Integrated Spatial Multi-Omics Reveals Metabolic Hotspots of Paclitaxel Biosynthesis in Taxus yunnanensis

Paclitaxel (Taxol) is a broad-spectrum anticancer drug, but its clinical application is limited by insufficient production and supply. Although previous studies have noted regional differences in paclitaxel metabolism within stem tissues, the spatial compartmentalization characteristics remain unclear due to the lack of effective analytical methods. Here, we first profiled stem segments of a single Taxus yunnanensis stem spanning six longitudinal positions from juvenile (P1) to mature (P6) tissue by DESI-MSI spatial metabolomics. We then integrated spatial metabolomics with Stereo-seq spatial transcriptomics and single-nucleus RNA-seq at position P2 using three biological replicates, through cross-modality alignment of adjacent sections at the bin level (40 μm). Our findings reveal that intermediate metabolites in the paclitaxel biosynthesis pathway exhibit positive spatial autocorrelation, forming two types of spatial microdomains (metabolic hotspots) with distinct accumulation patterns, one enriched in early scaffold-forming and the other in late side-chain intermediates. Compared with surrounding tissues, these hotspots had relatively higher relative abundancesof paclitaxel intermediates and elevated expression of paclitaxel biosynthesis-related genes, suggesting functional compartmentalization. At hotspot boundaries, ABC transporter genes (such as TyuABCG15) showed higher expression than either inside the hotspots or in the surrounding tissue, suggesting their possible involvement in intercellular transport of paclitaxel intermediates. This study therefore proposes a spatially coordinated multicellular model for paclitaxel biosynthesis, providing new insights into the spatial regulation of plant secondary metabolite biosynthesis and transport.