Physiological Characteristics and Related Gene Expressions Associated with Moist Chilling-Induced Seed Dormancy Release in Zoysiagrass (Zoysia japonica)

Moist chilling is widely used to overcome seed dormancy in zoysiagrass (Zoysia japonica Steud.), but the coordinated physiological and molecular basis remains unclear. Here, freshly matured seeds were subjected to moist chilling at 4 °C in darkness for 0 (Control), 1 (CS1), 2 (CS2), 3 (CS3), or 4 weeks (CS4) and then transferred to germination conditions (30/20 °C, day/night). Prolonged moist chilling progressively improved dormancy release: final germination percentage increased from 40.5% (Control) to 73.5% (CS4), accompanied by a higher germination index and earlier, faster cumulative germination dynamics. Moist chilling also enhanced early seedling vigor, with stronger treatment differentiation in root elongation than in shoot growth. Physiologically, abscisic acid (ABA) content declined while gibberellic acid (GA) content increased, resulting in an elevated GA/ABA ratio with prolonged chilling. Metabolic activation was evidenced by increased α-amylase activity, greater soluble sugar and soluble protein accumulation, and stimulated oxygen uptake. In addition, CAT, SOD, and POD activities were enhanced under prolonged moist chilling, whereas H2O2 levels remained relatively stable, suggesting that redox adjustment during dormancy release was characterized by strengthened antioxidant buffering rather than pronounced oxidative accumulation. qRT-PCR supported a mechanistic transition from dormancy maintenance to germination execution, showing moist chilling-associated regulation of ABA/GA metabolism and signaling genes (e.g., NCED, CYP707A, ABI3/ABI5, and GA20ox) and downstream metabolic modules (e.g., GAMYB, AMY, ISA, INV, and HXK1), together with concurrent modulation of respiration- and ROS-related markers (e.g., AOX1a, RBOH, and CAT). Correlation analysis linked germination performance most strongly with α-amylase activity, oxygen uptake, and the GA/ABA ratio. Collectively, our data support a working model in which moist chilling rebalances the ABA–GA gate and activates downstream metabolic and redox adjustment modules to promote dormancy release and improve germination performance in zoysiagrass, providing practical markers for optimizing seed establishment through moist chilling treatment.