Transcriptomic analysis to understand the alternative splicing event on regulating secondary metabolism during flowering stages of Lonicera japonica

Lonicera japonica Thunb. is an important traditional Chinese medicinal resource. The synthetic pathways, as well as the functional genes involved in the biosynthesis of phenolic acids and flavonoids, have been intensively studied; however, the mechanism of post-transcriptional regulation during the flowering stages remains largely unexplained. In this study, the L. japonica transcriptome and genome were used to identify the alternative splicing (AS) events that occur during L. japonica flowering and the relevant genes. A total of 31,739, 30,752, 31,147, and 32,071 AS events were identified based on a comparison of flowering stages with the first flowering stage, and 20,916, 20,501, 20,632, and 21,020 genes were involved in the AS events. Exon skipping (ES) has been identified as the most frequent type of alternative splicing event, whereas mutually exclusive exons (MXE) are typically among the least frequently occurring types. Functional analysis of genes underlying the two types of AS events showed these genes were mainly related to protein metabolism, RNA metabolism, cell cycle, transport, signaling, and secondary metabolism. Further pathway mapping analysis demonstrated the occurrence of AS event in chalcone isomerase (CHI), 2'-hydroxyisoflavone reductase (IFR), tyrosine decarboxylase (TYDC), and strictosidine synthase (STR) genes, which are responsible for the production of liquiritigenin, vestitone, tyramine, and strychnine, respectively. PCR and sequencing confirmed these AS events, further validating our findings. Our study elucidates the correlation between alterations in secondary metabolism and alternative splicing events during L. japonica flowering, which enhances our understanding of the complex regulation of its secondary metabolism and provides a basis for subsequent in-depth exploration of the regulatory mechanisms of active ingredient synthesis in L. japonica.