The basic/helix-loop-helix 79 (bHLH79) - xyloglucan endotransglucosylase/hydrolase 28 (XTH28) regulatory module mediates flower opening in Osmanthus fragrans.

During the process of flower opening in Osmanthus fragrans, the epidermal cells of the petals undergo substantial expansion. While xyloglucan endotransglucosylase/hydrolases (XTHs) are known to regulate plant cell expansion, the exact role of OfXTHs remains unclear. In this study, O. fragrans 'Yanhonggui' was employed as the experimental material. Transcriptomic and qRT-PCR analyses revealed that OfXTH28 was markedly upregulated during the top bracts (TB) stage of flower opening. The OfXTH28 protein contained a conserved DExDxEFLG motif and localized to the cell wall. Heterologous overexpression of OfXTH28 in Arabidopsis thaliana resulted in enhanced enlargement of petals and epidermal cells. We also identified OfbHLH79, a bHLH transcription factor characterized by nuclear localization and a gene expression pattern similar to that of OfXTH28. Yeast one-hybrid (Y1H), dual-luciferase (Dual-LUC), and electrophoretic mobility shift assays (EMSA) confirmed that OfbHLH79 directly activates the expression of OfXTH28 by binding to the E-box motif within its promoter region. The transient overexpression of OfbHLH79 in the flower buds of O. fragrans led to a marked upregulation of OfXTH28 expression. In conclusion, this research provides a thorough framework for understanding the role of OfXTH and lays a strong groundwork for exploring the mechanisms underlying flower opening in O. fragrans.