MYC2 Transcription Factors TwMYC2a and TwMYC2b Negatively Regulate Triptolide Biosynthesis in Tripterygium wilfordii Hairy Roots.
MYC2 Transcription Factors TwMYC2a and TwMYC2b Negatively Regulate Triptolide Biosynthesis in Tripterygium wilfordii Hairy Roots.
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DOI：
10.3390/plants10040679
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发表时间：
2021-04-01
期刊：
影响因子：
--
通讯作者：
Zhu C
中科院分区：
文献类型：
--
作者：
Huo Y;Zhang J;Zhang B;Chen L;Zhang X;Zhu C
雷公藤内酯醇是从雷公藤属植物中提取的一种重要的生物活性二萜类化合物，具有多种药理活性。MYC 2转录因子（TF）在植物体内多种次生代谢产物的调控中起着重要作用。然而，MYC 2 TF是否能调节雷公藤内酯醇在T. Wilfordii仍然是未知的。本研究从T. Wilfordii毛状根和功能特征。系统发育树和亚细胞定位分析表明，它们与其他功能性MYC 2蛋白一起被归为bHLH超家族的IIIe分支，并定位于细胞核中。此外，酵母单杂交和GUS反式激活试验表明，TwMYC2a和TwMYC2b通过与启动子中的E-box（CACACATG）和T/G-box（CACGTT）基序结合，抑制了米替雷烯合酶基因TwTPS27a和TwTPS27b的启动子活性。转基因结果显示，RNA干扰TwMYC 2a/B基因可显著提高雷公藤甲素在毛状根和液体培养基中的积累，其作用机制是通过上调几个关键生物合成基因的表达，包括TwMS（TwTPS27 a/B）、TwCPS（TwTPS7/9）、TwDXR和TwHMGR 1。总之，我们的研究结果表明，TwMYC2a和TwMYC2b作为两个负调节雷公藤内酯醇的生物合成在T.为雷公藤内酯醇代谢工程的研究提供了新的思路。
Triptolide, an important bioactive diterpenoid extracted from the plant Tripterygium wilfordii, exhibits many pharmacological activities. MYC2 transcription factor (TF) plays an important role in the regulation of various secondary metabolites in plants. However, whether MYC2 TF could regulate the biosynthesis of triptolide in T. wilfordii is still unknown. In this study, two homologous MYC2 TF genes, TwMYC2a and TwMYC2b, were isolated from T. wilfordii hairy roots and functionally characterized. The analyses of the phylogenetic tree and subcellular localization showed that they were grouped into the IIIe clade of the bHLH superfamily with other functional MYC2 proteins and localized in the nucleus. Furthermore, yeast one-hybrid and GUS transactivation assays suggested that TwMYC2a and TwMYC2b inhibited the promoter activity of the miltiradiene synthase genes, TwTPS27a and TwTPS27b, by binding to the E-box (CACATG) and T/G-box (CACGTT) motifs in their promoters. Transgenic results revealed that RNA interference of TwMYC2a/b significantly enhanced the triptolide accumulation in hairy roots and liquid medium by upregulating the expression of several key biosynthetic genes, including TwMS (TwTPS27a/b), TwCPS (TwTPS7/9), TwDXR, and TwHMGR1. In summary, our findings show that TwMYC2a and TwMYC2b act as two negative regulators of triptolide biosynthesis in T. wilfordii hairy roots and also provide new insights on metabolic engineering of triptolide in the future.
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