AaEIL2 and AaERF059 are involved in fruit coloration and ripening by crossly regulating ethylene and auxin signal pathway in Actinidia arguta

In kiwifruit (Actinidia arguta), fruit coloration is typically accompanied by ripening; however, the intrinsic connection between these two processes remains unclear. In this study, we found that ethylene and auxin accelerated and suppressed fruit coloration and ripening, respectively, in A. arguta. ETHYLENE INSENSITIVE 3-LIKE 2 (AaEIL2) and ETHYLENE RESPONSIVE FACTOR (AaERF059) were mined and identified using yeast two-hybrid library screening and transcriptome analysis. AaEIL2, specifically induced to high expression by ethylene, was confirmed as a transcription factor that positively regulates coloration and ripening by targeting AaLDOX (leucoanthocyanidin dioxygenase involved in anthocyanin biosynthesis) and AaPG18 (polygalacturonase involved in cell wall degradation) based on its subcellular localization in Arabidopsis protoplasts, stable genetic transformation in transgenic tomato, and yeast one-hybrid and luciferase activity assays. AaERF059 also responds to ethylene and regulates ethylene-/auxin-mediated fruit coloration and ripening by targeting the downstream genes AaACS2 (ACC synthase, which is involved in ethylene biosynthesis) and AaGH3 (Gretchen-Hagen 3, which is involved in the auxin pathway). Overall, AaEIL2 and AaERF059 regulate ethylene-and auxin-mediated fruit coloration and ripening by maintaining a dynamic balance in a positive and negative regulatory manner. Our results not only identified key genes but also established an intrinsic connection between fruit coloration and ripening in A. arguta.