Crosstalk between hormones and secondary metabolites regulates the interactions between plants and stress. However, little is known about the effects of hormone crosstalk on the concentration of flavonoids in seeds. In this study, we identified abscisic acid (ABA) as a negative regulator of flavonoid accumulation in soybean seeds under drought-stress conditions. Alterations in flavonoid accumulation at several intensities of water stress, followed by a recovery period, were measured during the soybean seed-filling stage. Low soil moisture (SM 10%) significantly decreased the total flavonoid content in seeds. The decline in flavonoid content was proportional to the severity of drought stress and was dependent on the activities of phenylalanine ammonia-lyase (PAL) and chalcone synthase (CHS), two key phenylpropanoid pathway enzymes. The expression of phenylalanine ammonia-lyase 1 (GmPAL1), chalcone isomerase 1A (GmCHI1A), and chalcone synthase 8 (GmCHS8) was associated with phenolic and flavonoid accumulation in soybean seeds of plants subjected to drought stress. Interestingly, the expression levels of GmCHS8 were highly correlated with flavonoid levels under drought stress and water recovery conditions. Cinnamic acid, which is a biosynthesis precursor shared by both phenylpropanoid metabolism and salicylic acid (SA) biosynthesis, decreased under drought stress conditions. Notably, exogenous ABA suppressed the expression of GmPAL1, which encodes the first rate-limiting enzyme in the phenylpropanoid biosynthesis pathway and affects downstream products such as SA and flavonoids. In conclusion, drought stress altered the phenylpropanoid-derived compounds, at least with regard to flavonoid and SA accumulation in seeds, which was regulated by antagonistic interactions with ABA.
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