Correlation Research of Corolla Non-Opening Phenotype with Cell Morphology and XTH/EXP-PME Expression Characteristics in Lonicera macranthoides
Dried flower buds or newly opened flowers of Lonicera macranthoides are used as Shanyinhua (Lonicerae Flos) for medicinal purposes. The Xianglei cultivar, developed from a natural mutant, exhibits a distinctive phenotype characterized by non-opening corolla and prolonged bud stage. This study aimed to elucidate the cellular and molecular mechanisms underlying this unique corolla phenotype. The flower of conventional cultivar (WT) and the Xianglei cultivar (XL) of L. macranthoides were used as materials. Corolla adaxial epidermal cells and longitudinal sections were observed using light microscopy. The expression patterns of xyloglucan endotransglucosylase/hydrolase genes (LmXTH6, LmXTH7, LmXTH33), expansin gene (LmEXP7), and pectin methylesterase genes (LmPME28, LmPME45) were analyzed by qRT-PCR. Protein-protein interaction networks were predicted using the STRING database. Ruthenium red staining coupled with Image J quantitative analysis was performed to assess the degree of pectin de-methylesterification in corolla cell walls. In the WT, the adaxial cells of corolla displayed a spatially sequential expansion, with the basal cells expanding first, followed by the middle cells. Moreover, the expansion rate of the adaxial cells is higher than that of the abaxial cells. In contrast, the expansion rate of adaxial cells in XL was significantly reduced, and the degree of cell expansion was not obvious. At the molecular level, LmXTH6, LmXTH7, and LmXTH33 were highly expressed in WT during the pre-anthesis and anthesis stages, but their expression was significantly inhibited in XL. LmEXP7 expression peaked before corolla opening in WT, while its expression was lower in XL during the same period. Ruthenium red staining results indicated that the overall degree of de-methylesterification in XL was higher than that in WT, consistent with the expression trends of LmPME28 and LmPME45. In conclusion, the non-opening corolla of the Xianglei cultivar is closely related to the inhibition of corolla cell expansion. The low expression of XTHs and EXP collectively leads to a decrease in the expansion capacity of Xianglei corolla cells, while the up-regulation of PMEs increases pectin de-methylesterification in the cell wall, potentially enhancing cell wall rigidity. The synergy of XTH/EXP-PME may be an important reason for the non-opening corolla of Xianglei, providing experimental evidence for the study of the regulatory mechanism of the excellent phenotype of Xianglei L. macranthoides and offering new insights into the cell wall regulatory mechanisms of plant floral organ morphogenesis.