A TE-derived regulatory variation in RsOFP2.3 underlies morphological diversification of radish taproots through the conserved OFP-TRM module

Abstract Radish (Raphanus sativus L.) represents a major root vegetable crop exhibiting remarkable variation in taproot morphology, yet the underlying genetic and molecular mechanisms remain poorly understood. Through a structural variant-based genome-wide association study, we identified RsOFP2.3, encoding an OVATE Family Protein, as a major determinant of fleshy taproot shape. Comprehensive expression profiling and RNA in situ hybridization revealed that RsOFP2.3 is broadly expressed, with the highest transcript accumulation in cambial tissues. Notably, RsOFP2.3 expression was markedly higher in the round-rooted accession than in the long-rooted one. Population-wide analysis showed that RsOFP2.3 expression was negatively correlated with taproot length and shape index, but positively correlated with taproot width. A 312-bp transposable element (TE) insertion in the RsOFP2.3 promoter repressed its expression and was strongly associated with taproot shape variation, revealing a TE-mediated cis-regulatory mechanism underlying morphological divergence. Functional analyses showed that RsOFP2.3 overexpression promotes radial expansion by enhancing cambial cell division and xylem differentiation, resulting in thicker and shorter taproots, whereas silencing RsOFP2.3 produced opposite phenotypic effects. Mechanistically, RsOFP2.3 physically interacts with the TONNEAU1-recruiting motif protein RsTRM4, recruiting it from microtubules to the cytoplasm. Downregulation of RsTRM4 reduced taproot length, while RsTRM4 overexpression partially alleviated the shortened organ phenotypes caused by RsOFP2.3 overexpression, indicating an antagonistic relationship that fine-tunes organ morphology. These findings uncover two coordinated regulatory mechanisms involving TE-mediated cis-regulatory variation and a conserved OFP-TRM interaction module that jointly shape the balance between radial and longitudinal growth during radish taproot development, providing valuable molecular targets for precision breeding of storage root crops.