Trypanosoma cruzi epimastigotes internalize macromolecules avidly by endocytosis. Previously, we identified a tubule-vesicular network likely to correspond to the early-endosomes. However, a detailed ultrastructural characterization of these endosomes was missing. Here, we combined endocytosis assays with ultrastructural data from high-resolution electron microscopy to produce a 3D analysis of epimastigote endosomes and their interactions with endocytic organelles. We showed that endocytic cargo was found in carrier vesicles budding from the cytopharynx. These vesicles appeared to fuse with a tubule-vesicular network of early endosomes identified by ultrastructural features including the presence of intermembrane invaginations and coated membrane sections. Within the posterior region of the cell, endosomes localized preferentially on the side nearest to the cytopharynx microtubules. At 4 °C, cargo accumulated at a shortened cytopharynx, and subsequent temperature shift to 12 °C led to slow cargo delivery to endosomes and, later, to reservosomes. Bridges between reservosomes and endosomes resemble heterotypic fusion. Reservosomes are excluded from the posterior end of the cell, with no preferential cargo delivery to reservosomes closer to the nucleus. Our 3D analysis indicates that epimastigotes accomplish high-speed endocytic traffic by cargo transfer to a bona fide early-endosome and then directly from endosomes to reservosomes, via multiple and simultaneous heterotypic fusion events.

克氏锥虫鞭毛体通过胞吞作用使大分子内在化。以前，我们确定了可能与早期内体相对应的肾小管-囊泡网络。但是，缺少这些内体的详细超微结构表征。在这里，我们将胞吞作用测定与高分辨率电子显微镜的超微结构数据相结合，以产生对鞭毛鞭毛体内体及其与内吞细胞器相互作用的3D分析。我们表明，在从细胞咽部出芽的载体囊泡中发现了内吞性货物。这些囊泡似乎与早期内体的肾小管-囊泡网络融合，这些囊泡通过超微结构特征（包括膜间内陷和包被的膜部分）得以鉴定。在细胞的后部区域，内体优先位于最靠近细胞咽部微管的一侧。在4°C时，货物聚集在缩短的咽中，随后温度移至12°C，导致货物缓慢递送到内体，然后再递送到储库。储体和内体之间的桥梁类似于异型融合。储库被排除在细胞的后端之外，没有优先的货物运送到更靠近细胞核的储库。我们的3D分析表明，前鞭毛体通过多次和同时发生的异型融合事件，通过货物转移到真正的早期内体，然后直接从内体传递到储库，来完成高速内吞运输。随后温度移至12°C，导致向内体和随后向储体的货物运输缓慢。储体和内体之间的桥梁类似于异型融合。储库被排除在细胞的后端之外，没有优先的货物运送到更靠近细胞核的储库。我们的3D分析表明，前鞭毛体通过多次和同时发生的异型融合事件，通过货物转移到真正的早期内体，然后直接从内体传递到储库，来完成高速内吞运输。随后温度移至12°C，导致向内体和随后向内体的货物运输缓慢。储体和内体之间的桥梁类似于异型融合。储库被排除在细胞的后端之外，没有优先的货物运送到更靠近细胞核的储库。我们的3D分析表明，前鞭毛体通过多次和同时发生的异型融合事件，通过货物转移到真正的早期内体，然后直接从内体传递到储库，来完成高速内吞运输。