Photoperiod drives cannabinoid and flavonoid biosynthesis via coordinated transcriptional and gene-metabolite regulatory networks in industrial hemp (Cannabis sativa L.) flowers

Incorporating cannabis into contemporary agriculture requires a detailed understanding of how photoperiods regulate flowering time and production of specialized metabolites. This study employed a multiomic approach to examine the effects of different photoperiods (8, 10, and 12 hours of light) on cannabinoid and flavonoid accumulation in industrial hemp ( Cannabis sativa L. cv. LN6) flowers under a constant photosynthetic photon flux density (PPFD). Key cannabinoids (CBD, CBDA, CBDV), total flavonoid, and proanthocyanidin contents significantly (p < 0.05) increased with increasing light exposure. Differentially accumulated metabolites (DAMs) and differentially expressed genes (DEGs) were identified across all photoperiod comparison groups, with the 8 h versus 12 h (S8F vs. S12F) comparison showing the largest number of integrated DEG-DAM pairs. Flavonoid and phenylpropanoid biosyntheses, along with sesquiterpenoid and triterpenoid, were found to be the most significantly photoperiod-responsive pathways. A co-expression network analysis of 276 DEGs identified three major transcriptional modules (ME1-ME3), with ME1 strongly correlated with proanthocyanidin (r = 0.808) and total flavonoid (r = 0.527) accumulation and enriched for major flavonoid biosynthetic genes, which was subsequently confirmed by qRT-PCR. A gene-metabolite correlation network of 120 nodes and 1367 edges showed a dense topology, with genes linked to the terpenoid, phenylpropanoid, and flavonoid pathways, along with their related metabolites, reflecting coordination among multiple biosynthetic pathways. Many triterpenoid/cannabinoid metabolites showed a strong photoperiod-responsive accumulation pattern that correlated with the expression of biosynthetic genes. These findings provide a deeper understanding of how photoperiod modulates plant metabolic networks and offer practical implications for enhancing the synthesis of high-value metabolites through photoperiod management in controlled systems.