Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109-1048, USA.
Plant Physiol. 2010 Sep;154(1):301-10. doi: 10.1104/pp.110.159566. Epub 2010 Jul 1.
The resin of Cistus creticus subsp. creticus, a plant native to Crete, is rich in labdane-type diterpenes with significant antimicrobial and cytotoxic activities. The full-length cDNA of a putative diterpene synthase was isolated from a C. creticus trichome cDNA library. The deduced amino acid sequence of this protein is highly similar (59%-70% identical) to type B diterpene synthases from other angiosperm species that catalyze a protonation-initiated cyclization. The affinity-purified recombinant Escherichia coli-expressed protein used geranylgeranyl diphosphate as substrate and catalyzed the formation of copal-8-ol diphosphate. This diterpene synthase, therefore, was named CcCLS (for C. creticus copal-8-ol diphosphate synthase). Copal-8-ol diphosphate is likely to be an intermediate in the biosynthesis of the oxygen-containing labdane-type diterpenes that are abundant in the resin of this plant. RNA gel-blot analysis revealed that CcCLS is preferentially expressed in the trichomes, with higher transcript levels found in glands on young leaves than on fully expanded leaves, while CcCLS transcript levels increased after mechanical wounding. Chemical analyses revealed that labdane-type diterpene production followed a similar pattern, with higher concentrations in trichomes of young leaves and increased accumulation upon wounding.
乳香树 creticus 亚种 creticus 的树脂富含具有显著抗菌和细胞毒性活性的角鲨烯型二萜。从乳香树的毛状体 cDNA 文库中分离出一种假定的二萜合酶的全长 cDNA。该蛋白的推导氨基酸序列与其他被子植物物种中的 B 型二萜合酶高度相似（59%-70%相同），这些酶催化质子化引发的环化。亲和纯化的重组大肠埃希菌表达的蛋白以香叶基香叶基二磷酸为底物，并催化考伯醇-8-二醇二磷酸的形成。因此，这种二萜合酶被命名为 CcCLS（乳香树考伯醇-8-二醇二磷酸合酶）。考伯醇-8-二醇二磷酸可能是该植物树脂中丰富的含氧角鲨烯型二萜生物合成的中间体。RNA 凝胶印迹分析显示，CcCLS 优先在毛状体中表达，在幼叶的腺毛中表达水平较高，而在完全展开的叶片中表达水平较低，而 CcCLS 转录本水平在机械损伤后增加。化学分析表明，角鲨烯型二萜的产生遵循类似的模式，幼叶毛状体中的浓度较高，损伤后积累增加。
