Berberine bridge enzymes (BBEs), functioning as carbonate oxidases, enhance disease resistance in and tobacco. However, the understanding of BBEs' role in monocots against pathogens remains limited. This study identified 81 TaBBEs with FAD_binding_4 and BBE domains. Phylogenetic analysis revealed a separation of the BBE gene family between monocots and dicots. Notably, RNA-seq showed 's significant induction by both pathogen-associated molecular pattern treatment and f. sp. () infection at early stages. Subcellular localization revealed TaBBE64 at the cytoplasmic membrane. Knocking down compromised wheat's resistance, reducing reactive oxygen species and promoting fungal growth, confirming its positive role. Molecular docking and enzyme activity assays confirmed TaBBE64's glucose oxidation to produce HO. Since relies on living wheat cells for carbohydrate absorption, TaBBE64's promotion of glucose oxidation limits fungal growth and resists pathogen infection. This study sheds light on BBEs' role in wheat resistance against biotrophic fungi.
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