Melatonin improves cold storage tolerance in cucumber via CsMYB44-mediated transcriptional activation of the polyamine biosynthesis gene family

Cucumber fruit is susceptible to chilling injury (CI) under cold storage conditions. The application of melatonin can mitigate CI in cucumber fruit; however, the underlying molecular mechanisms are not well understood. Polyamines (PAs) are important biomolecules that have regulatory roles in cold stress responses, and this study identified 15 PAs-encoding gene members in cucumber by using bioinformatics techniques and their genome attributes were characterized. The cucumber fruit was treated with 100 µM melatonin for 15 min and stored for 15 d at 4 °C. The results showed that melatonin reduced the CI levels. Similarly, melatonin treatment reduced the incidence of decay and inhibited malonaldehyde content and ion leakage. Among the identified PAs member genes, CsADC, CsODC1, CsODC2, CsSAMDC1, CsSAMDC2, CsSPDS1, CsSPDS2, CsSPDS3, CsSPDS4, CsSPDS5, CsACL5–1, CsACL5–2, and CsACL5–3 were upregulated in melatonin-treated fruits. Besides, CsMYB44 expression is upregulated by cold stress and melatonin treatment. Thus, CsMYB44 TF was cloned from cucumber fruit. Furthermore, subcellular localization experiments showed that CsMYB44 is localized in the cell nucleus. The yeast two-hybrid assay revealed the self-activation capacity of CsMYB44. Additionally, a dual-luciferase assay revealed that CsMYB44 acts as a transcriptional activator of the corresponding promoters of CsADC, CsODC2, and CsSPDS5. Similarly, luciferase complementation imaging revealed intense fluorescence in the CsMYB44-injected tobacco leaves. These findings suggest that CsMYB44 plays a transcriptional activator role in PAs genes, alleviating CI, and providing helpful insights into postharvest cold storage tolerance in cucumber fruit.