title: Molecular cloning and functional characterization of the lycopene ε-cyclase gene via virus-induced gene silencing and its expression pattern in Nicotiana tabacum.
abstract: Lycopene ε-cyclase (ε-LCY) is a key enzyme that catalyzes the synthesis of α-branch carotenoids through the cyclization of lycopene. Two cDNA molecules encoding ε-LCY (designated Ntε-LCY1 and Ntε-LCY2) were cloned from Nicotiana tabacum. Ntε-LCY1 and Ntε-LCY2 are encoded by two distinct genes with different evolutionary origins, one originating from the tobacco progenitor, Nicotiana sylvestris, and the other originating from Nicotiana tomentosiformis. The two coding regions are 97% identical at the nucleotide level and 95% identical at the amino acid level. Transcripts of Ntε-LCY were detectable in both vegetative and reproductive organs, with a relatively higher level of expression in leaves than in other tissues. Subcellular localization experiments using an Ntε-LCY1-GFP fusion protein demonstrated that mature Ntε-LCY1 protein is localized within the chloroplast in Bright Yellow 2 suspension cells. Under low-temperature and low-irradiation stress, Ntε-LCY transcript levels substantially increased relative to control plants. Tobacco rattle virus (TRV)-mediated silencing of ε-LCY in Nicotiana benthamiana resulted in an increase of β-branch carotenoids and a reduction in the levels of α-branch carotenoids. Meanwhile, transcripts of related genes in the carotenoid biosynthetic pathway observably increased, with the exception of β-OHase in the TRV-ε-lcy line. Suppression of ε-LCY expression was also found to alleviate photoinhibition of Potosystem II in virus-induced gene silencing (VIGS) plants under low-temperature and low-irradiation stress. Our results provide insight into the regulatory role of ε-LCY in plant carotenoid biosynthesis and suggest a role for ε-LCY in positively modulating low temperature stress responses.
authors.affiliation: Department of Chemistry, Zhengzhou University, Zhengzhou 450001, China. symzgh@163.com.
authors.forename: Yanmei
authors.initials: Y
authors.lastname: Shi
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. wangranljj2010@163.com.
authors.forename: Ran
authors.initials: R
authors.lastname: Wang
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. zhiwubingdu@sohu.com.
authors.forename: Zhaopeng
authors.initials: Z
authors.lastname: Luo
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. jin_lf@126.com.
authors.forename: Lifeng
authors.initials: L
authors.lastname: Jin
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. spesge@163.com.
authors.forename: Pingping
authors.initials: P
authors.lastname: Liu
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. nanzhichan@gmail.com.
authors.forename: Qiansi
authors.initials: Q
authors.lastname: Chen
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. ibi.zefeng@gmail.com.
authors.forename: Zefeng
authors.initials: Z
authors.lastname: Li
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. likite2002@163.com.
authors.forename: Feng
authors.initials: F
authors.lastname: Li
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. ztrisam@126.com.
authors.forename: Chunyang
authors.initials: C
authors.lastname: Wei
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. mingzhuwus@126.com.
authors.forename: Mingzhu
authors.initials: M
authors.lastname: Wu
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. weipan83@126.com.
authors.forename: Pan
authors.initials: P
authors.lastname: Wei
authors.affiliation: Molecular Breeding Group, Yunnan Academy of Tobacco Agricultural Sciences, Kunming 650031, China. xieh@tobacco.gov.cn.
authors.forename: He
authors.initials: H
authors.lastname: Xie
authors.affiliation: Department of Chemistry, Zhengzhou University, Zhengzhou 450001, China. qulingbo@zzu.edu.cn.
authors.forename: Lingbo
authors.initials: L
authors.lastname: Qu
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. fuchenglin@zju.edu.cn.
authors.forename: Fucheng
authors.initials: F
authors.lastname: Lin
authors.affiliation: National Tobacco Gene Research Center, Zhengzhou Tobacco Research Institute, Zhengzhou 450001, China. yangjun@ztri.com.cn.
authors.forename: Jun
authors.initials: J
authors.lastname: Yang
mesh_terms: D002338:Carotenoids / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods*; D020868:Gene Silencing / Q000502:physiology*; D019753:Intramolecular Lyases / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D014780:Viruses / Q000235:genetics
chemical_list: D002338:Carotenoids; D019753:Intramolecular Lyases; C070925:lycopene cyclase-isomerase