Melatonin soaking promotes germination of aged Atractylodes macrocephala seeds by GSH cycle activation and phenylpropanoid reprogramming to support superior mitochondrial function

Abstract

Seed aging restricts seedling propagation and sustainable cultivation of Atractylodes macrocephala Koidz. (A. macrocephala), a genuine herb from Zhejiang. While melatonin (MT) alleviates germination inhibition under abiotic stress, its mechanisms in rejuvenating aged A. macrocephala seeds remain unclear. Naturally aged A. macrocephala seeds were soaked with 0.1 mM MT or distilled water for 24 h before germination assays. MT-soaked seeds showed 43.9%, 66.7%, and 136.7% higher germination rates, germination potential, and vigor index, respectively, indicating rejuvenation in aged seeds. Transcriptomic and biochemical analysis revealed that MT upregulated genes and enzyme activities involved in the reduced glutathione (GSH) cycle and reprogrammed phenylpropanoid metabolism toward flavonoid biosynthesis. The resultant increases in GSH, flavonoid, and proanthocyanidin levels enhanced antioxidant defense, which lowered O₂^•⁻ production rate, H₂O₂ and malondialdehyde contents. Restored mitochondrial ultrastructure was also observed in MT-soaked seeds. Additionally, most differentially expressed genes related to starch catabolism, glycolysis, the citrate cycle, and oxidative phosphorylation were upregulated, accompanied by elevated ATP contents. Results indicate that MT-induced rejuvenation on A. macrocephala aged seeds is associated with reinforced antioxidant defense, repaired mitochondrial structure, and improved ATP supply. This work provides a theoretical and practical basis for improving seedling quality and promoting the sustainable cultivation of medicinal herbs.