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. 2022 Apr 11;41:136. doi: 10.1186/s13046-022-02346-w

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© The Author(s) 2022

Open AccessThis 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/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 6 — SETD3 regulates the sorting of hY4F into EVs through K264 methylation on YBX1. A Schematic representation of the functional domains of YBX1. B-E Biotin-labelled hY4F RNA pulldown analysis with cell lysate from A549 cells overexpressing FLAG-tagged truncated or mutated YBX1. F Quantitative PCR data of hY4F in EVs from YBX1-KO A549 cells transfected with wild type (WT) or K264A mutant YBX1. G Schematic representation of the pulldown assay using biotin-labelled unmodified or mono/di/tri-methyl-lysine modified YBX1-K264 peptide. H The quantitative mass spectrometry results of peptide pulldown assay shown as heatmap. I Verification of siRNA-mediated knockdown of SETD3 in A549 cells by qPCR. J The effect of siRNA-mediated SETD3 knockdown on hY4F level in EVs and cell lysates from A549 cells transfected with SETD3 siRNA. K Methylation of lysine K264 identified by di-methyl-lysine (Kme2) antibody. L The effect of SETD3 knockdown on YBX1 methylation level was assessed by western blot. M Analysis of YBX1 methylation by SETD3 in vitro using endogenous immunoprecipitation assay using SETD3 antibody in the presence of wild-type or K264A mutant. N QPCR analysis of EVs-hY4F levels in A549 co-transfected with YBX1 expression plasmid and siRNA targeting SETD3. Data from three independent experiments are shown as the mean ± SD (error bars). *P < 0.05, **P < 0.01, ***P < 0.001 (Student’s t-test)