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Elsässer et al. reply

Nature volume 548, pages E7–E9 (2017)Cite this article

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replying to G. Wolf et al. Nature 548, 10.1038/nature23277 (2017)

In Elsässer et al.1, we reported that the histone H3 variant H3.3 has a role in the maintenance of interstitial heterochromatin and transcriptional silencing at specific classes of endogenous retroviral elements (ERVs) in mouse embryonic stem (ES) cells. Additionally, we reported an increase in recent intracisternal A-type particle (IAP) ERV insertions in H3.3-knockout ES cells compared to wild-type ES cells (figure 4d of ref. 1). In the accompanying Comment2, Wolf et al. suggest that the majority of elements identified as new insertion events matched polymorphic ERV sites between mouse strains (personal communication with D. L. Mager, see Comment by Wolf et al.2). The H3.3-knockout and wild-type ES cells used in our study1 were generated from a mixed background (129 and C57BL/6) ES cell line.

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Figure 1: PCR analysis of non-annotated IAP insertions.
Figure 2: RNA-seq differential expression analysis using DESeq.

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References

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Author notes

  1. Simon J. Elsässer and Laura A. Banaszynski: These authors contributed equally to this work.

Authors and Affiliations

  1. Division of Translational Medicine and Chemical Biology, Department of Medical Biochemistry and Biophysics, Science for Life Laboratory, Karolinska Institutet, Stockholm, S-171 21, Sweden

    Simon J. Elsässer

  2. Laboratory of Chromatin Biology and Epigenetics, The Rockefeller University, 1230 York Avenue, New York, 10065, New York, USA

    Kyung-Min Noh, Nichole Diaz & C. David Allis

  3. Cecil H. and Ida Green Center for Reproductive Biology Science and Children’s Medical Center Research Institute, UT Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, 75390, Texas, USA

    Laura A. Banaszynski

Authors
  1. Simon J. Elsässer
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  2. Kyung-Min Noh
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  3. Nichole Diaz
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  4. C. David Allis
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  5. Laura A. Banaszynski
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Corresponding authors

Correspondence to Simon J. Elsässer or Laura A. Banaszynski.

Extended data figures and tables

Extended Data Figure 1 H3K9me3 levels after H3.3 depletion.

a, Levels of H3K9me3, H3.1/2, H3.3 and total H3 at IAP, ETn, MusD ERVs and LINE elements in wild-type (control) and H3.3-depleted (KO/KD) cells. H3K9me3 ChIP–seq was done under native conditions, all other ChIPs with formaldehyde (FA) crosslinking. Data are standardized and represented as enrichment over matched input (native and FA-crosslinked, respectively). Antibodies used were H3K9me3 (ab8898, Abcam), H3 (ab1791, Abcam) and H3.1/2 (ABE154, Millipore). b, Enrichment of H3K9me3 in unique flanking sites of IAP, ETn, MusD ERVs and LINE elements in wild-type (control) and H3.3-depleted (KO/KD) cells. Average profiles were aligned and aggregated at the 5′ and 3′ boundaries of 800 annotated elements from standardized unique read count coverage tracks. The profiles are directional with the 5′ ends on the left and 3′ on the right. Compare to extended data figure 4 of ref. 1.

Extended Data Figure 2 Correlation matrix.

a, b, Pearson’s and Spearman’s correlation between all the datasets used in the H3.3-knockout or depletion (a) and Daxx-knockout (b) differential expression analyses.

Extended Data Figure 3 Expression analysis after H3.3 or DAXX depletion.

a, Volcano plot of DESeq analysis of genes and repeats in two H3.3-knockout and two H3.3-knockout/depleted datasets versus two control datasets (GEO accession numbers: GSE59104 and GSE42154). Fold change plotted against −log10(P) value. Genes are plotted in grey, repeats are plotted in black and differentially expressed (P < 0.05) repeat families are shown in red. b, Volcano plot of DESeq analysis of genes and repeats in three pairs of Daxx-knockout and control datasets (GEO accession numbers: GSE73881, GSE70965 and GSE70850). Genes are plotted in grey, repeats are plotted in black and differentially expressed (P < 0.05) repeat families are shown in red.

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Elsässer, S., Noh, KM., Diaz, N. et al. Elsässer et al. reply. Nature 548, E7–E9 (2017). https://doi.org/10.1038/nature23278

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