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. 2018 Aug 28;9:3492. doi: 10.1038/s41467-018-05449-1

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© The Author(s) 2018

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. 5 — Phosphorylation of ULK1 influences the STX17 complex. a HeLa cells were starved and the lysates were performed with IP assay via STX17 antibody and analyzed with ULK1 antibody showed in the right top panel. IPed ULK1 is quantified and showed in the right bottom panel. b ULK1 S423D decreased binding to STX17 compared to WT and S423A by IP assay. In addition, IPed STX17 was quantified. c, d GFP-STX17, Flag-LC3, and Myc-ULK1 variants were cotransfected and detected by immunostaining in HeLa cells (n = 20), and the images were quantified (d). Scale bars, 10 μm. e, f ULK1 S423D affects interactions between STX17 and SNAP29 or VAMP8 with cotransfection and IP assay. The results were analyzed with the indicated antibodies. Bound SNAP29 and VAMP8 were quantified. g myc-ULK1 variant transfected HeLa cells were treated with CQ (100 nM) or PBS. IP assay was performed with STX17 antibody or IgG and detected by indicated antibodies. IPed proteins were quantified. h Lysates from the livers of fasting or normal chew mice were performed by IP assay with STX17 antibody and analyzed with ULK1 antibody. All values are means ± SEM of three independent experiments. Student’s t-test (unpaired); *p < 0.05, **p < 0.01, ***p < 0.001. Baf: bafilomycin a1, WCL: whole-cell lysate