Imaging analysis of six human histone H1 variants reveals universal enrichment of H1.2, H1.3, and H1.5 at the nuclear periphery and nucleolar H1X presence
- Molecular Biology Institute of Barcelona (IBMB-CSIC), Spain
Figures
Figure 1 with 1 supplement
H1 variants are differentially enriched toward the nuclear periphery in T47D cells.
(A) Confocal immunofluorescence of H1 variants (green) and DNA staining (blue). Bottom panels show the intensity profiles of H1 variants and DNA along the arrows depicted in the upper panel. Scale …
Figure 1—figure supplement 1
Endogenous H1 variants immunofluorescence controls and co-localization with heterochromatin protein HP1.
(A) To discard that the peripheral nuclear enrichment seen by immunofluorescence for some of the H1 variants is due to saturating antibody concentrations, we performed H1.3 immunofluorescence using …
Figure 2 with 3 supplements
Super-resolution imaging shows that H1 variants occupy different regions in single cells.
(A) Super-resolution radial fluctuation (SRRF) images of H1 variants (green) and DNA (red). Bottom-left: Insets show the Reference confocal image in each case. Scale bar: 2 µm. (B) Percentage of …
Figure 2—figure supplement 1
H1 variants co-localization at confocal resolution.
(A) Confocal immunofluorescence of the indicated H1 variants (green) with H1.0 (red) and DNA staining (blue). Zoom-in insets highlight H1.0-peripheral enrichment territories. Scale bar: 5 µm. (B) …
Figure 2—figure supplement 2
H1 variants distribution patterns along mitosis phases.
Immunofluorescence of H1 variants and LaminA along the distinct mitotic phases. In all cases, DNA staining is also shown. (A) Replication-independent H1.0 and H1X variants. (B) H1.5 and Lamin A. (C) …
Figure 2—figure supplement 3
H1 variants phosphorylation during mitosis.
(A) Immunofluorescence of H1.2-pT165 and H1.4pT146. A Z-stack maximum projection is shown. Mitotic cells are marked by a red arrow. Scale bar: 10 µm. Immunofluorescence of phosphorylated H1.2-pT165 …
Figure 3 with 1 supplement
H1 variants presence within lamina-associated domains (LADs) and nucleoli.
(A) Confocal (left) and super-resolution (right) images of a T47D cells stained for H1.2, H1.3, or H1.5 (in green) and Lamin A (in red) obtained using SRRF. Full nuclei (upper panel) and zoomed …
Figure 3—figure supplement 1
H1 variants within lamina-associated domains (LADs) and nucleoli under basal and upon ribosomal DNA (rDNA) transcription inhibition.
(A) Representative confocal immunofluorescence images of H1 variants (green), H3K9me2 (red), and DNA (blue). (B) Immunofluorescence of H1X and DNA in T47D H1Xsh −/+ Dox. A Z-stack of five …
Figure 4 with 1 supplement
Chromatin structural changes upon H1 depletion.
(A) Representative super-resolution radial fluctuation (SRRF) images of DNA staining in the different H1 knock-downs (KD) conditions indicated (multi-H1, H1.2, H1.4, and H1X Dox-inducible shRNAs). …
Figure 4—figure supplement 1
Super-resolution imaging of DNA upon different H1 knock-downs (KD) conditions.
(A) Representative images of DNA staining visualized with wide-field or super-resolution (SRRF) microscopy in a T47D nucleus. Bottom panels show a zoom-in of the perinucleolar region. Scale bar: 5 …
Figure 5 with 5 supplements
Nuclear distribution of H1 variants across multiple human cell lines.
(A) Immunofluorescence analysis of H1 variants (green) with DNA staining (blue) in different cancer cell lines. Merged images are shown. H1.3 and H1.0 grids in HeLa cells are empty, as HeLa cells do …
Figure 5—figure supplement 1
H1 protein and mRNA complement across cell lines.
(A) Immunoblot analysis of H1 variants in histones extracts from different tumoral and non-tumoral cell lines. Histone H3 is added as a nuclear control and Coomassie staining is shown. (B) Coomassie …
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Figure 5—figure supplement 1—source data 1
PDF containing original scans of the relevant Western blot analysis shown in Figure 5—figure supplement 1A, C with highlighted bands.
