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. 2019 Mar 7;9:3831. doi: 10.1038/s41598-019-40358-3

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© The Author(s) 2019

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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Iws1 is specifically required for H3K36 trimethylation in mouse preimplantation embryos. (A) The Immunocytochemical analysis shows that H3K36me3 and Setd2 are present during preimplantation development. With the exception of the zygote stage, in which H3K36me3 appears only in the maternal genome, this modification appears on chromatin during all analyzed stages. Setd2 is present at all stages of preimplantation development. (B) Knockdown of Setd2, Iws1 or Supt6 dramatically reduces the level of H3K36me3 compared to that in the siControl group but does not affect the level of H3. DNA was counterstained with DAPI. (C) Iws1 knockdown does not affect H3K36me2 or -me1 at E3.5 (80 hr after siRNA introduction). (D) Immunofluorescence staining shows that Iws1 knockdown does not significantly affect the nuclear level of Setd2. (E) PLA shows that Iws1 is necessary for the interactions of Setd2 with H3K36me2 and Pol II CTD Ser2p. The fluorescence intensities of the PLA signals were dramatically reduced in siIws1 embryos compared to siControl embryos. At least 20 embryos were used for each experiment. Scale bars: 20 μm.