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. 2014 Apr 15;25(8):1327–1337. doi: 10.1091/mbc.E13-08-0447

FIGURE 2:

FIGURE 2:

VPS33A and VPS16 translocate to STX17-positive autophagosomes. (A) MEFs stably coexpressing Myc-STX17 and either GFP-VPS33A or GFP-VPS16 were cultured in regular medium or starvation medium for 2 h. Cells were costained with anti-Myc antibody and anti-LAMP1, anti-LC3, or anti-ATG16L1 antibody and analyzed by immunofluorescence microscopy. Structures positive for VPS33A, STX17, and LC3 are indicated by blue arrows, structures positive for VPS33A and STX17 but not for LAMP1 are indicated by white arrows, structures positive for VPS33A and LAMP1 but not for STX17 are indicated by white arrowheads, and structures positive for VPS16, STX17, and LC3 are indicated by yellow arrows. Signal color is indicated by the color of the text. Scale bars,10 μm. (B) HeLa cells stably expressing GFP-VPS33A were treated with siRNA against luciferase or STX17. After 72 h, cells were starved for an additional 2 h. Then cells were fixed, stained with anti-GFP and anti-LC3 antibodies, and analyzed by confocal microscopy. Structures positive for GFP-VPS33A and LC3 are indicated by white arrows. Scale bars, 10 μm. Quantification of the colocalization rate between endogenous LC3 and GFP-VPS33A is shown in the graph. Data represent mean ± SEM of at least 10 different cells. *p < 0.05 compared with the luciferase siRNA–treated group.