异戊烯基焦磷酸转运对雷公藤甲素生物合成的影响

Effect of isopentenyl pyrophosphate translocation on the biosynthesis of triptolide

摘要:

雷公藤甲素是一种具有显著抗炎、抗肿瘤和免疫抑制活性的天然产物，市场需求量大，临床应用前景广阔。文中以雷公藤悬浮细胞为实验材料，通过对不同培养时期 (7 d、14 d) 的细胞外源性添加d,l-甘油醛 (DLG)以阻断异戊烯基焦磷酸 (IPP) 转运，分析诱导前后的细胞活性及生物量、雷公藤甲素累积量及其生物合成上游途径关键酶基因的变化规律，探究IPP转运在雷公藤甲素生物合成过程中的时空特点及其影响机制。通过实验结果可知：IPP转运参与雷公藤甲素的生物合成；在细胞培养早期，IPP转运主要经由质体 (MEP途径) 转运到细胞质 (MVA途径) 中，而细胞培养晚期则相反；阻断IPP转运，可反馈调节雷公藤甲素生物合成上游途径关键酶基因的表达，进而影响雷公藤甲素的累积。上述研究填补了雷公藤甲素生物合成过程中IPP转运特点和机制方面的空白，为雷公藤甲素合成生物学关键技术的开发奠定了理论基础，具有重要的现实意义。

Abstract:

Triptolide has wide clinical applications due to its anti-inflammatory, anti-tumor and immunosuppressive activities. In this study, we investigated the effect of blocking isopentenyl pyrophosphate (IPP) translocation on the biosynthesis of triptolide by exogenously adding d,l-glyceraldehyde (DLG) to the suspension cells of Ttripterygium wilfordii at different stages (7 d, 14 d). Subsequently, the cell viability, biomass accumulation, triptolide contents, as well as the profiles of the key enzyme genes involved in the upstream pathway of triptolide biosynthesis, were analyzed. The results showed that IPP translocation is involved in the biosynthesis of triptolide. IPP is mainly translocated from the plastid (containing the MEP pathway) to the cytoplasm (containing the MVA pathway) in the early stage of the culture, but reversed in the late stage. Blocking the translocation of IPP affected the expression of key enzyme genes involved in the upstream pathway of triptolide, which in turn affected the accumulation of triptolide. Understanding the characteristics and mechanism of IPP translocation provides a theoretical basis for further promoting triptolide biosynthesis through synthetic biology.