title: Down-regulation of β-carotene hydroxylase increases β-carotene and total carotenoids enhancing salt stress tolerance in transgenic cultured cells of sweetpotato.
abstract: Sweetpotato (Ipomoea batatas Lam.) is an important industrial crop and source of food that contains useful components, including antioxidants such as carotenoids. β-Carotene hydroxylase (CHY-β) is a key regulatory enzyme in the beta-beta-branch of carotenoid biosynthesis and it catalyzes hydroxylation into both β-carotene to β-cryptoxanthin and β-cryptoxanthin to zeaxanthin. To increase the β-carotene content of sweetpotato through the inhibition of further hydroxylation of β-carotene, the effects of silencing CHY-β in the carotenoid biosynthetic pathway were evaluated. A partial cDNA encoding CHY-β was cloned from the storage roots of orange-fleshed sweetpotato (cv. Shinhwangmi) to generate an RNA interference-IbCHY-β construct. This construct was introduced into cultured cells of white-fleshed sweetpotato (cv. Yulmi). Reverse transcription-polymerase chain reaction analysis confirmed the successful suppression of IbCHY-β gene expression in transgenic cultured cells. The expression level of phytoene synthase and lycopene β-cyclase increased, whereas the expression of other genes showed no detectable change. Down-regulation of IbCHY-β gene expression changed the composition and levels of carotenoids between non-transgenic (NT) and transgenic cells. In transgenic line #7, the total carotenoid content reached a maximum of 117 μg/g dry weight, of which β-carotene measured 34.43 μg/g dry weight. In addition, IbCHY-β-silenced calli showed elevated β-cryptoxanthin and zeaxanthin contents as well as high transcript level P450 gene. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity (DPPH) in transgenic cells was more than twice that in NT cells. RNA-IbCHY-β calli increased abscisic acid (ABA) content, which was accompanied by enhanced tolerance to salt stress. In addition, the production of reactive oxygen species measured by 3,3'-diaminobenzidine (DAB) staining was significantly decreased in transgenic cultured cells under salt stress. Taken together, the present results indicate that down-regulation of IbCHY-β increased β-carotene contents and total carotenoids in transgenic plant cells and enhanced their antioxidant capacity.
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-806, Republic of Korea.
authors.forename: Sun Ha
authors.initials: SH
authors.lastname: Kim
authors.forename: Young Ock
authors.initials: YO
authors.lastname: Ahn
authors.forename: Mi-Jeong
authors.initials: MJ
authors.lastname: Ahn
authors.forename: Haeng-Soon
authors.initials: HS
authors.lastname: Lee
authors.forename: Sang-Soo
authors.initials: SS
authors.lastname: Kwak
mesh_terms: D000040:Abscisic Acid / Q000378:metabolism; D019883:Alkyl and Aryl Transferases / Q000378:metabolism; D000975:Antioxidants / Q000378:metabolism; D001713:Biphenyl Compounds / Q000378:metabolism; D002338:Carotenoids / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D018076:DNA, Complementary; D015536:Down-Regulation; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D017343:Genes, Plant; D051232:Geranylgeranyl-Diphosphate Geranylgeranyltransferase; D006900:Hydroxylation; D019753:Intramolecular Lyases / Q000378:metabolism; D027723:Ipomoea batatas / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D010851:Picrates / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D018517:Plant Roots; D035281:Plant Tubers / Q000201:enzymology; D030821:Plants, Genetically Modified; D034622:RNA Interference; D017382:Reactive Oxygen Species / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055049:Salt Tolerance / Q000235:genetics*; D013312:Stress, Physiological; D019207:beta Carotene / Q000378:metabolism*
chemical_list: D000975:Antioxidants; D001713:Biphenyl Compounds; D018076:DNA, Complementary; D010851:Picrates; D010940:Plant Proteins; D017382:Reactive Oxygen Species; D019207:beta Carotene; D002338:Carotenoids; D000040:Abscisic Acid; D003577:Cytochrome P-450 Enzyme System; C004931:1,1-diphenyl-2-picrylhydrazyl; D006899:Mixed Function Oxygenases; C079232:beta-carotene hydroxylase; D019883:Alkyl and Aryl Transferases; D051232:Geranylgeranyl-Diphosphate Geranylgeranyltransferase; D019753:Intramolecular Lyases; C070925:lycopene cyclase-isomerase