Map-based cloning of Zmccr3 and its network construction and validation for regulating maize seed germination.

key messageMap-based cloning of Zmccr3 for regulate SG and its molecular regulatory pathway was performed and validated. WGCNA, target genes/pathways during the process of seed dormancy formation were obtained. Seed dormancy (SD) and pre-harvest sprouting (PHS) affect the grain yield and quality of grain in cereal and hybrid seed production. Although the benefits of studying SD and seed germination (SG) during seed development are well established, research into the genetic variation and molecular regulation of SD, particularly during the transition from SD to SG, remains very limited. In this study, bulked segregant analysis (BSA) and linkage analysis were used to map the QTL for the maize vp16 mutant of PHS. Using genetic and biological methods, the candidate gene was identified as Zmccr3, encoding cinnamoyl-CoA reductase 3 (ccr3), which is involved in the phenylalanine pathway of lignin metabolism and affects SG. Based on RNA-seq (RNA sequencing) at two stages of grain development with extreme PHS traits, a weighted gene coexpression network analysis (WGCNA) related to SD and SG formation was constructed, and ten target genes and three pathways during the transition from SD to SG were identified. Simultaneously, the Zmccr3 pathway was established and validated, involving upstream lipid metabolism, redox modification and degradation of cell wall oligosaccharides (as electrophilic compounds), regulation of GA signaling and intracellular ROS homeostasis, and downstream oxidation of cell wall lignin units and synthesis of phenolic compounds that affect endosperm weakening and cell wall loosening, ultimately regulating SG or SD. Therefore, we propose the Zmccr3 hypothesis to elucidate its possible functions. These findings have important theoretical and practical implications for understanding the genetic basis of PHS and SD in maize, increasing genetic resources and improving traits.