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data1-v1.pdf
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Figure 5—figure supplement 1—source data 2
Original file for the Western blot analysis in Figure 5—figure supplement 1A, left panels (H1X).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data2-v1.jpg
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Figure 5—figure supplement 1—source data 3
Original file for the Western blot analysis in Figure 5—figure supplement 1A, left panels (H1.0).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data3-v1.jpg
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Figure 5—figure supplement 1—source data 4
Original file for the Western blot analysis in Figure 5—figure supplement 1A, left panels (H1.2).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data4-v1.jpg
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Figure 5—figure supplement 1—source data 5
Original file for the Western blot analysis in Figure 5—figure supplement 1A, left panels (H1.3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data5-v1.jpg
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Figure 5—figure supplement 1—source data 6
Original file for the Western blot analysis in Figure 5—figure supplement 1A, left panels (H1.4).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data6-v1.jpg
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Figure 5—figure supplement 1—source data 7
Original file for the Western blot analysis in Figure 5—figure supplement 1A, left panels (H1.5).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data7-v1.jpg
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Figure 5—figure supplement 1—source data 8
Original file for the Western blot analysis in Figure 5—figure supplement 1A, left panels (H3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data8-v1.jpg
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Figure 5—figure supplement 1—source data 9
Original file for the Western blot analysis in Figure 5—figure supplement 1A, right panels (H1X).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data9-v1.jpg
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Figure 5—figure supplement 1—source data 10
Original file for the Western blot analysis in Figure 5—figure supplement 1A, right panels (H1.0).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data10-v1.jpg
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Figure 5—figure supplement 1—source data 11
Original file for the Western blot analysis in Figure 5—figure supplement 1A, right panels (H1.2).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data11-v1.jpg
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Figure 5—figure supplement 1—source data 12
Original file for the Western blot analysis in Figure 5—figure supplement 1A, right panels (H1.3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data12-v1.jpg
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Figure 5—figure supplement 1—source data 13
Original file for the Western blot analysis in Figure 5—figure supplement 1A, right panels (H1.4).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data13-v1.jpg
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Figure 5—figure supplement 1—source data 14
Original file for the Western blot analysis in Figure 5—figure supplement 1A, right panels (H1.5).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data14-v1.jpg
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Figure 5—figure supplement 1—source data 15
Original file for the Western blot analysis in Figure 5—figure supplement 1A, right panels (H3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data15-v1.jpg
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Figure 5—figure supplement 1—source data 16
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H1X).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data16-v1.tif
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Figure 5—figure supplement 1—source data 17
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H1.0).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data17-v1.tif
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Figure 5—figure supplement 1—source data 18
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H1.2).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data18-v1.tif
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Figure 5—figure supplement 1—source data 19
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H1.3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data19-v1.tif
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Figure 5—figure supplement 1—source data 20
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H1.4).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data20-v1.tif
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Figure 5—figure supplement 1—source data 21
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H1.5).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data21-v1.tif
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Figure 5—figure supplement 1—source data 22
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data22-v1.tif
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Figure 5—figure supplement 1—source data 23
Original file for the Western blot analysis in Figure 5—figure supplement 1C, left panels (H4).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data23-v1.tif
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Figure 5—figure supplement 1—source data 24
Original file for the Western blot analysis in Figure 5—figure supplement 1C, right panels (H1X).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data24-v1.tif
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Figure 5—figure supplement 1—source data 25
Original file for the Western blot analysis in Figure 5—figure supplement 1C, right panels (H1.0).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data25-v1.tif
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Figure 5—figure supplement 1—source data 26
Original file for the Western blot analysis in Figure 5—figure supplement 1C right panels (H1.2).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data26-v1.tif
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Figure 5—figure supplement 1—source data 27
Original file for the Western blot analysis in Figure 5—figure supplement 1C, right panels (H1.3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data27-v1.tif
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Figure 5—figure supplement 1—source data 28
Original file for the Western blot analysis in Figure 5—figure supplement 1C, right panels (H1.4).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data28-v1.tif
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Figure 5—figure supplement 1—source data 29
Original file for the Western blot analysis in Figure 5—figure supplement 1C, right panels (H1.5).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data29-v1.tif
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Figure 5—figure supplement 1—source data 30
Original file for the Western blot analysis in Figure 5—figure supplement 1C, right panels (H3).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data30-v1.tif
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Figure 5—figure supplement 1—source data 31
Original file for the Western blot analysis in Figure 5—figure supplement 1C, right panels (H4).
- https://cdn.elifesciences.org/articles/91306/elife-91306-fig5-figsupp1-data31-v1.tif
Figure 5—figure supplement 2
H1 variants expression regulation by DNA methylation.
(A) Scatterplots between H1 variants expression (Y-axis) and H1 gene methylation (X-axis) from NCI-60 public data. Gene expression from total RNA-Seq data is expressed in log2(FPKM + 1) while gene …
Figure 5—figure supplement 3
H1 variants nuclear distribution in non-tumoral cell lines.
Confocal immunofluorescence of H1 variants (green) and DNA staining (blue) in 293T (A) or IMR-90 (B) cells. Bottom panels show the intensity profiles of H1 variants and DNA along the arrows depicted …
Figure 5—figure supplement 4
Nuclear radial distribution of H1.4 and H1.0 across cell lines.
Graphs show immunofluorescence quantification signal of (A) H1.4 and (B) H1.0 in different cell lines. As illustrated in Figure 1B, four sections of an equivalent area and convergent to the nuclear …
Figure 5—figure supplement 5
Cell lines lacking H1.3 and H1.5 show high basal expression of repetitive elements in comparison with cell lines with a complete H1 somatic repertoire.
(A) Reverse-transcriptase-quantitative PCR (RT-qPCR) of interferon stimulated genes (ISGs) and repetitive elements in different melanoma cell lines. Expression data are corrected by GAPDH and …
Figure 6
Summary of H1 variants expression and distribution specificities.
The table summarizes the main aspects of H1 variants expression (blue; top part) and distribution (green/orange; bottom part). Regarding distribution characteristics (from microscopy and ChIP-Seq …
Additional files
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Supplementary file 1
Oligonucleotides for semiquantitative PCR.
Forward (F) and reverse (R) oligonucleotides for the indicated genes are shown.
- https://cdn.elifesciences.org/articles/91306/elife-91306-supp1-v1.docx
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MDAR checklist
- https://cdn.elifesciences.org/articles/91306/elife-91306-mdarchecklist1-v1.docx
