title: Multifunctional oxidosqualene cyclases and cytochrome P450 involved in the biosynthesis of apple fruit triterpenic acids.
abstract: Apple (Malus × domestica) accumulates bioactive ursane-, oleanane-, and lupane-type triterpenes in its fruit cuticle, but their biosynthetic pathway is still poorly understood. We used a homology-based approach to identify and functionally characterize two new oxidosqualene cyclases (MdOSC4 and MdOSC5) and one cytochrome P450 (CYP716A175). The gene expression patterns of these enzymes and of previously described oxidosqualene cyclases were further studied in 20 apple cultivars with contrasting triterpene profiles. MdOSC4 encodes a multifunctional oxidosqualene cyclase producing an oleanane-type triterpene, putatively identified as germanicol, as well as β-amyrin and lupeol, in the proportion 82 : 14 : 4. MdOSC5 cyclizes 2,3-oxidosqualene into lupeol and β-amyrin at a ratio of 95 : 5. CYP716A175 catalyses the C-28 oxidation of α-amyrin, β-amyrin, lupeol and germanicol, producing ursolic acid, oleanolic acid, betulinic acid, and putatively morolic acid. The gene expression of MdOSC1 was linked to the concentrations of ursolic and oleanolic acid, whereas the expression of MdOSC5 was correlated with the concentrations of betulinic acid and its caffeate derivatives. Two new multifuntional triterpene synthases as well as a multifunctional triterpene C-28 oxidase were identified in Malus × domestica. This study also suggests that MdOSC1 and MdOSC5 are key genes in apple fruit triterpene biosynthesis.
authors.affiliation: Department of Environmental Research and Innovation, Luxembourg Institute of Science and Technology, Avenue des Hauts-Fourneaux, L-4362, Esch/Alzette, Luxembourg.
authors.forename: Christelle M
authors.identifier: 0000-0003-0239-5774
authors.initials: CM
authors.lastname: Andre
authors.affiliation: Department of Environmental Research and Innovation, Luxembourg Institute of Science and Technology, Avenue des Hauts-Fourneaux, L-4362, Esch/Alzette, Luxembourg.
authors.forename: Sylvain
authors.initials: S
authors.lastname: Legay
authors.affiliation: Department of Environmental Research and Innovation, Luxembourg Institute of Science and Technology, Avenue des Hauts-Fourneaux, L-4362, Esch/Alzette, Luxembourg.|Institut des Sciences de la Vie, UCLouvain, B-1348, Louvain-la-Neuve, Belgium.
authors.forename: Amélie
authors.initials: A
authors.lastname: Deleruelle
authors.affiliation: The New Zealand Institute for Plant & Food Research Limited, Mt Albert Research Centre, Private Bag 92 169, Auckland, 1142, New Zealand.
authors.forename: Niels
authors.initials: N
authors.lastname: Nieuwenhuizen
authors.affiliation: The New Zealand Institute for Plant & Food Research Limited, Mt Albert Research Centre, Private Bag 92 169, Auckland, 1142, New Zealand.
authors.forename: Matthew
authors.initials: M
authors.lastname: Punter
authors.affiliation: The New Zealand Institute for Plant & Food Research Limited, Mt Albert Research Centre, Private Bag 92 169, Auckland, 1142, New Zealand.
authors.forename: Cyril
authors.initials: C
authors.lastname: Brendolise
authors.affiliation: The New Zealand Institute for Plant & Food Research Limited, Ruakura, Hamilton, 3240, New Zealand.
authors.forename: Janine M
authors.initials: JM
authors.lastname: Cooney
authors.affiliation: Walloon Agricultural Research Centre, Rue de Liroux, B-5030, Gembloux, Belgium.
authors.forename: Marc
authors.initials: M
authors.lastname: Lateur
authors.affiliation: Department of Environmental Research and Innovation, Luxembourg Institute of Science and Technology, Avenue des Hauts-Fourneaux, L-4362, Esch/Alzette, Luxembourg.
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Hausman
authors.affiliation: Institut des Sciences de la Vie, UCLouvain, B-1348, Louvain-la-Neuve, Belgium.
authors.forename: Yvan
authors.initials: Y
authors.lastname: Larondelle
authors.affiliation: The New Zealand Institute for Plant & Food Research Limited, Mt Albert Research Centre, Private Bag 92 169, Auckland, 1142, New Zealand.
authors.forename: William A
authors.initials: WA
authors.lastname: Laing
mesh_terms: D000595:Amino Acid Sequence; D053898:Biosynthetic Pathways / Q000235:genetics; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D005638:Fruit / Q000201:enzymology* / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D019751:Intramolecular Transferases / Q000378:metabolism*; D027845:Malus / Q000201:enzymology* / Q000235:genetics; D010802:Phylogeny; D018515:Plant Leaves / Q000378:metabolism; D030821:Plants, Genetically Modified; D025341:Principal Component Analysis; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D013185:Squalene / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics; D014315:Triterpenes / Q000737:chemistry / Q000378:metabolism*
chemical_list: D014315:Triterpenes; C520393:oxidosqualene; D013185:Squalene; D003577:Cytochrome P-450 Enzyme System; D019751:Intramolecular Transferases; C021924:lanosterol synthase
keywords: Malus × domestica; apple; betulinic acid; cytochrome P450; germanicol; oxydosqualene cyclase; triterpene; ursolic acid