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. 2018 Oct 12;37(21):e99753. doi: 10.15252/embj.201899753

Figure 2. ESCRT‐I, ESCRT‐II, ESCRT‐III, ALIX, and VPS4A are recruited to endolysosomes upon LLOMe treatment.

Figure 2

To screen for ESCRT proteins that are involved in the endolysosomal repair process, cells were incubated with 250 μM LLOMe for 30 min and processed for immunofluorescence. TSG101, a component of the ESCRT‐I complex, and EAP30, an ESCRT‐II protein, are clearly recruited to the sites of damage compared to the DMSO control (Ctrl). As shown, the ESCRT‐III complex together with ALIX is recruited to damaged endolysosomes. In contrast, HRS, a component of the ESCRT‐0 complex, and HD‐PTP show no recruitment to damaged endomembranes. Number of foci per cell was quantified from >85 cells per condition from three independent experiments and are presented as mean ± SD. Statistical significance for number of foci per cell in Ctrl versus LLOMe treatment was determined using Student's t‐test, the P‐values for which are as follows: HRS P = 0.9257, GAL3 P = 0.0050; TSG101 P = 0.0243, GAL3 P = 0.0489; EAP30 P = 0.0006, GAL3 P = 0.0307; CHMP2A P = 0.0284, GAL3 P = 0.0260; CHMP4B P = 0.0028, GAL3 P = 0.0414; VPS4A P = 0.0309, GAL3 P = 0.0461; ALIX P = 0.0249, GAL3 P = 0.0471; HD‐PTP P = 0.4898, GAL3 P = 0.0105. Scale bars: 5 μm and 1 μm (inset).