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

Figure 5. PI(3,5)P2 is localized to lysosomes and catalyzed by INPP5E.

Figure 5

  1. NIE‐115 cells stably expressing mSt‐2xML1N or ‐2xPLCδ1 PH were cultured in growth medium with or without 200 nM Torin1. Cells were fixed and stained with anti‐LAMP1 or anti‐LC3 antibodies and then analyzed by immunofluorescence microscopy. Insets show the boxed areas at high magnification. Scale bar, 10 μm.
  2. NIE‐115 cells stably expressing mSt‐2xML1N treated with siControl or siINPP5Es were cultured in growth medium. Cells were fixed and stained with anti‐LAMP1 antibodies and then analyzed by immunofluorescence microscopy. Insets show the boxed areas at high magnification. Scale bar, 10 μm.
  3. Quantitation of signal intensities in (B) showing mSt‐2xML1N colocalizing with LAMP1 (means ± s.d.; n > 100 cells from three independent experiments). **P < 0.01; *P < 0.05.
  4. N1E‐115 cells stably expressing GFP‐2xML1N treated with siControl or siINPP5E were transiently transfected with mSt‐INPP5E (WT, D477N). Quantitation of signal intensities showing GFP‐2xML1N colocalizing with LAMP1 (means ± s.d.; n > 20 cells from three independent experiments). **P < 0.01; *P < 0.05.
  5. NIE‐115 cells stably expressing mCherry‐2xFYVE treated with siControl or siINPP5Es were cultured in growth medium. Cells were fixed and stained with anti‐LAMP1 antibodies and then analyzed by immunofluorescence microscopy. Insets show the boxed areas at high magnification. Scale bar, 10 μm.
  6. Quantitation of signal intensities in (E) showing mCherry‐2xFYVE colocalizing with LAMP1 (means ± s.d.; n > 100 cells from three independent experiments). **P < 0.01.
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