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Figure 11 | Journal of Molecular Signaling

Figure 11

From: Defining regulatory and phosphoinositide-binding sites in the human WIPI-1 β-propeller responsible for autophagosomal membrane localization downstream of mTORC1 inhibition

Figure 11

PIKfyve inhibition elevates the number of cells that display WIPI-1 puncta, and WIPI-1 downregulation decreases LC3 lipidation. (A) Human G361 cells were treated for 3 h either with YM201636 (YM) or wortmannin (WM) diluted in complete control medium (CM) with or without serum (FCS) or in serum- and amino acid-free medium (NF), followed by immunostaining of endogenous WIPI-1 using anti-WIPI-1/Alexa 488 antibodies and quantitative fluorescence microscopy. Results are expressed as the number of WIPI-1 puncta-positive cells (150 cells per condition, n = 3). P-value: ** < 0.01. (B) G361 cells were transfected with GFP-labelled control shRNA or shRNA targeting WIPI-1, and GFP-expressing cells were sorted into two groups with regard to high or low GFP intensities. Anti-WIPI-1 and anti-LC3 ECL analysis was conducted and signal intensities normalized over GAPDH. Representative results are shown (n = 2).

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Journal of Molecular Signaling

ISSN: 1750-2187