Fig. 3. Endocytosis is perturbed in BigEye cells. (A) In uninduced TbCLH RNAi cells at 4°C, lectin concanavalin A (Con A) is retained at the flagellar pocket, at 12°C the lectin is internalized to the endocytic system and at 37°C the lectin is delivered to the terminal lysosome. Induced cells demonstrate a severe defect in uptake. At 37°C, the lectin remains associated with the flagellar pocket. (B) Enlarged image of induced bloodstream form (BSF) clathrin RNAi cells (from A), showing that Con A is localized to the enlarged flagellar pocket at 37°C. (C) At 4°C, FM 4-64 accumulates in the flagellar pocket in both the parental and induced clathrin RNAi cell line. Note that the staining appears rather more extensive in the RNAi cells due to the enlargement of the pocket. At 37°C, the fluorophore has been internalized in the parental cells, but remains associated with the enlarged pocket structure in the RNAi cell line. (A) Left, phase; right, DAPI (DNA) in blue, lectin in green. (C) Left, phase; right, DAPI (DNA) in blue, FM 4-64 in red. Scale bar: 2 µm. Par, BSF 90-13 parental cells; –Tet, uninduced BSFp2T7^TiCLH cells; +Tet, BSFp2T7^TiCLH cells induced for 16 h.