分析技术，筛选到了可调控三角褐指藻生长、岩藻黄素及油脂生物合成的关键基因，构建了三角褐指藻基因枪遗传转化体系及关键调控基因过表达与沉默型转基因藻株，反向验证了调控基因在三角褐指藻岩藻黄素生物合成过程中的功能。本项目成果为解析岩藻黄素的生物合成过程与调控机制提供了新思路，也为微藻岩藻黄素产业化分子育种提供了依据，具有重要的应用研究前景。