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. 2024 Jan 20;15:631. doi: 10.1038/s41467-024-44978-w

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© The Author(s) 2024

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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — a Co-localization of LDs with mCherry-ATG14 in HeLa cells treated with or without 200 μM OA. Mock (17 cells) and OA-treated cells (51 cells) from three independent experiments were analyzed. b Co-localization of LDs with ATG14-mCherry in HeLa cells treated with 200 μM OA. c Co-localization of LDs with mCherry-ATG14 in Vero-E6 cells treated with 200 μM OA. d HeLa cells were treated with OA for 12 h. PNS, cytoplasm, total membrane and LDs fractions were isolated via ultracentrifugation and were analyzed via western blot. e Immunogold electron micrograph of GFP or GFP-ATG14 expressed HeLa cells. Blue arrows mark GFP-ATG14 dots are enriched on the surface of LDs. Red arrows mark GFP-ATG14 dots are enriched on the surface of autophagosomes (AP). f, g Co-localization of LDs with GFP, GFP-ATG14, or GFP-ATG14^WFY-RRR in HeLa cells treated with 200 μM OA, n = 20 cells from three independent experiments were analyzed. h Co-localization of LDs with mCherry-ATG14^△C10aa in HeLa cells treated with 200 μM OA. i HeLa cells were transfected with or without ATG14 siRNA for 48 h and analyzed via western blot. For imaging assay, LDs were labeled with BODIPY-493/503. The nuclei were stained with DAPI. Number of LDs in each cell was counted from 50 cells of three independent experiments. j, k HeLa cells expressing mCherry-ATG14 or its mutant were treated with 200 μM OA. Cells were also treated with 100 μM CQ for 6 h or 400 nM Baf-A1 for 4 h. LDs were labeled with BODIPY-493/503. The nuclei were stained with DAPI. Yellow ROIs indicate cells expressing ATG14. Number of LDs was counted from 50 cells of three independent experiments. l The concentration of triglyceride in HeLa cells of three independent experiments was analyzed. Data information: Scar bar represents 500 nm in e and represents 10 μm for all fluorescence images. Error bars, mean ± SD of three independent experiments. Two-tailed Unpaired Student’s t test. Source data are provided as a Source Data file.