Distinct nanoscale membrane organizations of mucins and trans-sialidases in Trypanosoma cruzi

Trypanosoma cruzi, the causative agent of Chagas disease, relies on surface sialylation to evade host immunity and invade cells. This process is mediated by trans-sialidases (TS) and mucins, an enzyme-substrate pair anchored to distinct lipid environments. Yet, how these molecules are organized at the nanoscale in the parasite's membrane remains unknown. Using dual-color super-resolution microscopy, we show that ~60% of mucins and TS are segregated into nanoclusters (~100 nm) that rarely contact each other, distributed with a nonrandom separation distance, indicating that these abundant domains follow a specific order in the plasma membrane and are unlikely to serve as primary sites of sialylation. In contrast, the ~40% fraction of non-clustered mucins and TS exhibits significantly shorter-than-random separation distances and appear ordered as in a shared fibrillar network. Additionally, we describe a distinct structural organization within these domains: mucins, residents of detergent-resistant domains (DRDs), organize into high molecular-weight complexes, whereas TS (which are excluded from DRDs) do not, suggesting a potential mechanism for domain-specific protein localization.