The mannose 6-phosphate receptor system is ancient, widespread in eukaryotes, and functionally diversified in the human parasite Trichomonas vaginalis
In mammals, the mannose 6-phosphate (M6P) pathway tags newly synthesized hydrolases for delivery to the lysosomes. The pathway underlies healthy lysosomal function, but whether an equivalent system operates outside animals remains largely unexplored. Surveying 53 representative species, across eukaryotic supergroups we show that the M6P receptor (MPR), the phosphorylating enzyme GNPT, and the uncovering enzyme (UCE) form a statistically robust, co-occurring module across extant eukaryotic supergroups. In the human parasite Trichomonas vaginalis, this module has diversified into an expanded MPR family of seven paralogs, the largest reported within these 53 species, alongside a structurally minimal single GNPT gene, despite the complete absence of UCE. Using co-immunoprecipitation (co-IP), mass spectrometry and confocal microscopy, we demonstrate that MPRs and GNPT localise to the Golgi apparatus, and their interactomes show paralog-specific enrichment of distinct hydrolase cargo, adaptor complexes and the retromer complex proteins for retrieval of MPRs, presenting a full receptor-recycling cycle. Critically, mass spectrometry based glycomics directly detects mature M6P-modified N-glycans in T. vaginalis built on unique Man5GlcNAc2 structure with no trace of GlcNAc-capped intermediate, seen in mammalian cells. Together, these results provide evidence that M6P pathway is an ancient and widespread eukaryotic system, with its functionally diversified variant operating in T. vaginalis for lysosomal biogenesis.