Low temperature is involved in regulating plant growth, development, and quality formation. The mechanism by which low temperature affects sucrose accumulation in oriental melon fruit is currently unclear. Here, high-sucrose (HS) melons were used as the research materials and subjected to two temperature regimes of 30℃/18℃ (day/night) and 22℃/10℃ (day/night) at the onset of ethylene production. Low temperature significantly inhibited ethylene release and sucrose accumulation in melon fruit, while ethephon treatment at low temperature partially restored the ethylene production and sucrose content. Using yeast one-hybrid (Y1H), GUS activity analysis, and luciferase assay, we found that the transcription factor CmCBF3 could bind to CmACO1 (ACC oxidase 1) promoter and inhibit its activity, thereby suppressing ethylene production. Overexpression of CmCBF3 under low temperature significantly inhibited the synthesis of ethylene and sucrose. Further research showed that low temperature promoted CmERFV-2 expression, and CmERFV-2 could bind to CmCBF3 promoter to further inhibit ethylene synthesis. In addition, CmMYB44, a transcription factor that negatively regulated fruit ethylene production and sucrose accumulation, repressed the expression of CmACO1 and CmSPS1 (sucrose phosphate synthase 1). CmERFV-2 further affected the expression of CmACO1 and CmSPS1 by binding to CmMYB44 promoter, thereby regulating ethylene and sucrose content at low temperature. In summary, this study revealed the mechanism by which CmERFV-2 affects ethylene release and sucrose accumulation in oriental melon fruit, laying a foundation for breeding high-quality melon varieties suitable for low-temperature cultivation.

The flesh color of oriental melons is an important commercial trait that affects consumer preferences. To explore the mechanisms underlying the flesh color formation and regulation during fruit ripening, carotenoid-targeted metabolomic and RNA-seq analysis were conducted between white-fleshed (WF) and orange-fleshed (OF) oriental melon cultivars at different stages. The carotenoid-targeted metabolomic analysis indicated that β-carotene was the major metabolite that caused differences in flesh color between the two cultivars. Additionally, through KEGG pathway enrichment and weighted gene co-expression network (WGCNA) analysis, metabolic pathways and related transcription factors that are associated with carotenoid metabolism were selected and transcriptome data was verified using RT-qPCR. Finally, the yeast one hybrid and luciferase activity showed that the transcription factor CmWRKY49 could directly bind to the CmPSY1 promoter to activate its expression in the ‘OF’ cultivar. Transient overexpression of CmWRKY49 in ‘OF’ cultivar increased the β-carotene content, while the β-carotene content decreased when it was silenced in the same cultivar. This study provides insights into the underlying regulatory network of carotenoid metabolism in oriental melon fruit.