The interaction of CpEBF1 with CpMADSs is involved in cell wall degradation during papaya fruit ripening

Ethylene plays a pivotal role in climacteric fruit ripening; whereas 1-MCP, a non-toxic antagonist of ethylene, prevents ethylene-dependent responses and fruit ripening. In this study, a short-term treatment (1 h) with 400 nL L−1 1-MCP delayed the ripening of harvested papaya. However, long-term application of 1-MCP (400 nL L−1, 16 h) resulted in abnormal fruit ripening, with the fruits exhibiting normal yellowing without softening, significantly higher cellulose and lignin contents, and intact cell walls (CW). Furthermore, we found that long-term treatment with 1-MCP significantly inhibited the expression of CpEBF1, an EIN3-binding F-box-1 gene. A protein interaction analysis using yeast two-hybrid, BiFC and GST pull-down assays showed that CpEBF1 interacts with the CpMADS1/3 and CpEIL1 proteins. The interaction of CpEBF1 with CpMADS1/3 further activated the activities of CW-degradation gene promoters. Subcellular localization showed that these proteins were localized in the nucleus. Additionally, the expression levels of CpMADS1/3, CpEIL1, and several CW-degradation-related genes were significantly downregulated by long-term 1-MCP treatment. Therefore, we propose that the inhibited expression of CpEBF1 and CpMADS1/3 resulted in the repressed activation of CW-degradation-related genes via their interaction, thereby resulting in fruit softening disorders.