Institute of Soil and Water Conservation, CAS & MWR, Yangling 712100, China; University of Chinese Academy of Sciences, Beijing 100049, China.
College of Life Sciences, Key Laboratory of Plant Secondary Metabolism and Regulation of Zhejiang Province, Zhejiang Sci-Tech University, Hangzhou 310018, China.
Plant Sci. 2018 Nov;276:229-238. doi: 10.1016/j.plantsci.2018.07.016. Epub 2018 Jul 31.
The bHLH transcription factors have important role in regulation of plant growth, development, and secondary metabolism. Tanshinones are the major pharmaceutical components present in Salvia miltiorrhiza Bunge. It has been reported that bHLHs have functions in terpenoids biosynthesis. Here, we got a bHLH family member named SmbHLH10 which could positively regulate tanshinones biosynthesis in S. miltiorrhiza hairy roots. In the SmbHLH10-overexpressing line 6, four major tanshinones contents were reaching 2.51-fold (dihydrotanshinone I), 2.84-fold (cryptotanshinone), 2.89- fold (tanshinone I), 2.68-fold (tanshinone II A) of WT, respectively. The variation in tanshinones biosynthetic pathway gene transcription was generally consistent with tanshinones content. DXS2, DXS3 and DXR of MEP pathway were induced substantially, reaching 10-fold, 3-fold, 5.74-fold higher of the WT, respectively. The downstream pathway genes CPS1, CPS5 and CYP76AH1 were highest in line OE-SmbHLH10-6, reached 4.93, 16.29 and 3.27-fold of the WT, respectively, while KSL1's expression was highest in line OE-SmbHLH10-4, 4.64-fold of WT. Yeast one-hybrid assays results showed that SmbHLH10 could binds the predicted G-box motifs within the promoters of DXS2, CPS1 and CPS5. These findings indicated that SmbHLH10 could directly binds to G-box in the pathway genes' promotor, activate their expression and then upregulate tanshinones biosynthesis.
bHLH 转录因子在植物生长、发育和次生代谢的调节中起着重要作用。丹参酮是丹参中主要的药物成分。据报道，bHLH 参与萜类生物合成。在这里，我们得到了一个 bHLH 家族成员，命名为 SmbHLH10，它可以正向调节丹参毛状根中的丹参酮生物合成。在 SmbHLH10 过表达系 6 中，四种主要丹参酮的含量分别达到 WT 的 2.51 倍（二氢丹参酮 I）、2.84 倍（隐丹参酮）、2.89 倍（丹参酮 I）和 2.68 倍（丹参酮 II A）。丹参酮生物合成途径基因转录的变化与丹参酮含量的变化基本一致。MEP 途径中的 DXS2、DXS3 和 DXR 基因被显著诱导，分别达到 WT 的 10 倍、3 倍和 5.74 倍。下游途径基因 CPS1、CPS5 和 CYP76AH1 在 OE-SmbHLH10-6 中最高，分别达到 WT 的 4.93 倍、16.29 倍和 3.27 倍，而 KSL1 的表达在 OE-SmbHLH10-4 中最高，达到 WT 的 4.64 倍。酵母单杂交实验结果表明，SmbHLH10 可以结合 DXS2、CPS1 和 CPS5 启动子中的预测 G 盒基序。这些结果表明，SmbHLH10 可以直接结合途径基因启动子中的 G 盒，激活其表达，从而上调丹参酮的生物合成。
