Ji Pingping, Lin Maoyi, Chen Mengying, Kashif Muhammad Haneef, Fan Yuling, Ali Tahir, Dai Ruixian, Peng Chongsheng, Wang Zhiqing, Liu Zhong
School of Pharmacy, Shanghai Jiao Tong University, Shanghai 200240, China.
School of Pharmacy, Shanghai Jiao Tong University, Shanghai 200240, China; School of Life Sciences, Beijing University of Chinese Medicine, Beijing 102488, China.
Plant Physiol Biochem. 2023 Aug;201:107855. doi: 10.1016/j.plaphy.2023.107855. Epub 2023 Jul 5.
Asarum sieboldii Miq. possesses remarkable medicinal value due to its essential oil enriched with phenylpropenes (e.g., methyleugenol and safrole). Although the biosynthesis of phenylpropenes shares a common pathway with lignin, the regulation mechanisms in carbon flux allocation between them are unclear. This study is the first to genetically verify the carbon flux regulation mechanism in A. sieboldii roots. We regulated the expression of Caffeoyl-coenzyme A O-methyltransferase (CCoAOMT), an essential enzyme in the common pathway, to investigate carbon flux allocation in vegetative organs. Here, the lignin and phenylpropene content fluctuation was analyzed by wet chemistry and GC-MS methods. A bona fide CCoAOMT gene from A. sieboldii was firstly cloned and verified. Preliminary heterologous expression validation in transgenic Arabidopsis thaliana showed that RNAi-induced CCoAOMT down-regulation significantly decreased lignin content by 24% and increased the S/G ratio by 30%; however, AsCCoAOMT over-expression in A. thaliana resulted in a 40% increase in lignin content and a 20% decrease in the S/G ratio when compared to the wild type. Similar trends were noted in homologous transformation in A. sieboldii, although the variations were not conspicuous. Nevertheless, the transgenic A. sieboldii plants displayed substantial differences in the level of phenylpropene compounds methyleugenol and safrole leading to a 168% increase in the methyleugenol/safrole ratio in the over-expression line and a 73% reduction in RNAi-suppression line. These findings suggest that the biosynthesis of phenylpropene constituents methyleugenol and safrole seems to be prioritized over lignin. Furthermore, this study indicated that suppression of AsCCoAOMT resulted in marked root susceptibility to pathogenic fungal disease, implying a significant additional role of CCoAOMT in protecting plant vegetative parts from diseases. Overall, the present study provides important references and suggests that future research should be aimed at elucidating the detailed mechanisms of the carbon flux allocation between phenylpropenes and lignin biosynthesis, as well as the disease resistance competency.
细辛因其富含苯丙烯类化合物（如甲基丁香酚和黄樟素）的精油而具有显著的药用价值。尽管苯丙烯类化合物的生物合成与木质素共享一条共同途径，但它们之间碳通量分配的调控机制尚不清楚。本研究首次从基因层面验证了细辛根中的碳通量调控机制。我们调节了共同途径中的关键酶咖啡酰辅酶A O-甲基转移酶（CCoAOMT）的表达，以研究营养器官中的碳通量分配。在此，通过湿化学和GC-MS方法分析了木质素和苯丙烯类化合物含量的波动情况。首先克隆并验证了来自细辛的一个真正的CCoAOMT基因。在转基因拟南芥中的初步异源表达验证表明，RNA干扰诱导的CCoAOMT下调使木质素含量显著降低了2４%，S/G比值增加了30%；然而，与野生型相比，拟南芥中细辛CCoAOMT的过表达导致木质素含量增加了40%，S/G比值降低了20%。在细辛的同源转化中也观察到了类似的趋势，尽管变化不明显。然而，转基因细辛植株在苯丙烯类化合物甲基丁香酚和黄樟素的水平上表现出显著差异，导致过表达系中甲基丁香酚/黄樟素比值增加了168%，RNA干扰抑制系中降低了73%。这些发现表明，甲基丁香酚和黄樟素等苯丙烯类成分的生物合成似乎优先于木质素。此外，本研究表明，抑制细辛CCoAOMT会导致根部对病原真菌病害的易感性显著增加，这意味着CCoAOMT在保护植物营养器官免受病害方面具有重要的额外作用。总体而言，本研究提供了重要参考，并表明未来的研究应旨在阐明苯丙烯类化合物与木质素生物合成之间碳通量分配的详细机制以及抗病能力。
