Hansen Nikolaj L, Heskes Allison M, Hamberger Britta, Olsen Carl E, Hallström Björn M, Andersen-Ranberg Johan, Hamberger Björn
Plant Biochemistry Laboratory, Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, Copenhagen, DK-1871, Denmark.
Center for Synthetic Biology 'bioSYNergy' and Copenhagen Plant Sciences Centre, Copenhagen, Denmark.
Plant J. 2017 Feb;89(3):429-441. doi: 10.1111/tpj.13410. Epub 2017 Feb 14.
Tripterygium wilfordii (Celastraceae) is a medicinal plant with anti-inflammatory and immunosuppressive properties. Identification of a vast array of unusual sesquiterpenoids, diterpenoids and triterpenoids in T. wilfordii has spurred investigations of their pharmacological properties. The tri-epoxide lactone triptolide was the first of many diterpenoids identified, attracting interest due to the spectrum of bioactivities. To probe the genetic underpinning of diterpenoid diversity, an expansion of the class II diterpene synthase (diTPS) family was recently identified in a leaf transcriptome. Following detection of triptolide and simple diterpene scaffolds in the root, we sequenced and mined the root transcriptome. This allowed identification of the root-specific complement of TPSs and an expansion in the class I diTPS family. Functional characterization of the class II diTPSs established their activities in the formation of four C-20 diphosphate intermediates, precursors of both generalized and specialized metabolism and a novel scaffold for Celastraceae. Functional pairs of the class I and II enzymes resulted in formation of three scaffolds, accounting for some of the terpenoid diversity found in T. wilfordii. The absence of activity-forming abietane-type diterpenes encouraged further testing of TPSs outside the canonical class I diTPS family. TwTPS27, close relative of mono-TPSs, was found to couple with TwTPS9, converting normal-copalyl diphosphate to miltiradiene. The phylogenetic distance to established diTPSs indicates neo-functionalization of TwTPS27 into a diTPS, a function not previously observed in the TPS-b subfamily. This example of evolutionary convergence expands the functionality of TPSs in the TPS-b family and may contribute miltiradiene to the diterpenoids of T. wilfordii.
雷公藤（卫矛科）是一种具有抗炎和免疫抑制特性的药用植物。在雷公藤中鉴定出大量不同寻常的倍半萜、二萜和三萜，这激发了对其药理特性的研究。三环氧内酯雷公藤内酯醇是最早鉴定出的众多二萜之一，因其生物活性谱而备受关注。为了探究二萜多样性的遗传基础，最近在叶片转录组中发现了II类二萜合酶（diTPS）家族的扩张。在根部检测到雷公藤内酯醇和简单二萜支架后，我们对根部转录组进行了测序和挖掘。这使得能够鉴定出TPS的根特异性互补序列，并发现I类diTPS家族的扩张。对II类diTPS的功能表征确定了它们在形成四种C-20二磷酸中间体中的活性，这些中间体是一般代谢和特殊代谢的前体，也是卫矛科的一种新型支架。I类和II类酶的功能对导致形成三种支架，这解释了雷公藤中发现的一些萜类多样性。缺乏形成枞酸型二萜的活性促使对经典I类diTPS家族之外的TPS进行进一步测试。发现单TPS的近亲TwTPS27与TwTPS9偶联，将正常的柯巴基二磷酸转化为丹参二烯。与已确定的diTPS的系统发育距离表明TwTPS27新功能化为diTPS，这是TPS-b亚家族中以前未观察到的功能。这种进化趋同的例子扩展了TPS-b家族中TPS的功能，并可能为雷公藤的二萜类化合物贡献丹参二烯。
