Skip to main content
. 2016 Jun 23;35(17):1853–1867. doi: 10.15252/embj.201593148

Figure 6. INPP5E enhances actin polymerization on lysosomes through cortactin activation.

Figure 6

  1. N1E‐115 cells treated with siControl or siINPP5Es were cultured in growth medium. Cells were fixed and stained with anti‐LAMP1 antibodies and phalloidin and then analyzed by immunofluorescence microscopy. Insets show the boxed areas at high magnification. Scale bar, 10 μm. Quantitation of signal intensities showing phalloidin colocalizing with LAMP1 (means ± s.d.; n > 40 cells from three independent experiments). **P < 0.01.
  2. N1E‐115 cells treated with siControl or siINPP5Es were cultured in growth medium. Cells were fixed and stained with anti‐phospho‐Y421 cortactin and anti‐LAMP1 antibodies and then analyzed by immunofluorescence microscopy. Insets show the boxed areas at high magnification. Scale bar, 10 μm. Quantitation of signal intensities showing cortactin (pY421) colocalizing with LAMP1 (means ± s.d.; n > 40 cells from three independent experiments). **P < 0.01.
  3. N1E‐115 cells were treated with 0.5 μM latrunculin A (Lat A) in growth medium for 6 h. Cells were fixed and stained with anti‐LC3 antibodies and then analyzed by immunofluorescence microscopy. Scale bar, 10 μm. Quantitation of the number of LC3 puncta per cell (mean ± s.d.; n > 100 cells from three independent experiments). **P < 0.01.
  4. NIE‐115 cells stably expressing LAMP1‐mCherry were treated with DMSO or 0.5 μM Lat A in growth medium for 4 h, subsequently cultured with 200 nM Torin1 and protease inhibitors (10 μg/ml pepstatin A and 10 μg/ml E‐64‐d) for 2 h (Lat A treatment for total 6 h). Cells were fixed and stained with anti‐LC3 antibodies and then analyzed by immunofluorescence microscopy. Insets show the boxed areas at high magnification. Scale bar, 10 μm. Percentages of colocalization of LC3 dots with LAMP1 dots (mean ± s.d.; n > 20 cells from three independent experiments). **P < 0.01.