青蒿2-C-甲基-D-赤藓醇-4-磷酸胱氨酰转移酶基因克隆与分析  

Molecular cloning and characterization of the 2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase gene from Artemisia annua L.

作　　者：张曼[1,2] 向礼恩[1,2] 王辉[1,2] 兰小中[3] 陈敏[4] 廖志华[1,2] 

机构地区：[1]西南大学生命科学学院,重庆400715 [2]西南大学-西藏农牧学院药用植物联合研发中心,重庆400715 [3]西藏大学农牧学院药用植物研究中心,西藏林芝860000 [4]西南大学药学院,重庆400715

出　　处：《药学学报》2016年第8期1334-1339,共6页Acta Pharmaceutica Sinica

基　　金：国家863计划项目(2011AA100605);教育部新世纪优秀人才计划(NCET-12-0930);西南大学中央高校基本科研业务费专项(XDJK2013A024)

摘　　要：MEP途径定位于植物细胞的质体中,为包括青蒿素在内的萜类合成提供基本前体。2-C-甲基-D-赤藓醇-4-磷酸胱氨酰转移酶(2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase,MCT)是该途径的第3个关键酶,催化2-C-甲基-D-赤藓醇-4-磷酸生成4-(5'-焦磷酸胞苷)-2-C-甲基-D-赤藓醇。本文首次克隆了青蒿MCT基因全长cDNA(AaMCT)并进行了相关生物信息学分析。基因表达结果表明:AaMCT在青蒿分泌型腺体中大量表达,在叶、花、茎和根中少量表达;同时发现,AaMCT表达受到MeJA强烈诱导。亚细胞定位结果显示:AaMCT融合GFP特异性定位在叶绿体中,与MEP途径定位于质体吻合。最后在拟南芥中超量表达AaMCT,拟南芥中叶绿素a、叶绿素b和类胡萝卜素含量得到显著提高,表明AaMCT在萜类物质的生物合成中起重要作用。The plastidial methylerythritol phosphate（MEP） pathway provides 5-carbon precursors to the biosynthesis of isoprenoid（including artemisinin）. 2-C-Methyl-D-erythritol-4-phosphate cytidylyltransferase（MCT） is the third enzyme of the MEP pathway, which catalyzes 2-C-methyl-D-erythritol-4-phosphate to form 4-（cytidine 5＇-diphospho）-2-C-methyl-D-erythritol. The full-length MCT cDNA sequence（AaMCT） was cloned and characterized for the first time from Artemisia annua L. Analysis of tissue expression pattern revealed that AaMCT was highly expressed in glandular secretory trichome and poorly expressed in leaf, flower, root and stem. AaMCT was found to be a methyl jasmonate（Me JA）-induced genes, the expression of AaMCT was significantly increased after MeJA treatment. Subcellular localization indicated that the GFP protein fused with AaMCT was targeted specifically in chloroplasts. The transgenic plants of Arabidopsis thaliana with AaMCT overexpression exhibited a significantly increase in the content of chlorophyll a, chlorophyll b and carotenoids, demonstrating that AaMCT kinase plays an influential role in isoprenoid biosynthesis.

关 键 词：青蒿 2-C-甲基-D-赤藓醇-4-磷酸胱氨酰转移酶 超量表达 亚细胞定位 萜类 

分 类 号：R931[医药卫生—生药学]