Jin Baolong, Cui Guanghong, Guo Juan, Tang Jinfu, Duan Lixin, Lin Huixin, Shen Ye, Chen Tong, Zhang Huabei, Huang Luqi
State Key Laboratory Breeding Base of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China (B.J., G.C., J.G., J.T., H.L., Y.S., T.C., H.Z., L.H.); and.
Key Laboratory of Plant Molecular Physiology, Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China (L.D.).
Plant Physiol. 2017 Jun;174(2):943-955. doi: 10.1104/pp.17.00202. Epub 2017 Apr 5.
-kaurene diterpenoids are the largest group of known diterpenoids. Among them, oridonin is accumulated in the leaves, and is the most frequently studied compound because of its antitumor and antibacterial activities. We have identified five copalyl diphosphate synthase (CPS) and six kaurene synthase-like (KSL) genes by transcriptome profiling of leaves. An in vitro assay assigns ten of them to five different diterpene biosynthesis pathways, except IrCPS3 that has a mutation in the catalytic motif. The Lamiaceae-specific clade genes (IrCPS1 and IrCPS2) synthesize the intermediate copalyl diphosphate (normal-CPP), while IrCPS4 and IrCPS5 synthesize the intermediate -copalyl diphosphate (-CPP). IrKSL2, IrKSL4, and IrKSL5 react with -CPP to produce an -isopimaradiene-like compound, -atiserene and -kaurene, respectively. Correspondingly, the Lamiaceae-specific clade genes IrKSL1 or IrKSL3 combined with normal-CPP led to the formation of miltiradiene. The compound then underwent aromatization and oxidization with a cytochrome P450 forming two related compounds, abietatriene and ferruginol, which were detected in the root bark. IrKSL6 reacts with normal-CPP to produce isopimaradiene. IrKSL3 and IrKSL6 have the γβα tridomain structure, as these proteins tend to possess the bidomain structure of IrKSL1, highlighting the evolutionary history of KSL gene domain loss and further elucidating chemical diversity evolution from a macroevolutionary stance in Lamiaceae.
贝壳杉烯二萜类化合物是已知二萜类化合物中最大的一类。其中，冬凌草甲素积累在叶片中，因其抗肿瘤和抗菌活性而成为研究最为频繁的化合物。我们通过对叶片进行转录组分析，鉴定出了5个柯巴基二磷酸合酶（CPS）基因和6个贝壳杉烯合酶类似物（KSL）基因。体外实验将其中10个基因分配到5条不同的二萜生物合成途径中，不过IrCPS3在催化基序上存在突变。唇形科特有的进化枝基因（IrCPS1和IrCPS2）合成中间产物柯巴基二磷酸（正常-CPP），而IrCPS4和IrCPS5合成中间产物异柯巴基二磷酸（-CPP）。IrKSL2、IrKSL4和IrKSL5分别与-CPP反应生成类似异海松二烯的化合物、-阿替斯烯和-贝壳杉烯。相应地，唇形科特有的进化枝基因IrKSL1或IrKSL3与正常-CPP结合导致形成丹参新醌二烯。该化合物随后通过细胞色素P450进行芳构化和氧化，形成两种相关化合物，枞酸三烯和铁锈醇，在根皮中被检测到。IrKSL6与正常-CPP反应生成异海松二烯。IrKSL3和IrKSL6具有γβα三结构域结构，因为这些蛋白倾向于拥有IrKSL1的双结构域结构，突出了KSL基因结构域丢失的进化历史，并从宏观进化的角度进一步阐明了唇形科化学多样性的进化。
