Genome-wide analysis of expansin and aquaporin genes and their association with auxin-related metabolites during flower opening in Lonicera macranthoides.

INTRODUCTION: In flowering plants, expansins and aquaporins function coordinately to promote cell expansion, thereby driving flower opening. Lonicera macranthoides is a widely used medicinal plant rich in chlorogenic acid (CGA) and saponins. Its flowers have high medicinal value; however, the contents of these bioactive compounds decrease after flower opening, and open flowers are prone to abscission. To date, the expansin and aquaporin gene families in L. macranthoides have not been systematically studied, and their roles in flower opening remain unclear.

METHODS: Expansin and aquaporin genes were identified from a chromosome-level genome using hidden Markov models. Phylogenetic relationships, chromosomal localization, protein sequence features, gene structures, and cis-regulatory elements were further analyzed. Gene expression patterns were examined using transcriptomic data and quantitative real-time PCR (qRT-PCR), and auxin-related metabolites were quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS).

RESULTS AND DISCUSSION: Here, 34 expansin and 43 aquaporin genes were identified and systematically characterized. Expression analyses showed that LmEXPA26 (EXPA subfamily) and LmPIP2-2 (PIP2 subfamily) exhibited expression patterns that closely matched flower developmental progression during opening in L. macranthoides, with peak expression in S4 flowers of the flowering wild type (WT) but much lower expression in the bud-type cultivar 'Yulei 1'. Promoter analysis revealed abundant MYB-, bZIP-, and ARF-binding sites, together with multiple ABA-, JA-, Auxin-, and ethylene-responsive elements in the promoters of expansin and aquaporin genes. Transcriptome analysis revealed significant enrichment of auxin-responsive genes in WT S4 flowers. Auxin-related metabolite profiling showed higher accumulation of MEIAA, a reversibly inactive auxin form convertible to IAA, in WT S4 flowers, whereas irreversibly inactivated OxIAA accumulated predominantly in S4 flowers of 'Yulei 1'. Based on these results, the present study suggests that enhanced expression of expansins and aquaporins, especially LmEXPA26 and LmPIP2-2, may be associated with the flower-opening process in L. macranthoides, which is likely primarily influenced by MEIAA-mediated auxin activation, although other hormones may also contribute. This study provides new insights into the molecular basis of flower opening in L. macranthoides.