The 14-3-3 Protein SlTFT1 Accelerates Tomato Fruit Ripening by Binding and Stabilising YFT1 in the Ethylene Signalling Pathway.

Ethylene plays a pivotal role during the fruit ripening process in tomato (Solanum lycopersicum). Previously, we have reported that the tomato EIN2-like protein YELLOW-FRUITED TOMATO1 (YFT1), a core component in the ethylene signal transduction pathway, exerts a critical regulatory function in the tomato fruit ripening process. However, the molecular mechanism of YFT1-mediated ethylene signalling during tomato ripening still remains largely unknown. In this study, we performed yeast two-hybrid screens of a cDNA expression library from tomato fruits and obtained a new YFT1-interacting protein, SlTFT1 (TOMATO FOURTEEN-THREE PROTEIN 1), and confirmed their interaction in vitro and in vivo. Further bimolecular fluorescence complementation (BiFC) assays indicated that SlTFT1 can specifically bind the canonical and non-canonical on the carboxyl terminus of YFT1 (YFT1-C). Triple response assays and ethylene responsive gene expression analysis demonstrated that SlTFT1 positively regulates the ethylene signalling pathway in a YFT1-dependent manner. Phenotypical and biochemical analysis of SlTFT1 overexpression lines and loss-of-function mutants showed that SlTFT1 significantly accelerated ethylene emission, chromoplast development, lycopene accumulation and fruit ripening rates; however, the deletion of YFT1 and/or SlTFT1 lesion generated the opposite results, suggesting that SlTFT1 positively regulates tomato fruit ripening also in a YFT1-dependent manner. Co-expression and western blotting assays showed that SlTFT1 can efficiently prevent YFT1 from protein degradation mediated by an F-box protein SlETP2-like3, suggesting that SlTFT1 can stabilise YFT1 through their physical interaction. Collectively, our results reveal a genetic and molecular framework of the SlTFT1-YFT1 complex, which modulates tomato fruit ripening by regulating ethylene signalling.