Decoding pilocarpine biosynthesis and its roles in Pilocarpus microphyllus through a comparative transcriptomics approach

Pilocarpus microphyllus, widely known as jaborandi, faces a decline in its natural population due to unsustainable harvesting for pilocarpine extraction, an imidazole alkaloid with significant pharmacological properties. This study presents the first comparative transcriptomic analysis in jaborandi by investigating gene expression across four different tissues (leaflets, rachis, root, and stem) and exploring potential pathways and functions involved in pilocarpine biosynthesis. The comparisons involving the root had the highest number of DEGs, including root vs. leaflets, rachis vs. root, and stem vs. root. In contrast, no DEGs were identified in the comparison between stem and leaflets. We observed that the root exhibited the most diverse functional profile, including an abundance of TFs involved in alkaloid biosynthesis. Functional enrichment analysis of root-overexpressed genes revealed associations with cell transport, regulatory processes, and defense responses. In contrast, aerial tissues exhibited enrichment for photosynthesis and oxidative stress response pathways, with antioxidant enzymes highly expressed in the leaflets—the primary site of pilocarpine accumulation in its final form—suggesting a potential link between pilocarpine production and the plant’s antioxidative response. The presence of enzymes potentially involved in pilocarpine biosynthesis in both aerial and root tissues supports the idea that its biosynthesis is a multi-tissue process. This study provides important insights into the metabolic pathways for advancing conservation strategies and promoting sustainable management of this species.