title: Branch Pathway Blocking in Artemisia annua is a Useful Method for Obtaining High Yield Artemisinin.
abstract: There are many biosynthetic pathways competing for the metabolic flux with the artemisinin biosynthetic pathway in Artemisia annua L. To study the relationship between genes encoding enzymes at branching points and the artemisinin biosynthetic pathway, β-caryophyllene, β-farnesene and squalene were sprayed on young seedlings of A. annua. Transient expression assays indicated that the transcription levels of β-caryophyllene synthase (CPS), β-farnesene synthase (BFS) and squalene synthase (SQS) were inhibited by β-caryophyllene, β-farnesene and squalene, respectively, while expression of some artemisinin biosynthetic pathway genes increased. Thus, inhibition of these genes encoding enzymes at branching points may be helpful to improve the artemisinin content. For further study, the expression levels of four branch pathway genes CPS, BFS, germacrene A synthase (GAS) and SQS were down-regulated by the antisense method in A. annua. In anti-CPS transgenic plants, mRNA levels of BFS and ADS were increased, and the contents of β-farnesene, artemisinin and dihydroartemisinic acid (DHAA) were increased by 212, 77 and 132%, respectively. The expression levels of CPS, SQS, GAS, amorpha-4,11-diene synthase (ADS), amorphadiene 12-hydroxylase (CYP71AV1) and aldehyde dehydrogenase 1 (ALDH1) were increased in anti-BFS transgenic plants and, at the same time, the contents of artemisinin and DHAA were increased by 77% and 54%, respectively, and the content of squalene was increased by 235%. In anti-GAS transgenic plants, mRNA levels of CPS, BFS, ADS and ALDH1 were increased. The contents of artemisinin and DHAA were enhanced by 103% and 130%, respectively. In anti-SQS transgenic plants, the transcription levels of BFS, GAS, CPS, ADS, CYP71AV1 and ALDH1 were all increased. Contents of artemisinin and DHAA were enhanced by 71% and 223%, respectively, while β-farnesene was raised to 123%. The mRNA level of artemisinic aldehyde Δ11(13) reductase (DBR2) had changed little in almost all transgenic plants.
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Zongyou
authors: Z
authors: Lv
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Fangyuan
authors: F
authors: Zhang
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Qifang
authors: Q
authors: Pan
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Xueqing
authors: X
authors: Fu
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Weimin
authors: W
authors: Jiang
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Qian
authors: Q
authors: Shen
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Tingxiang
authors: T
authors: Yan
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Pu
authors: P
authors: Shi
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Xu
authors: X
authors: Lu
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China.
authors: Xiaofen
authors: X
authors: Sun
authors: Joint International Research Laboratory of Metabolic & Developmental Sciences, Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Shanghai Jiao Tong University, Shanghai 200240, China kxtang@sjtu.edu.cn.
authors: Kexuan
authors: K
authors: Tang
mesh_terms: D031172:Artemisia annua / Q000187:drug effects / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D037621:Artemisinins / Q000737:chemistry / Q000378:metabolism*; D053898:Biosynthetic Pathways / Q000187:drug effects / Q000235:genetics; D013186:Farnesyl-Diphosphate Farnesyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D017343:Genes, Plant; D007783:Lactones / Q000737:chemistry / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D000081223:Polycyclic Sesquiterpenes; D011755:Pyrophosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D012717:Sesquiterpenes / Q000494:pharmacology; D013185:Squalene / Q000494:pharmacology; D013729:Terpenes / Q000494:pharmacology
chemical_list: D037621:Artemisinins; D007783:Lactones; D010940:Plant Proteins; D000081223:Polycyclic Sesquiterpenes; D012717:Sesquiterpenes; D013729:Terpenes; C062671:beta-farnesene; D013185:Squalene; C024714:caryophyllene; D013186:Farnesyl-Diphosphate Farnesyltransferase; D011755:Pyrophosphatases; C109459:beta-farnesene synthase; C075875:artemisin
keywords: Antisense; Artemisia annua L; Artemisinin biosynthesis; Germacrene A synthase (GAS); Squalene synthase (SQS); β-Caryophyllene synthase (CPS); β-farnesene synthase (BFS)