Unraveling the target genes of RIN transcription factor during tomato fruit ripening and softening

BACKGROUND: The RIN transcription factor is one of the MADS box family members and predominantly controls fruit ripening. In this study, effort was made to demonstrate the regulation network of RIN transcription factor during tomato fruit ripening and softening. Novel RIN target geneswere identified by proteomics, electrophoresis mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) analysis. RESULTS: Over 700 protein spots were achieved by two-dimensional gel electrophoresis, and 42 proteins were successfully identified. Among them, 1-aminocyclopropane-1-carboxylate oxidase (LeACO4, ethylene synthesis, spot 3) and alpha-galactosidase-like isoform 2 (alpha-Gal, cell wall metabolism, spot 26) exhibited varied expression levels in different tomato fruits. Particularly high expression levels of LeACO4 and alpha-Gal were observedin wild type but not in the rin mutant. Additionally, CArGbox, aRIN-binding site, was discovered in the promoter regions of both LeACO4 and alpha-Gal genes, suggesting that RIN possibly directly regulates their transcriptions, and this assumption was further confirmed by EMSA and ChIP assay. CONCLUSION: Functional annotations of RIN target genes demonstrated the specific role of RIN in the process of fruit ripening and softening, especially in cell wall degradation and ethylene biosynthesis. This study will further illuminate the mechanism of tomato ripening and softening. (C) 2016 Society of Chemical Industry