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. 2020 Mar 2;11:1146. doi: 10.1038/s41467-020-14948-z

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© The Author(s) 2020

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 Mass spectrometry analysis of the potential Flag-STAT1-binding proteins. The number of the identified peptides from the interacting proteins, including HOIP (RNF31), were shown as indicated. CON: control vectors. b Immunoprecipitation (IP) and immunoblotting (IB) analysis of the interaction between Flag-HOIP and Myc-STAT1 in HEK293T cells. c Immunofluorescence analysis of the interaction between Flag-HOIP and Myc-STAT1 in HeLa cells. Scale bar: 1 µm. d Immunoprecipitation analysis of the interaction between endogenous STAT1 and HOIP in mouse primary liver, spleen, and lung cells. e Immunoprecipitation analysis of linear ubiquitination of endogenous STAT1 in HEK293T cells transfected with either control shRNAs (–) or two different shRNAs against HOIP (shHOIP #1, #2). Ub ubiquitination. f Immunoprecipitation analysis of linear ubiquitination of STAT1 in HEK293T cells cotransfected with Flag-HOIL-1L and Flag-Sharpin, together with Flag-HOIP wild-type or its catalytically inactivated mutants (C885A and C916A). g Immunoprecipitation analysis of linear ubiquitination of endogenous STAT1 in HOIP-wild-type (WT) or HOIP-knockout (KO) HEK293T cells. Data are representative of three independent experiments.