Transcriptomic Responses to Chilling Reveal Potential Chilling Tolerance Mechanisms in Cucumber

Chilling is a devastating stress that has led to a crisis of production for cucumber (Cucumis sativus L.). To determine the molecular mechanisms underlying chilling responses in cucumber, we investigated physiological changes and transcriptomic responses to chilling stress in the chilling-tolerant inbred line CC and chilling-susceptible inbred line R1461. Physiological analysis showed that CC had a higher survival rate, lower H₂O₂ accumulation, and ion leakage than R1461 after chilling treatment. RNA-seq analysis identified 938 differentially expressed genes (DEGs) in response to chilling and revealed that chilling stress regulated the transcript levels of genes related to hormones, including auxin, salicylic acid (SA), jasmonic acid (JA), and ethylene. RT-qPCR and pharmacological analysis suggested that cucumber chilling tolerance was associated with variation in the gene expression involved in ethylene biosynthesis and signaling. Exogenously applying 1-aminocyclopropane-1-carboxylic acid (ACC), the precursor of ethylene, improved the chilling tolerance of cucumber, while the exogenous application of the ethylene inhibitor AgNO₃ impaired the chilling tolerance of cucumber. After ACC treatment, the difference in chilling tolerance between CC and R1461 disappeared, suggesting that the different chilling tolerance level between CC and R1461 is dependent on the ethylene biosynthesis and signaling pathway. In addition, a comparison of cucumber lines with different chilling tolerances revealed that chilling tolerance is highly associated with the up-regulation of C-repeat binding factor (CBF) genes, while natural variation in the promoter of CsCBF1 is associated with chilling response. This study thus provides information on transcriptomic responses in different varieties of chilling-tolerant cucumber and reveals potential chilling tolerance mechanisms that could be used to improve chilling tolerance in cucumber.