SlNAP2promoted fruit ripening by directly binding theACS2promoter and interacting withEIL3

Abstract

AbstractThe ripening process of tomato fruit is affected by a variety of environmental factors and genetic regulators. NAC transcription factors (TFs) function in a multitude of biological processes, while the current knowledge on the participation of NAC TFs in the regulatory network of fruit ripening is relatively limited. In this study, we isolated a NAC TFs, NAP2, which acts as a positive transcription activator in tomato fruit ripening. We also observed a notable delay in the ripening process ofSlNAP2silenced and knockout mutant fruit. In particular, ethylene production was obviously inhibited inSlNAP2mutant fruit. Y1H and DLR assays showed that SlNAP2 directly binds to the promoter ofSlACS2and activates its transcriptional activity. Furthermore, SlNAP2 and SlEIL3 physically interaction was demonstrated by yeast two-hybrid (Y2H), luciferase complementation (LUC), bimolecular fluorescence complementation (BiFC) and coimmunoprecipitation analysis (CoIP) assays. Meanwhile, the pigment content, firmness and the transcript levels of genes associated with carotenoid and chlorophyll metabolism and cell wall metabolism were also potentially affected by theSlNAP2deletion; however, it has remained unclear whether these genes are also directly regulated bySlNAP2. Therefore, our findings indicate that SlNAP2 directly binds toSlACS2promoter to activate its expression and promote ethylene generation, which in turn interacts with EIL3 to enhance the function of ethylene in tomato fruit ripening. Collectively, our data contribute to understanding the interaction of NAC TFs and ethylene in tomato fruit ripening, thereby enhancing our knowledge of the ripening regulatory network that governs tomato fruit maturation.