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. 2024 Feb 23;43(7):1164–1186. doi: 10.1038/s44318-024-00055-y

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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/. Creative Commons Public Domain Dedication waiver http://creativecommons.org/publicdomain/zero/1.0/ applies to the data associated with this article, unless otherwise stated in a credit line to the data, but does not extend to the graphical or creative elements of illustrations, charts, or figures. This waiver removes legal barriers to the re-use and mining of research data. According to standard scholarly practice, it is recommended to provide appropriate citation and attribution whenever technically possible.

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(A) SILAC enrichment distributions, comparing: WT iBMDMs treated with 1 mM CuOOH to untreated (UT) (A I), Ninj1^−/− iBMDMs treated with 1 mM CuOOH to untreated (UT) (A II), and WT iBMDMs treated with 1 mM CuOOH to Ninj1^−/− iBMDMs treated with 1 mM CuOOH (A III). CuOOH treatment for 2 h. Selected DAMPs released in a NINJ1-dependent manner upon ferroptosis induction are indicated (full list in Dataset EV1). (B) Heatmap depicting the distribution of protein sizes detected with a logarithmic enrichment larger than 0.5 in WT and Ninj1^−/− iBMDMs treated with 1 mM CuOOH for 2 h. Bin centers were chosen in 20 kDa increments, ranging from 10 kDa (i.e., 0 to 20 kDa) to 210 kDa (i.e., larger than 200 kDa). (C, D) Cellular Localization (C) and Biological Process (D) GO-term analyses of the NINJ1-dependent (WT iBMDMs) and NINJ1-independent (Ninj1^−/− iBMDMs) ferroptotic secretome from iBMDMs treated with 1 mM CuOOH for 2 h. Bars depict the percentage of proteins that are annotated with a given GO-term among all proteins identified in the sample. Data information: p-values were calculated by Chi-square test, assuming an even distribution. Data are pooled from two independent experiments (A–D), requiring presence of a given protein in both datasets to ensure reliability.