Metabolomics analysis reveals Embden Meyerhof Parnas pathway activation and flavonoids accumulation during dormancy transition in tree peony

Abstract

Background: Bud dormancy is a sophisticated strategy which plants evolve to survive in tough environments. Endodormancy is a key obstacle for anti-season culture of tree peony, and sufficient chilling exposure is an effective method to promote dormancy release in perennial plants including tree peony. However, the mechanism of dormancy release is still poorly understood, and there are few systematic studies on the metabolomics during chilling induced dormancy transition.

Results: The tree peony buds were treated with artificial chilling, and the metabolmics analysis was employed at five time points after 0-4 °C treatment for 0, 7, 14, 21 and 28 d, respectively. A total of 535 metabolites were obtained and devided into 11 groups including flavonoids, amino acid and its derivatives, lipids, organic acids and its derivates, nucleotide and its derivates, alkaloids, hydroxycinnamoyl derivatives, carbohydrates and alcohols, phytohormones, coumarins and vitamins. Totally, 118 differential metabolites (VIP ≥ 1, P < 0.05) during chilling treatment process were detected, and their KEGG pathways involved in several metabolic pathways related to dormancy. Sucrose was the most abundant carbohydrate in peony bud. Starch was degradation and Embden Meyerhof Parnas (EMP) activity were increased during the dormancy release process, according to the variations of sugar contents, related enzyme activities and key genes expression. Flavonoids synthesis and accumulation were also promoted by prolonged chilling. Moreover, the variations of phytohormones (salicylic acid, jasmonic acid, abscisic acid, and indole-3-acetic acid) indicated they played different roles in dormancy transition.

Conclusion: Our study suggested that starch degradation, EMP activation, and flavonoids accumulation were crucial and associated with bud dormancy transition in tree peony.

Keywords: Dormancy transition; EMP activation; Flavonoids accumulation; Metabolomics; Tree peony.

Fig. 1

Metabolomics analysis of tree peony buds. a Bud morphology of tree peony buds during the chilling duration. The scale bar indicated 1 cm. b PCA plot of the tree peony buds at five stages. 0 d, 7 d, 14 d, 21 d and 28 d indicated days for chilling duration. QC, quality control samples. c The KEGG pathway annotation of metabolites. Black texts indicated the first hierarchy of KEGG Pathway database, and colored texts indicated the second hierarchy of KEGG Pathway database. The global and overview is a secondary metabolism name, containing eight metabolic pathways: Metabolic pathways, Biosynthesis of secondary metabolites, Microbial metabolism in diverse environments, Carbon metabolism, 2-Oxocarboxylic acid metabolism, Fatty acid metabolism, Biosynthesis of amino acids, Degradation of aromatic compounds

Fig. 2

Expression dynamics and comparative analysis of metabolites in tree peony buds chilled at different stages. a Statistical map of intergroup metabolites (P < 0.05, VIP ≥ 1). Red and blue represented up- and down-regulated metabolites, respectively. b Venn diagram of metabolites among four comparison groups. c The KEGG pathway annotation of all DMs

Fig. 3

Changes of 50 DMs related to dormancy release in tree peony. a The clustering heatmap of 50 DMs at 14 d vs 7 d and 21 d vs 7 d. b The top 20 of KEGG enrichment pathways for DMs among 14 d vs 7 d and 21 d vs 7 d

Fig. 4

Dynamics of carbon metabolic pathways throughout the chilling duration process. The metabolite amounts were shown in heatmaps as 0, 7, 14, 21, and 28 d from left to right, respectively. G6P (glucose 6-phosphate), F6P (fructose 6-phosphate), GAP (3-phosphoglyceraldehyde), PEP (phosphoenolpyruvate)

Fig. 5

Changes of carbohydrates during dormancy transition induced by the chilling in tree peony. a The cluster heatmap of carbohydrates. b The variations of several sugars content. c The relative expression levels of sucrose synthase and sucrose invertase genes. d The variations of starch and amylases activity. The mean ± SD in three biological replicates was shown. *, ** and *** indicated significant differences of one-way ANOVA at P < 0.05, P < 0.01, and P < 0.001, respectively

Fig. 6

The changes of flavonoids content and the related genes expression during the chilling duration. a The flavonoids and anthocyanin pathway. CHS, chalcone synthase; CHI, chalcone isomerase; F3H, flavanone 3-hydroxylase; DFR, dihydroflavonol 4-reductase; ANS, anthocyanin synthase; 3GT, anthocyanidin 3-O-glycosyltransferase; 5GT, anthocyanidin 5-O-glycosyltransferase; MT, anthocyanidin methyltransferase; 7GT, anthocyanidin 7-O-glycosyltransferase; FLS, flavonol synthase; FNS, flavone synthase; Lu, luteolin; Ap, apigenin; Is, isorhamnetin; Qu, quercetin; Km, kaempferol; Cy, cyanidin; Pg, pelargonidin; Glu, glucoside; Hex, hexoside; Neo, neohesperidoside; Rut, rutinoside; Rob, robinobioside; Syr, syringic acid. b The relative content of anthocyanins in tree peony flower buds after 28 d chilling duration based on the results of LC-ESI-MS/MS. c The relative expression levels of key genes involving in anthocyanin biosynthesis. Data were represented as mean of three different determinations ± SD. Asterisks indicate statistically significant differences (one-way ANOVA, *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001)

Fig. 7

The changes of phytohormone contents and the expression patterns of related genes during chilling duration. a The cluster heatmap of phytohormone during the chilling duration. b Phytohormone contents in tree peony flower buds during the chilling duration by LC-MS/MS. c The qRT-PCR results of JA related gene PsMYC2, and two auxin receptors genes PsAFBs, and PsTIR1. The mean ± SD (n = 3) were shown. Asterisks indicated statistically significant differences (one-way ANOVA, *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001)

Fig. 8

Model of metabolite role during dormancy transition induced by chilling in tree peony. Starch degradation and EMP activation provide energy and material basis for flavonoid accumulation during endodormancy release. Flavonoid and anthocyanin accumulation might promote flower bud development at ecodormancy stage. Meanwhile, the accumulation of anthoyanin may be regulated by JA