The transcription factors TaABI4 and TaNAC‐A1 fine‐tune TaCYP94‐A1 transcription to modulate seed dormancy in wheat

Seed dormancy (SD) is the primary genetic determinant of pre‐harvest sprouting (PHS) resistance. However, the molecular mechanisms underlying SD remain incompletely understood. Here, we identified a wheat cytochrome P450 gene, TaCYP94‐A1 , that is expressed at significantly higher levels in weak‐dormancy varieties than in strong‐dormancy varieties. TaCYP94‐A1 expression increased during SD release and decreased during dormancy establishment. Knockout of TaCYP94‐A1 markedly enhanced SD and PHS resistance without adversely affecting yield‐related traits. Two key single‐nucleotide polymorphisms (T/C at –1,895 bp and T/C at –1,225 bp) in the TaCYP94‐A1 promoter were significantly associated with SD variation, with the TaCYP94‐A1 1,895C and TaCYP94‐A1 1,225C allele combination (haplotype Hap4 ) strongly associated with enhanced dormancy. Two transcription factors, TaABI4 and TaNAC‐A1, bind directly to the 5′‐ACCGC‐3′ (C, –1,895 bp) and 5′‐GACTTC‐3′ (C, –1,225 bp) motifs in the TaCYP94‐A1 promoter, respectively, and regulate its transcription through antagonistic protein–protein interactions in the nucleus. Physiological, biochemical, and gene expression analyses revealed that the TaABI4/TaNAC‐A1–TaCYP94‐A1 module regulates SD through crosstalk with the gibberellic acid, abscisic acid, and jasmonic acid pathways. Together, these findings uncover a previously uncharacterized regulatory module controlling SD and provide valuable genetic resources and molecular markers for developing PHS‐resistant wheat cultivars through molecular design breeding.