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    • Figure 1. 

      Effect of MT on ABA content, activity and relative mRNA expression of CsNCED in cucumber seedlings under normal conditions (25 °C /18 °C). Cucumber seedlings grown in a solar greenhouse were foliar sprayed with 75 μM MT, 100 μM p-chlorophenylalanine (p-CPA, an inhibitor of melatonin synthesis) and deionized water at the two-leaf stage, and samples from the second leaves (from the bottom) were collected at 0, 3, 6, 9, 12, 18, and 24 h for content, activity and relative mRNA expression. All values shown are mean ± SD (n = 3). Different letters indicate significant differences among treatments (p < 0.05).

    • Figure 2. 

      Effects of ABA on MT content and activities and RNA expression of key MT biosynthesis enzymes in cucumber seedlings under normal conditions (25 °C /18 °C). (a, b) TDC and ASMT activities, respectively; (c, d) TDC and ASMT mRNA expression levels; (e) MT content. Cucumber seedlings grown in a solar greenhouse were foliar sprayed with 100 μM ABA, 3 mM Na2WO4 (an inhibitor of ABA synthesis) and deionized water at the two-leaf stage, and samples from the second leaves (from the bottom) were collected at 0, 3, 6, 9, 12, 18, and 24 h for content, activity and relative mRNA expression. All values shown are mean ± SD (n = 3). Different letters indicate significant differences among treatments (p < 0.05).

    • Figure 3. 

      Effects of MT and ABA on chilling tolerance of cucumber seedlings under low temperature stress. (a) phenotype; (b) daily increasing leaf area; (c) daily dry weight; (d) MDA content and (e) electrolyte leakage of cucumber seedlings under chilling stress. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 48 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are mean ± SD (n = 3). Different letters indicate significant differences among treatments (p < 0.05).

    • Figure 4. 

      Effects of different treatments on ROS content in cucumber seedlings under chilling stress. (a) H2O2 content; (b) O2·− content; (c, d) H2O2 and O2·− inverted fuorescence microscope imaging. Bars = 1000 μm. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O After 24 h, seedlings were exposed to 8 °C /5 °C for 48 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are mean ± SD (n = 3). Different letters indicate significant differences among treatments (p < 0.05).

    • Figure 5. 

      Interactive effects of MT and ABA on activities and relative mRNA expression of antioxidant enzymes in cucumber seedlings under chilling stress. (a–c) Activities of superoxide dismutase (SOD), peroxidase (POD), ascorbate and peroxidase (APX), respectively. (d–f) Relative mRNA expression of SOD, POD, APX, respectively. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 48 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are mean ± SD (n = 3). Different letters indicate significant differences among treatments (p < 0.05).

    • Figure 6. 

      Effects of different treatments on the contents of antioxidant substances in cucumber seedlings under chilling stress. (a) AsA content, (b) GSH content, (c) the ratio of AsA and DHA, (d) the ratio of GSH and GSSG. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 48 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are mean ± SD (n = 3). Different letters indicate significant differences among treatments (p < 0.05).

    • Figure 7. 

      Interaction between MT and ABA on the chlorophyll content of cucumber seedlings exposed to chilling stress. (a) Content of chlorophyll a; (b) Content of chlorophyll b; (c) Content of carotenoids. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 24 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are mean ± SD (n = 3). Different letters indicate significant differences among treatments (p < 0.05).

    • Figure 8. 

      Interaction between MT and ABA on gas exchange parameters in cucumber seedlings under chilling stress. (a) photosynthetic rate; (b) intercellular CO2 concentration; (c) stomatal conductance; and (d) transpiration rate. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 24 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are means ± SD (n = 3). Different letters indicate that the mean values are significantly different among the samples (p < 0.05).

    • Figure 9. 

      Interactive effects of MT and ABA on the relative expression of rbcL and RCA in mRNA, and the activity of Rubisco and Rubisco activase (RCA) and protein levels in cucumber seedlings under chilling stress. (a, b) Relative mRNA expression for rbcL and RCA, (c) Rubisco activity, (d) RCA activity and (e, f) protein levels for RbcL and RCA. Rubisco (RBC) protein is shown as an equal loading control by Coomassie Brilliant Blue staining. The value above each line is the relative enrichment of the protein. Western blotting was performed three times with three independent biological samples and similar results were obtained. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 24 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are means ± SD (n = 3). Different letters indicate that the mean values are significantly different among the samples (p < 0.05).

    • Figure 10. 

      Interactive effects of MT and ABA on photoprotection in chilling stressed cucumber seedlings. (a) ΦPSII; (b) Fv/Fm; (c) qP; (d) NPQ. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 24 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are means ± SD (n = 3). Different letters indicate that the mean values are significantly different among the samples (p < 0.05).

    • Figure 11. 

      Effects of MT and ABA on chlorophyll a fluorescence kinetics of the OJIP curve and PSII -related protein level of cucumber under chilling stress. (a) OJIP curve; (b) Vt; (c) ΔVt; (d) PIABS (e) ФD0; (f) ψ0; (g) D1 protein; (h) PsbS protein; (i) VDE protein. Rubisco (RBC) protein is shown as an equal loading control by Coomassie Brilliant Blue staining. The value above each line is the relative enrichment of the protein. Western blotting was performed three times with three independent biological samples and similar results were obtained. At two-leaf stage, cucumber seedlings were foliar sprayed with 75 μM MT, 100 μM ABA, 100 μM p-CPA+100 μM ABA, 3 mM Na2WO4+75 μM MT, 50 μM Flu+75 μM MT and H2O. After 24 h, seedlings were exposed to 8 °C /5 °C for 24 h. The half of H2O pretreated seedlings under normal temperature (25 °C /18 °C) were as the control. All values shown are means ± SD (n = 3). Different letters indicate that the mean values are significantly different among the samples (p < 0.05).