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. 2021 May 11;12:2673. doi: 10.1038/s41467-021-22882-x

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — a Relative Shiga toxin (STX; 5 μg/ml) uptake in HeLa cells treated with siRNAs against the indicated proteins. STX uptake into SCR-siRNA-treated control cells (n = 50 cells, set to 1) or into cells depleted of INPP5A (n = 47 cells), PIP5K1B (n = 86 cells), or PI3KC2γ (PI3K2CG, n = 47 cells). One sample t-test with Benjamini–Hochberg correction: PIP5K1B: p = 0.0343, t = 5.261, df = 2. INPP5A: p = 0.0343, t = 5.279, df = 2. PI3K2CG: p = 0.0138, t = 14.7, df = 2. Data represent mean ± SEM, n = 3 independent experiments. b Representative immunoblot from three independent experiments for INPP5A and β-actin of HeLa cells treated with control (SCR) or INPP5A siRNA (5AKD). Asterisk denotes non-specific background band decorated by INPP5A antibodies. c Representative confocal images of HeLa cells treated with siRNA control (SCR) or INPP5A siRNA (5AKD) and incubated with labeled STX (magenta) for 15 min at 37 °C (uptake) or kept at 4 °C for 45 min (surface binding). Blue, DAPI-stained nuclei. Scale bar, 25 μm. d Quantification of cells similar to those shown in c. Data for SCR-control siRNA-treated cells were set to 1. One sample t-test for each pair. Uptake: p = 0.0026, t = 19.66, df = 2. Surface: p = 0.0019, t = 22.81, df = 2. Data represent mean ± SEM from three independent experiments, [number of cells for uptake: (n1) SCR: 329, INPP5A: 243, (n2) SCR: 341, INPP5A: 261 (n3) SCR: 295, INPP5A: 344, Surface: (n1) SCR: 54, INPP5A: 104, (n2) SCR: 340, INPP5A: 183 (n3) SCR: 243, INPP5A: 205]. e Uptake of EGF or transferrin in HeLa cells treated with control (SCR, set to 1) or INPP5A siRNA (5AKD). One sample t-test, EGF: p = 0.4458, t = 0.8754, df = 3. Trf: p = 0.5912, t = 0.5830, df = 3. Data are mean ± SEM from n = 4 (EGF uptake) or 5 (Tfr uptake) independent experiments. Numerical source data and unprocessed blots are reported in the Source Data file.