红花黄酮生物合成途径糖基转移酶基因CtUGT25的功能研究  

Functional study of glycosyltransferase genes CtUGT25 in the flavone biosynthesis pathway of Carthamus tinctorius L.

作　　者：齐书艺 王璐暖 何贝轩 高越 郭美丽 QI Shu-yi;WANG Lu-nuan;HE Bei-xuan;GAO Yue;GUO Mei-li(Department of Pharmacognosy,College of Pharmacy,Naval Medical University,Shanghai 200082,China;Changhai Clinical Research Unit,the First Affiliated Hospital of Naval Medical University,Shanghai 200082,China)

机构地区：[1]海军军医大学药学系生药学教研室,上海200082 [2]海军军医大学第一附属医院临床研究中心,上海200082

出　　处：《药学学报》2024年第6期1854-1863,共10页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金项目(81973421);国家重点研发计划(2019YFC1711100).

摘　　要：糖基转移酶(UDP-glycosyltransferase,UGT)是黄酮糖苷类化合物形成的末端修饰酶。本研究通过对红花花冠转录组数据库中糖基转移酶基因在四个花期的表达量与红花代谢组数据中主要成分的含量进行皮尔斯相关性分析,获得了2个与红花黄酮类化合物合成密切相关的糖基转移酶基因CtUGT25、CtUGT18,并对二者的基因及蛋白序列进行生物信息学分析。表达模式分析发现CtUGT25主要在花冠中表达,在开花第3天表达量最高;CtUGT18主要在根中表达,在开花第1天表达量最高。通过农杆菌介导的花粉管道法转基因手段在红花体内进行了功能验证,证明CtUGT25促进了山柰酚-3-O-葡萄糖苷及羟基红花黄色素A(HSYA)的积累,CtUGT18促进了山柰酚-3-O-葡萄糖苷、荭草苷的积累,二者可能是红花黄酮类化合物合成的糖基修饰酶。同时,体外实验证明CtUGT25蛋白对柚皮素、槲皮素、芹菜素、山柰酚、木犀草素、2-羟基柚皮素、高良姜素均具有催化活性。本研究为下一步通过分子生物技术调控红花的品质,特别是红花中特有成分HSYA等的工业化生产提供参考,也为其他植物的相关基因的研究提供资料。UDP glycosyltransferase(UGT)is a terminal modifying enzyme for the formation of flavonoid glycosides.In this study,we obtained two glycosyltransferase genes,CtUGT25 and CtUGT18,which are closely related to the synthesis of safflower flavonoids,through the Pierce correlation analysis of the expression of glycosyltransferase genes in the safflower corolla transcriptome database at different developmental stages with the contents of the major constituents of safflower metabolome database,and bioinformatically analyzed the gene and protein sequences of the two genes.Expression pattern analysis revealed that CtUGT25 was mainly expressed in the corolla,with the highest expression on day 3 of flowering stage;CtUGT18 was mainly expressed in the root,with the highest expression on day 1 of flowering stage.Functional validation was verified in safflower by Agrobacterium-mediated pollen-tube pathway transgenesis method,demonstrating that CtUGT25 promoted the accumulation of kaempferol-3-O-glucoside and hydroxysafflor yellow A(HSYA),and CtUGT18 promoted the accumulation of kaempferol-3-O-glucoside and orientin,both of which may be the glycosyl-modifying enzymes for the synthesis of safflower flavonoids.Meanwhile,in vitro experiments demonstrated the catalytic activity of CtUGT25 protein on naringenin,quercetin,apigenin,kaempferol,luteoin,2-hydroxynaringenin and galangin.This study serves as reference for future advancements in regulating the quality of safflower using molecular biotechnology,particularly focuses on the industrial production of safflower exclusive component HSYA.Additionally,it offers valuable insights for researching related genes in other plants.

关 键 词：红花 黄酮类化合物 糖基转移酶 生物合成途径 羟基红花黄色素A 

分 类 号：R931[医药卫生—生药学]