A Pair of Tabersonine 16-Hydroxylases Initiates the Synthesis of Vindoline in an Organ-Dependent Manner in Catharanthus roseus
A Pair of Tabersonine 16-Hydroxylases Initiates the Synthesis of Vindoline in an Organ-Dependent Manner in Catharanthus roseus
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DOI：
10.1104/pp.113.222828
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发表时间：
2013-12-01
期刊：
影响因子：
7.4
通讯作者：
Courdavault, Vincent
中科院分区：
文献类型：
--
作者：
Besseau, Sebastien;Kellner, Franziska;Courdavault, Vincent
在他勃松宁16-羟化酶（T16 H）的催化下，他勃松宁在C-16位的羟基化，启动了文多灵的合成，文多灵构成了在长春花叶中积累的主要生物碱。在过去的十年中，该反应与从未分化的C.玫瑰细胞在这项研究中，我们分离出第二个细胞色素P450（CYP 71 D351）显示T16 H活性。生化特性表明，CYP 71 D12和CYP 71 D351都表现出对他勃松宁的高亲和力和窄的底物特异性，使T16 H，据我们所知，第一个生物碱生物合成酶显示两个异构体编码的不同基因的日期在C。罗斯然而，这两个基因显着分歧的转录本分布在植物。虽然CYP 71 D12（T16 H1）表达仅限于花和未分化细胞，但CYP 71 D351（T16 H2）表达谱与其他文多灵生物合成基因相似，在幼叶中达到最大值。此外，转录本本地化carboreclavage和RNA原位杂交表明，CYP 71 D351信使RNA特异性地位于叶表皮，这也主机的下一步文多灵生物合成。文多灵高、低积累型C. roseus栽培品种也突出了CYP 71 D351转录物和文多灵水平之间的直接相关性。此外，由病毒诱导的基因沉默介导的CYP 71 D351下调减少了叶片中文多灵的积累，并将生物合成通量重新导向C-16位的未修饰生物碱的产生。所有这些数据表明，他勃松宁16-羟基化是由两个基因，包括CYP 71 D351，它编码的特定T16 H亚型在叶文多灵生物合成中发挥作用的器官依赖性的方式编排。
Hydroxylation of tabersonine at the C-16 position, catalyzed by tabersonine 16-hydroxylase (T16H), initiates the synthesis of vindoline that constitutes the main alkaloid accumulated in leaves of Catharanthus roseus. Over the last decade, this reaction has been associated with CYP71D12 cloned from undifferentiated C. roseus cells. In this study, we isolated a second cytochrome P450 (CYP71D351) displaying T16H activity. Biochemical characterization demonstrated that CYP71D12 and CYP71D351 both exhibit high affinity for tabersonine and narrow substrate specificity, making of T16H, to our knowledge, the first alkaloid biosynthetic enzyme displaying two isoforms encoded by distinct genes characterized to date in C. roseus. However, both genes dramatically diverge in transcript distribution in planta. While CYP71D12 (T16H1) expression is restricted to flowers and undifferentiated cells, the CYP71D351 (T16H2) expression profile is similar to the other vindoline biosynthetic genes reaching a maximum in young leaves. Moreover, transcript localization by carborundum abrasion and RNA in situ hybridization demonstrated that CYP71D351 messenger RNAs are specifically located to leaf epidermis, which also hosts the next step of vindoline biosynthesis. Comparison of high-and low-vindoline-accumulating C. roseus cultivars also highlights the direct correlation between CYP71D351 transcript and vindoline levels. In addition, CYP71D351 down-regulation mediated by virus-induced gene silencing reduces vindoline accumulation in leaves and redirects the biosynthetic flux toward the production of unmodified alkaloids at the C-16 position. All these data demonstrate that tabersonine 16-hydroxylation is orchestrated in an organ-dependent manner by two genes including CYP71D351, which encodes the specific T16H isoform acting in the foliar vindoline biosynthesis.