A comprehensive proteomic map revealing the regulation of the development of long-duration, red butterfly-shaped fruit in Euscaphis japonica

Fruit features are crucial for plant propagation, population growth, biodiversity preservation, and evolutionary survival. However, the synergistic regulatory mechanisms underlying the development of fruit traits such as color, shape and duration are unclear. Euscaphis japonica, whose fruits have a red-winged pericarp and persist for a long period of time, is an important ornamental plant in eastern Asia. In this study, we present a complete proteome spanning multiple time points and the phosphoproteome landscape of E. japonica fruit during the maturation and ripening phases. Quantitative evaluation via two-way proteomics analysis revealed three distinct phases that are consistent with the fruit maturation and ripening stages on a longitudinal time scale. The proteome and phosphoproteome analyses revealed functionally important biological events, including anthocyanin accumulation and phytohormone and light signal transduction. Importantly, our integrated analysis, along with experimental validation and phytohormone treatments, suggests that alterations in EjPHYBS21/S37 and EjPHOT1S394/S429 phosphorylation may lead to auxin accumulation and the inhibition of ethylene biosynthesis, thereby initiating the development of long-duration, red butterfly-shaped E. japonica fruit. Our study reveals a mechanism of E. japonica fruit formation that highlights plant adaptive strategies that potentially evolved through interactions with frugivores.