Metabolomic Variation And Genetic Basis Of Mini-core Collection For Tartary Buckwheat

Tartary buckwheat(Fagopyrum tataricum Gaertn.)is one of the cultivated of the Fagopyrum(buckwheat)genus and has been considered to originate in southwest China.Besides the well-balanced amino acid composition and rich content of dietary fiber,mineral elements and vitamins,Tartary buckwheat contains diverse bioactive compounds,such as flavonoids,anthraquinones and phenolic acids,which endues this gluten-free crop with medical values.Given the nutritional and medical values,Tartary buckwheat is often considered as an ideal functional food with diverse health benefits including pain relief,neuroprotection,anti-oxidation,anti-microbial,anti-inflammatory,anti-diabetic,anti-cancer,antihypertension and cholesterol level reduction.However,the metabolic pathways of buckwheat bioactive substances remain to be discovered and elucidated.In addition,previous studies have found that the content of flavonoids such as rutin and disease resistance are diminished during the domestication of buckwheat,but the genetic basis is not clear.We selected a mini-core collection for Tartary buckwheat,which consists of 11 wild relatives and 189 landraces based on multi-years of identification and evaluation of germplasm resources.The phylogenetic analysis divided the mini-core collection into three major monophyletic clades: group HW(Himalayan wild accessions)including 11 wild accessions and 3 landraces,group SL(Southwestern landraces)including 80 landraces,and group NL(Northern landraces)including 106 landraces.Using widelytargeted metabolomics,we constructed a metabolome map of buckwheat containing 1,092 metabolites,and 567 metabolites were annotated.These metabolites belong to flavonoids,lipids,phenolic acids,amino acids and their derivatives,and nucleotides and their derivatives,respectively.We found that the content of 245 metabolites were significantly different between HW and NL,and 265 between HW and SL,with98 metabolites showing similar trends.We conducted a subsequent mGWAS and found 1,253 signals.Among them,384 mGWAS signals corresponding to 336 metabolites were identified to be subjected to domestication.Some loci with significantly different genotype frequencies within different groups may be the genetic basis for the occurrence of directional selection for differential metabolites,including salicylic acid and emodin 8-O-glucoside.Salicylic acid is an important plant immune hormone.During crop domestication,its disease resistance is usually diminished,and the reduction in SA content may be one of the causal factors for this phenomenon.We compared the levels of SA in the buckwheat wild population HW and cultivated populations NL and SL and found that it was the highest in HW and the lowest in SL,suggesting that SA was negatively selected during domestication and affected the disease resistance of buckwheat.The mGWAS of SA identified the glycoside hydrolase gene FtSAGH1,which was able to hydrolyze SA 2-O-β-glucoside to SA in vitro.Overexpression of FtSAGH1 in buckwheat hairy roots increased SA content.Ectopic expression of FtSAGH1 in Arabidopsis increased resistance to Rhizoctonia solani AG-4 HGI.In addition,different FtSAGH1 haplotypes with different resistance to R.solani had different genotype frequencies in HW and SL.FtSAGH1 was located within the domestication sweep,indicating that FtSAGH1 controls the negative selection of SA,while reducing disease resistance during buckwheat domestication.Emodin is a bitter tasting compound widely present in Polygonaceae plants being the main functional ingredient of many herbs of the Polygonaceae.Glycosylation of emodin can increase both its solubility and bioavailability,but the catalytic process is unclear.The mGWAS of emodin 8-O-glucoside identified the UDP-glucosyltransferase gene FtUGT74L2,which was able to catalyze the glycosylation of emodin to generate emodin 8-O-glucoside in vivo and in vitro.In addition,the flavonol glycoside hydrolase gene FtFGH1 was found to be involved in the conversion between rutin,quercetin and other flavonol glycosides and quercetin.And the R2R3-MYB transcription factor FtMYB43 regulated buckwheat procyanidin synthesis by upregulating the expression of genes for procyanidin synthesis,such as FtANS,FtDFR and FtFLS.This study provides a systematic understanding of buckwheat metabolite variation and enrich the regulation mechanism of specialized metabolism of Tartary buckwheat.The uncover of genetic basis for controlling changes in buckwheat bioactive ingredient content contributes to genetic breeding and quality improvement of buckwheat.