Comprehensive identification of ripening-related RNA-binding proteins in tomatoes using improved plant phase extraction.

RNA-binding proteins (RBPs) have emerged as key players in posttranscriptional gene regulation, yet their full scale role in fruit ripening remains to be fully elucidated. However, due to the complex structure and composition of fruit tissue, exploring RBPs in fruits still faces many challenges. Here, we optimized the plant phase extraction method and successfully applied it to tomato fruits for the unbiased excavation of RBPs in fruits, this method were named as "plant phase extraction in tomato fruit" (termed tfPPE). We yielded a comprehensive candidate RNA-binding proteome (RBPome) composed of 230 proteins and disclosed that approximately 66% of them were unconventional RBPs. Validation of the RNA-binding activities of six candidate RBPs unveiled that metabolic enzymes function as moonlighting RBPs. Furthermore, combined with transcriptome analysis, we identified 41 candidate RBPs associated with fruit ripening. Remarkably, we proposed that SlER21 and SlFER1 play significant roles in fruit coloring and ripening process. Taken together, these results demonstrate that tfPPE was an impactful approach for unbiased excavation RBPs in fruits and pave the way for investigating RBP functions in fruit-ripening regulatory network.