Transcriptomic dissection of floral identity and trichome-derived metabolite pathways in Cannabis sativa L

Cannabis sativa L. has a long history of medicinal and industrial use, with female flowers as the primary source of bioactive cannabinoids and terpenoids synthesized in glandular trichomes. Despite its importance, the genetic mechanisms governing flower development, sex determination, and secondary metabolite biosynthesis remain incompletely understood. Here, we combined transcriptomic and comparative genomic analysis to elucidate the molecular networks underlying these traits. Integrating 117 RNA-Seq datasets and phylogenetic analyses across nine C. sativa genomes, we identified 31 orthogroups of MADS-box transcription factors. Expression profiling highlighted distinct candidates for male and female flower identity, consistent with the ABCDE model. AP3, PI/GLO, and MIKCS clades were preferentially expressed in male flowers, whereas AGL6, FLC-like, and Bsister genes predominated in female flowers. Analyses of pollen-associated genes underscored roles for sugar metabolism and transport in male fertility. Profiling cannabinoid and terpenoid biosynthetic genes confirmed strong trichome expression and revealed a key distinction: upstream polyketide, MEV, MEP, and the terpenoid pathways showed conserved expression across chemotypes, whereas the cannabinoid pathway displayed chemotype-specific profiles. Collectively, these findings provide an integrative framework for floral development and secondary metabolism in C. sativa, offering targets for functional validation and breeding to optimize agronomic and medicinal traits in modern cultivars.