Live cell imaging showing localization of Vps29-YFP to endosomal domains that contain CFP-Rab7 but not CFP-Rab5a. HeLa cells cotransfected with plasmids encoding Vps29-YFP (green) and either CFP-Rab7 (A–C) or CFP-Rab5a (D–F; red) were imaged by time-lapse fluorescence microscopy. Pictures in this figure were extracted from Videos 1–6 (available). Merging the green and red images generated the third picture in A and D and the images in the bottom rows of B, C, E, and F. Yellow indicates overlapping localization of green and red objects. Examples of colocalization are indicated by yellow arrows. The images in A and the series in B and C show the localization of Vps29-YFP to domains that contain CFP-Rab7 (indicated by green and red arrows in individual channel images or yellow arrows in merge images). The series in C shows an endosome in which Vps29-YFP (green arrows) is enriched on a detaching tubule that does not contain CFP-Rab7 (red arrows). The images in D and the series in E and F show that Vps29-YFP (green arrows) and CFP-Rab5a (red arrows) are localized to different endosomal domains. The series in F shows a Vps29-YFP–positive endosome with little or no CFP-Rab5 (green arrows). Time after the start of imaging (in seconds) is shown on the bottom left corner of each panel. Bars: (A and D) 5 μm; (B, C, E, and F) 2 μm.