Melatonin antagonizes ABA action to promote seed germination by regulating Ca²⁺ efflux and H₂O₂ accumulation

Seed germination is a vital stage in the plant life-cycle that greatly contributes to plant establishment. Melatonin has been shown to promote seed germination under various environmental stresses; however, the mechanism remains largely underexplored. Here, we reported that melatonin antagonized abscisic acid (ABA) to promote seed germination by regulating ABA and gibberellic acid (GA₃) balance. Transcriptomic analysis revealed that such a role of melatonin was associated with Ca²⁺ and redox signaling. Melatonin pretreatment induced Ca²⁺ efflux accompanied by an up-regulation of vacuolar H+/Ca²⁺ antiporter 3 (CAX3). AtCAX3 deletion in Arabidopsis exhibited reduced Ca²⁺ efflux. Inhibition of Ca²⁺ efflux in the seeds of melon and Arabidopsis mutant AtCAX3 compromised melatonin-induced germination under ABA stress. Melatonin increased H₂O₂ accumulation, and H₂O₂ pretreatment decreased ABA/GA₃ ratio and promoted seed germination under ABA stress. However, complete inhibition of H₂O₂ accumulation abolished melatonin-induced ABA and GA₃ balance and seed germination. Our study reveals a novel regulatory mechanism in which melatonin counteracts ABA to induce seed germination that essentially involves CAX3-mediated Ca²⁺ efflux and H₂O₂ accumulation, which, in turn, regulate ABA and GA₃ balance by promoting ABA catabolism and/or GA₃ biosynthesis.