Abscisic acid and hydrogen peroxide regulate proline homeostasis in melon seedlings under cold stress by forming a bidirectional closed loop

Low temperatures severely limit melon cultivation and production in winter and spring. Cold stress can induce proline accumulation in melon seedlings. However, the accumulation mechanism is unclear and the way by which abscisic acid (ABA) and hydrogen peroxide (H2O2) act as signaling molecules in the process remains unknown. We first identified key enzyme genes in the proline metabolism pathway, and identified six genes, namely, two CmP5CSs and one each of CmP5CR, CmOAT, ProDH, and CmP5CDH. Low temperature induced proline accumulation by up-regulating three synthase genes, CmP5CS1, CmP5CR, and CmOAT, while suppressing the expressions of two catabolic genes, namely, CmProDH and CmP5CDH. Cold stress triggered ABA and H2O2 synthase genes to activate their accumulation. Both H2O2 and ABA pretreatments enhanced melon seedling's cold tolerance, but the two behaved divergently in proline metabolism. Exogenous H2O2 enhanced or maintained the aforementioned induction efficiency of low temperature. In contrast, ABA only induced CmP5CR and CmOAT expression, with no effect on CmP5CS1, and synchronously induced CmProDH and CmP5CDH expressions, leading to a slight reduction in proline content. Alternatively, ABA controlled proline homeostasis under cold stress, at least in part, by inhibiting H2O2 generation. Overall, the results suggest that ABA and H2O2 formed a fine bidirectional closed loop to regulate proline homeostasis in melon seedlings.