Key message

MIR396b had been cloned and overexpressed in Salvia miltiorrhiza hairy roots. MiR396b targets SmGRFs, SmHDT1, and SmMYB37/4 to regulate cell growth and secondary metabolism in S. miltiorrhiza hairy roots.

Abstract

Danshen (Salvia miltiorrhiza Bunge) is a valuable medicinal herb with two kinds of clinically used natural products, salvianolic acids and tanshinones. miR396 is a conserved microRNA and plays extensive roles in plants. However, it is still unclear how miR396 works in S. miltiorrhiza. In this study, an smi-MIR396b has been cloned from S. miltiorrhiza. Overexpression of miR396b in danshen hairy roots inhibited hairy root growth, reduced salvianolic acid concentration, but enhanced tanshinone accumulation, resulting in the biomass and total salvianolic acids respectively reduced to 55.5 and 72.1% of the control and total tanshinones increased up to 1.91-fold of the control. Applied degradome sequencing, 5′RLM-RACE, and qRT-PCR, 13 targets for miR396b were identified including seven conserved SmGRF1-7 and six novel ones. Comparative transcriptomics and microRNomics analysis together with qRT-PCR results confirmed that miR396b targets SmGRFs, SmHDT1, and SmMYB37/4 to mediate the phytohormone, especially gibberellin signaling pathways and consequentially resulted in the phenotype variation of miR396b-OE hairy roots. Furthermore, miR396b could be activated by methyl jasmonate, abscisic acid, gibberellin, salt, and drought stresses. The findings in this study indicated that smi-miR396b acts as an upstream and central regulator in cell growth and the biosynthesis of tanshinones and salvianolic acids, shedding light on the coordinated regulation of plant growth and biosynthesis of active ingredients in S. miltiorrhiza.

中文翻译：

---

smi-miR396b 靶向 SmGRFs、SmHDT1 和 SmMYB37/4 协同调节丹参毛状根中的细胞生长和活性成分积累。

关键信息

MIR396b 已被克隆并在 丹参 毛状根中过表达。MiR396b 靶向 SmGRFs、SmHDT1 和 SmMYB37/4 ，以调节 丹参毛根中的细胞生长和次生代谢。

抽象的

丹参（Salvia miltiorrhiza Bunge）是一种名贵药材，有两种临床常用的天然产物丹酚酸和丹参酮。miR396 是一种保守的 microRNA，在植物中发挥着广泛的作用。然而，目前尚不清楚 miR396 如何在S. miltiorrhiza中发挥作用。在这项研究中，从丹参中克隆了一个 smi-MIR396b. 丹参毛状根中miR396b的过表达抑制毛状根生长，降低丹酚酸浓度，但增加丹参酮的积累，导致生物量和丹酚酸总量分别降低至对照的55.5％和72.1％，总丹参酮增加至对照的1.91倍控制。应用降解组测序、5'RLM-RACE 和 qRT-PCR，鉴定了 13 个 miR396b 靶标，包括 7 个保守SmGRF1-7和 6 个新靶标。比较转录组学和 microRNomics 分析以及 qRT-PCR 结果证实 miR396b 靶向SmGRFs、SmHDT1和SmMYB37/4介导植物激素，尤其是赤霉素信号通路，从而导致 miR396b-OE 毛状根的表型变异。此外，茉莉酸甲酯、脱落酸、赤霉素、盐和干旱胁迫可以激活 miR396b。本研究结果表明，smi-miR396b 在细胞生长和丹参酮和丹酚酸的生物合成中充当上游和中枢调节剂，揭示了丹参中植物生长和活性成分生物合成的协调调节。