茉莉酸诱导丹酚酸B合成的COI1-JAZ-MYB75信号转导单元及偶联机制研究

Jasmonates (JA) play a number of diverse roles in plant defense and development. CORONATINE INSENSITIVE1 (COI1) is a key regulator in the JA signaling pathway.Jasmonate ZIM-domain (JAZ) proteins are critical transcriptional repressors in the JA-induced plant responses. As JA co-recepter, COI1 and JAZ have recently been found to play important roles in the JA perception and JA-mediated transcriptional regulation. Our previous studies demonstrated that the expression of transcription factor MYB75 was significantly increased and the salvianolic acid B biosynthesis was also strongly induced in Salvia miltiorrhiza under the treatment of JA. However, the critical components of JA signaling pathway and their coupling mechanism are not clear. We speculated that a COI1-JAZ-MYB75 signaling module may mediate JA-induced salvianolic acid B biosynthesis. In this proposal, the phenotype, gene expression and salvianolic acid B contents of wildtype, COI1/JAZ-RNAi and JAZ overexpression transgenic plants with or without JA will be compared, to define the key function of COI1 and JAZ in JA-induced salvianolic acid B biosynthesis. Furthermore, for elucidating the coupling mechanism of COI1-JAZ-MYB75 signaling module in S. miltiorrhiza, the interactions that between COI1 and JAZ, or JAZ and MYB75, will be investigated using yeast two-hybrid system, GST-pull down combined with firefly luciferase complementation imaging assays. Success of the proposal is of great significance for the further study on the JA signaling pathway of medicinal plant secondary metabolism.

COI1是茉莉酸(JA)信号转导通路活化的开关，JAZ是一类转录抑制因子。作为JA的共受体，COI1和JAZ蛋白通过协同调控不同转录因子的活性实现JA功能。我们发现JA通过激活MYB75转录因子诱导丹参重要活性成分丹酚酸B的合成，但该信号转导通路的核心环节及其偶联的分子机制尚不清楚。我们推测丹参中存在COI1-JAZ-MYB75信号转导单元并介导了JA诱导的丹酚酸B积累。本项目在已获得COI1和JAZ沉默及过表达转基因株系的基础上，通过比较转基因与对照植株响应JA的表型、基因表达及丹酚酸B含量的变化，明确COI1和JAZ在JA诱导丹酚酸B合成信号转导中的关键作用；采用酵母双杂交、GST-pull down结合荧光素酶互补技术分析JAZ与COI1、MYB75的相互作用，阐明丹参COI1-JAZ-MYB75信号转导单元偶联的分子机制，为JA调控植物次生代谢的信号转导及应答机制提供了理论依据。

COI1（coronatine insensitive 1）蛋白是茉莉酸(JA)信号转导通路活化的开关，JAZ（Jasmonate ZIM domain）蛋白是一类转录抑制因子。作为JA的共受体，COI1和JAZ蛋白通过协同调控不同转录因子的活性而实现JA复杂的生理功能。本研究通过比较丹参COI1和JAZ基因降低或超量表达的转基因丹参植株响应JA处理的表型、基因表达差异和次生代谢产物的合成变化，明确了COI1和JAZ在JA诱导丹酚酸合成信号转导通路中的关键作用；通过转录组测序，结合荧光素酶互补和酵母双杂交实验，发现了与JAZ有相互作用的可能的候选转录因子MYC及下游调控因子LBD，进一步采用基因工程技术，在证明MYC和LBD参与了丹参茉莉酸信号转导和丹酚酸合成代谢通路的基础上，深入研究其在茉莉酸诱导丹参酚酸类成分积累中的具体功能，揭示了完整的[JA-COI1-JAZ-MYC2-LBD-丹酚酸合成（JA应答）]信号转导通路。本研究对揭示JA多样性生理功能内在机制具有重要意义，也为受JA调控的不同植物次生代谢的信号转导及应答机制提供了新的理论依据。本项目共发表基金标注SCI论文8篇，培养博士2名，硕士生4名，青年学科骨干1名。