CeBiTec Colloquium: 2008/08/20 Takayuki Tohge, RIKEN Plant Science Center Yokohama, Japan (currently MPI of Molecular Plant Physiology Golm, Germany)

2008/08/20, 17:00

CeBiTec Laboratory Building, Room G2-104

Comprehensive flavonol profiling and transcriptome coexpression analysis leading to decoding gene-metabolite correlations in Arabidopsis

Abstract:
To complete the metabolic map for an entire class of compounds, it is essential to identify gene-metabolite correlations of a metabolic pathway. The flavonoids produced by Arabidopsis thaliana wild-type and flavonoid biosynthetic mutant lines were identified using liquid chromatography-mass spectrometry (LC-MS). The structures of 15 newly identified and 8 known flavonols were deduced by LC-MS profiling of these mutants. Candidate genes presumably involved in the flavonoid pathway were delimited by transcriptome coexpression network analysis using public databases, leading to the detailed analysis of two flavonoid pathway genes, UGT89C1 (At1g06000), UGT78D3 (At5g17030),AtOMT1 (At5g54160) and RHM1 (At1g78570). No C-7 rhamnosylated flavonols were detected in either of two T-DNA ugt89c1 mutants. This specific metabolite deficiency in the mutants was complemented by stable transformation with the genomic fragment containing intact UGT89C1. Glutathione S-transferasefused recombinant UGT89C1 protein converted kaempferol 3-O-glucoside to kaempferol 3-O-glucoside-7-O-rhamnoside and recognized 3-O-glycosylated flavonols and UDP-rhamnose as substrates, but not flavonol aglycones, 3-O-glycosylated anthocyanins or other UDP-sugars. The levels of flavonol 3-O-arabinosides were reduced in ugt78d3 knockdown mutants, suggesting that UGT78D3 is a flavonol arabinosyltransferase. Recombinant UGT78D3 protein could convert quercetin to quercetin 3-O-arabinoside. The strict substrate specificity of UGT78D3 for flavonol aglycones and UDP-arabinose indicate that UGT78D3 is the first flavonol arabinosyltransferase characterized. The metabolic profiling of the omt1 mutant lacking a methyltransferase gene narrowed down by the coexpression analysis showed that AtOMT1 (At5g54160) is involved not only in the production of lignins and sinapoyl esters but also in the methylation of flavonols forming isorhamnetin. A comparison of flavonol profile in RHMs knockout mutants indicated that RHM1 play a major role in supplying UDP-rhamnose for flavonol modification. The rate of flavonol 3-O-glycosylation is more affected than those of 7-O-glycosylation by the supply of UDP-rhamnose. The precise identification of flavonoids in conjunction with transcriptomics thus led to the identification of a novel gene function and a more complete understanding of a plant metabolic network.