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. 2016 Jan 18;212(2):181–198. doi: 10.1083/jcb.201506081

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© 2016 Liu et al.

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Figure 8. — Human WDR91 and WDR81 are recruited to endosomes and are important for endolysosomal trafficking. (A) Colocalization of WDR91 with WDR81 (arrows) in immunostained HeLa cells. Insets show magnified (1.8×) views of the box in the merged image. (B and C) Coimmunostaining of WDR91 or WDR81 with EEA1 (B) or Rab7 (C). Insets show magnified (1.8×) views of the boxes in the merged images. Arrows indicate colocalization of WDR91 or WDR81 with EEA1 or Rab7. Bars: (A–C) 5 µm. (D) Time course of 2xFYVE::mCherry and GFP-WDR91 (top) and GFP-WDR81 (bottom) on endosomes. Bars, 2 µm. (E) Top: sgRNA targeting sites in WDR81 and WDR91 cDNAs. Numbers indicate nucleotides (nt) starting from the first ATG. Bottom right: Western blot of WDR91 in control (Ctrl), WDR91 knockout (KO-91), and WDR81 knockout (KO-81) HeLa cells. (F) Immunostaining of WDR91 or WDR81 in Ctrl, KO-91, and KO-81 cells. Red, WDR91 or WDR81 antibody; blue, DAPI. Bars, 10 µm. (G) Ctrl, KO-91, and KO-81 cells were treated with EGF for 10 min, and then EGFR levels were examined by Western blot (left). EGFR levels were quantified with ImageJ software (right). Data representing mean ± SEM are from three independent experiments.