title: CYP93G2 is a flavanone 2-hydroxylase required for C-glycosylflavone biosynthesis in rice.
abstract: C-Glycosylflavones are ubiquitous in the plant kingdom, and many of them have beneficial effects on human health. They are a special group of flavonoid glycosides in which the sugars are C-linked to the flavone skeleton. It has been long presumed that C-glycosylflavones have a different biosynthetic origin from O-glycosylflavonoids. In rice (Oryza sativa), a C-glucosyltransferase (OsCGT) that accepts 2-hydroxyflavanone substrates and a dehydratase activity that selectively converts C-glucosyl-2-hydroxyflavanones to 6C-glucosylflavones were recently described. In this study, we provide in vitro and in planta evidence that the rice P450 CYP93G2 protein encoded by Os06g01250 is a functional flavanone 2-hydroxylase. CYP93G2 is related to the CYP93B subfamily, which consists of dicot flavone synthase II enzymes. In the presence of NADPH, recombinant CYP93G2 converts naringenin and eriodictyol to the corresponding 2-hydroxyflavanones. In addition, CYP93G2 generates 2-hydroxyflavanones, which are modified by O-glycosylation in transgenic Arabidopsis (Arabidopsis thaliana). Coexpression of CYP93G2 and OsCGT in Arabidopsis resulted in the production of C-glucosyl-2-hydroxyflavanones in the dibenzoylmethane tautomeric form. The same structure was reported previously for the in vitro OsCGT reaction products. Thus, CYP93G2 generates 2-hydroxyflavanone substrates from flavanones for C-glucosylation by OsCGT in planta. Furthermore, knocking down Os06g01250 in rice (O. sativa subsp. japonica 'Zhonghua 11') preferentially depleted the accumulation of C-glycosylapigenin, C-glycosylluteolin, and C-glycosylchrysoeriol but did not affect the levels of tricin, which is frequently present as O-glycosides in cereals. Taken together, our work conclusively assigned CYP93G2 as the first enzyme that channels flavanones to C-glycosylflavone biosynthesis in rice.
authors.affiliation: School of Biological Sciences, The University of Hong Kong, Hong Kong, China.
authors.forename: Yegang
authors.initials: Y
authors.lastname: Du
authors.forename: Hung
authors.initials: H
authors.lastname: Chu
authors.forename: Ivan K
authors.initials: IK
authors.lastname: Chu
authors.forename: Clive
authors.initials: C
authors.lastname: Lo
mesh_terms: D017360:Arabidopsis / Q000235:genetics; D002853:Chromatography, Liquid; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics; D047309:Flavones / Q000096:biosynthesis* / Q000737:chemistry; D006027:Glycosides / Q000096:biosynthesis* / Q000737:chemistry; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000235:genetics; D012275:Oryza / Q000201:enzymology* / Q000235:genetics; D010802:Phylogeny; D018515:Plant Leaves / Q000378:metabolism; D030821:Plants, Genetically Modified; D011994:Recombinant Proteins / Q000378:metabolism; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D045868:Sorghum / Q000201:enzymology; D013379:Substrate Specificity
chemical_list: D004269:DNA, Bacterial; D047309:Flavones; D006027:Glycosides; D011994:Recombinant Proteins; C036483:T-DNA; D003577:Cytochrome P-450 Enzyme System; C114032:cytochrome P-450 CYP93B1 (Glycyrrhiza echinata)