| Plant secondary metabolites play an important role in plant growth, development, biotic and abiotic stress response. Derived from phenylalanine metabolism, flavonoids constitute a major group of plant phenolic compounds and involved in plant physiological processes, such as root growth, flower pollination, pathogens and UV stress. Flavonoids are strong antioxidants, which therefor has also been widely used for human disease treatment and health food. Phenolamides, however, is a large secondary metabolites derived from arginine and phenylalanine metabolism. It has been reported to have crucial roles in a number of physiological processes, such as seed germination, pollen fertility, and drought, cold, UV and pathogens stresses. Investigating accumulation of flavonoids and phenolamides in rice(Oryza sativa), one of mankind’s most important food crops, contributes to illustrate rice growth processes, stress response and nutritional quality traits.While extensively studied in Arabidopsis, profiling and naturally occurring variation of these compounds in rice, the monocot model plant, are less reported. Using a collection of rice germplasm, comprehensive profiling and natural variation of flavonoids were presented in this report. Application of a widely targeted metabolomics method facilitated the simultaneous identification and quantification of 91 flavonoids using liquid chromatography tandem mass spectrometry(LC-MS/MS), including 72 flavones, 9 flavanones, 6 flavonols and 4 anthocyaninis. Comparing flavonoid contents in various tissues during different developmental stages revealed tissue-specific accumulation of most flavonoids. Flag leaf contained the highest levels of most flavonoids, followed by culm, panicle and grain. Root, however, showed the lowest accumulation of most flavonoids. Comparing the levels of flavonoids with different glycosylated and acylated forms in rice subspecies, we found that that flavone mono-C-glycosides, malonylated flavonoid O-hexosides and some flavonoid O-glycosides accumulated at significant higher levels in indica than in japonica, while the opposite was observed for flavonoid 7 O-glycosides and aromatic acylated flavone C-hexosyl-O-hexosides. Studies of rice flavonoid biosynthetic pathway have enabled the identification of a few genes responsible for production of flavonoids. Besides, an association analysis between flavonoid accumulation and its biosynthetic gene sequence polymorphisms disclosed natural variation of flavonoids was probably caused by sequence polymorphisms in coding region of flavonoid biosynthetic genes.A total of 16 phenolamides, including different hydrocinnmoyl derivatives of agmatine, putrescine and spermidine, have also been characterized in rice using both LC-MS based widely targeted and targeted metabolomics method. Comprehensive metabolic profiling and natural variation analysis of phenolamides in a collection of rice germplasm were also presented. Spatio-temporal controlled accumulations were observed for most phenolamides. Root contained the highest levels of most phenolamides, followed by flag leaf and panicle. Differential accumulations between the two major subspecies of rice were also observed. spermidine conjugates displayed very high levels in japonica, while very low levels in indica. Coumaroylated agmatine and putrescine displayed higher levels in indica than those in japonica. Further metabolic genome-wide association study(m GWAS) in leaf leaf identified a few loci might be responsible for the natural variation of spermidine conjugates. Further in vivo metabolic analysis of the transgenic plants identified Os12g27220 and Os12g27254 as two spermidine hydroxycinnamoyl transferases that might be underlying the natural variation of levels of spermidine conjugates in rice. Our work demonstrates ‘gene-to-metabolite’ analysis by m GWAS provides a useful tool for functional gene identification and omics-based crop genetic improvement. |
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