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. 2023 Mar 16;14(3):199. doi: 10.1038/s41419-023-05723-1

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© The Author(s) 2023

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. 2 — A, B HCT-15 cells were transiently transfected with the plasmids encoding Lon or Lon-shRNA. Cell lysates were analyzed by immunoblotting using the indicated antibodies. GAPDH as the loading control. C HCT-15 cells transfected with the plasmids encoding Lon or empty were treated with SBI-0206965 (20 μM for 6 h) or not. Cell lysates were analyzed by immunoblotting using the indicated antibodies. GAPDH as the loading control. D HCT-15 cells were treated with CoCl₂ (200 μM for 18 h) or not in the presence or absence of SBI-0206965 (20 μM for 6 h). Cell lysates were analyzed by immunoblotting using the indicated antibodies. GAPDH as the loading control. E, F Mitochondrial Lon induces lysosome-mediated autophagy. E Mitochondrial Lon-induced lysosome-mediated autophagy were verified by immunofluorescence. HCT-15 cells were transfected with the plasmids encoding Lon or empty and treated with BafilomycinA1 (100 nM for 6 h) or not. The cells were fixed and immunostained by RFP-LC3B (red) and anti-LAMP1 (green) antibodies. Scale bars = 50 μm. F Mitochondrial Lon-induced lysosome-mediated autophagy were quantified by LC3B puncta/cell detection according to the immunofluorescence data in E. LC3B puncta/cell was quantified by selecting more than 50 cells per condition. n = 3 biological replicates. The error bars shown in the panel represent the standard deviation from three independent experiments. ***p < 0.001. G, H Mitochondrial Lon induces lysosome-mediated mitophagy. G Mitochondrial Lon-induced lysosome-mediated mitophagy were verified by confocal immunofluorescence. HCT-15 cells transiently expressing mito-QC were transfected with the plasmids encoding Lon or empty and treated with BafilomycinA1 (100 nM for 6 h) or not. The cells were fixed and immunostained by GFP (mitochondria, green), mCherry (mitolysosome, red), and anti-LAMP1 (lysosome, blue) antibodies. Scale bars = 10 μm. H Mitochondrial Lon-induced lysosome-mediated mitophagy were quantified by mcherry signals according to the immunofluorescence data in F. n = 3 biological replicates. The error bars shown in the panel represent the standard deviation from three independent experiments. ***p < 0.001.