A regulatory network underlying glume hardness revealed by transcriptomic analysis of an ems-induced mutant in synthetic hexaploid wheat

Abstract Background Tenacious glume a key trait limiting threshing efficiency in synthetic hexaploid wheat (SHW), represents an ancestral defense mechanism inherited from wild progenitors. Despite its evolutionary importance, the molecular basis of this domestication-associated trait remains poorly understood. To address this, we performed a comprehensive transcriptomic analysis of developing spikes from wild-type (tenacious glume)- and EMS-induced mutant (softglume) at 0, 5, and 10 days post-heading (DPH). Results Multi-factorial analysis identified 676 genes exhibiting significant developmental stage × glume type interactions, highlighting 5 DPH as a critical temporal window for trait establishment. Weighted gene co-expression network analysis (WGCNA) revealed a strongly correlated module (r = 0.92) that functions as the core transcriptional regulon underlying glume hardness. This module drives a dual-layered structural reinforcement program. Coordinated upregulation of CAD (Cinnamyl alcohol dehydrogenase) and COMT (Caffeic acid O-methyltransferase) in the phenylpropanoid pathway promotes lignin deposition, increasing mechanical rigidity, while activation of cuticle biosynthesis, lipid modification, and glycine-rich protein genes strengthens the epidermal barrier. This coordinated process is regulated by MYB and MIKC_MADS transcription factors acting as central hubs, and the co-enrichment of flavonoid biosynthesis and photoprotection pathways indicates that glume hardness is linked to integrated structural and chemical defense mechanisms. Conclusion Together, these findings delineate the molecular and temporal architecture of glume persistence in SHW. The identified transcriptional regulators provide potential genetic targets f to separate the tenacious glume phenotype from beneficial wild alleles, thereby facilitating the improvement of threshing efficiency without compromising adaptive traits.