Figure 7.

Endocytic compartments can be ordered using the shi^ts1 mutation. HRP-Boss was localized by EM in shi^ts1 eye disks after blocking endocytosis at 30°C. After 1 h, the majority of HRP activity was detected in multilammelar late endosomes or lysosomes (A). Only rarely did we detect remnants of HRP activity in MVBs (B). After 2 h at the nonpermissive temperature, strong staining in multilammelar late endosomes and lysosomes persisted (C–E), whereas labeled MVBs could not no longer be detected. Strikingly, after blocking shibire function, unstained MVBs were detected in R7 cells (A, D, and E, open arrows). A synchronized wave of endocytosis was released by holding shi^ts1 larvae at 30°C for 1 h and than returning them to 22°C for 15 min before visualizing HRP activity (F). The relevant genotypes of the larvae were w¹¹¹⁸ shi^ts1; P[w⁺, gen.HRP-Boss]¹⁷. Bar: A, C, and E, 250 nm; B, D, and F, 400 nm.