Enhanced triacylglycerol production in the diatom Phaeodactylum tricornutum by inactivation of a Hotdog-fold thioesterase gene using TALEN-based targeted mutagenesis

文献类型： 外文期刊

第一作者： Hao, Xiahui

作者： Hao, Xiahui;Luo, Ling;You, Lingjie;Chen, Hong;Wei, Fang;Gong, Yangmin;Hao, Xiahui;Luo, Ling;You, Lingjie;Chen, Hong;Wei, Fang;Gong, Yangmin;Jouhet, Juliette;Rebeille, Fabrice;Marechal, Eric;Hu, Hanhua;Pan, Yufang;Tan, Xiaoming;Chen, Zhuo

作者机构：

关键词： Acyl-CoA thioesterase; Acyl-CoA; Fatty acids; Phaeodactylum tricornutum; TALEN; Triacylglycerols

期刊名称：BIOTECHNOLOGY FOR BIOFUELS （ 影响因子：6.04； 五年影响因子：6.485 ）

ISSN： 1754-6834

年卷期： 2018 年 11 卷

页码：

收录情况： SCI

摘要： Background: In photosynthetic oleaginous microalgae, acyl-CoA molecules are used as substrates for the biosynthesis of membrane glycerolipids, triacylglycerol (TAG) and other acylated molecules. Acyl-CoA can also be directed to beta-oxidative catabolism. They can be utilized by a number of lipid metabolic enzymes including endogenous thioesterases, which catalyze their hydrolysis to release free fatty acids. Acyl-CoA availability thus plays fundamental roles in determining the quantity and composition of membrane lipids and storage lipids. Results: Here, we have engineered the model diatom Phaeodactylum tricornutum to produce significantly increased TAGs by disruption of the gene encoding a Hotdog-fold thioesterase involved in acyl-CoA hydrolysis (ptTES1). This plastidial thioesterase can hydrolyze both medium-and long-chain fatty acyl-CoAs, but has the highest activity toward long-chain saturated and monounsaturated fatty acyl-CoAs. The maximum rate was found with oleoyl-CoA, which is hydrolyzed at 50 nmol/min/mg protein. The stable and targeted interruption of acyl-CoA thioesterase gene was achieved using a genome editing technique, transcription activator-like effector nucleases (TALENs). Disruption of native ptTES1 gene resulted in a 1.7-fold increase in TAG content when algal strains were grown in nitrogen-replete media for 8 days, whereas the content of other lipid classes, including phosphoglycerolipids and galactoglycerolipids, remained almost unchanged. The engineered algal strain also exhibited a marked change in fatty acid profile, including a remarkable increase in 16: 0 and 16: 1 and a decrease in 20:5. Nitrogen deprivation for 72 h further increased TAG content and titer of the engineered strain, reaching 478 mu g/10(9) cells and 4.8 mg/L, respectively. Quantitative determination of in vivo acyl-CoAs showed that the total acyl-CoA pool size was significantly higher in the engineered algal strain than that in the wild type. Conclusions: This study supports the role of ptTES1 in free fatty acid homeostasis in the plastid of Phaeodactylum and demonstrates the potential of TALEN-based genome editing technique to generate an enhanced lipid-producing algal strain through blocking acyl-CoA catabolism.

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