Ethylene regulates organic acid metabolism in strawberry via the FaMAP3K-FaERF13-FaKAT module

The ripening of fleshy fruits is a highly complex process characterized by changes in quality attributes such as sugar, acid, and anthocyanin content. Among these, the levels of major organic acids (citric and malic acids) are crucial determinants of fruit taste quality. However, the mechanisms underlying the biosynthesis and metabolism of organic acids (citric and malic acids) in strawberries-especially those influenced by ethylene signaling-remain inadequately understood. This study elucidates the ethylene-mediated regulation of organic acid (citric and malic acids) metabolism in strawberry fruits through the FaMAP3K-FaERF13-FaKAT module. We identified FaERF13 as a key transcription factor that directly binds to and represses the FaKAT promoter (electrophoretic mobility shift assay, yeast 1-hybrid, luciferase reporter assay), thereby promoting the accumulation of total organic acids. Comparative analysis of red ("Benihoppe") and white ("Snow Princess") cultivars revealed that this regulatory mechanism is consistently observed, although post-translational modifications likely explain the acid reduction in ripening "Snow Princess" despite high FaERF13 expression. Protein interaction studies (yeast 2-hybrid, co-immunoprecipitation, bimolecular fluorescence complementation) demonstrated that FaMAP3K physically interacts with FaERF13, forming a signaling cascade that fine-tunes the homeostasis of organic acids (citric and malic acids). Functional validation in strawberry and tobacco systems confirmed that FaMAP3K enhances FaERF13-mediated FaKAT suppression. These findings establish a dual-layer regulatory network in which ethylene coordinates both transcriptional (FaERF13-FaKAT) and post-translational (FaMAP3K-FaERF13) control of acid metabolism during strawberry ripening.