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Reorganization of chromosome architecture in replicative cellular senescence

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Science Advances  05 Feb 2016:
Vol. 2, no. 2, e1500882
DOI: 10.1126/sciadv.1500882
  • Fig. 1 Hi-C interactions matrices.

    (A to C) Normalized heatmaps are shown for the q arm of chromosome 18 in proliferating, quiescent, and senescent cells. The color maps for relative interaction probability are displayed on the same scale for each heatmap. The PC1 signal was used to define the A and B compartments and is displayed below each heatmap (positive PC1 is shown in red and is designated as A compartments, and negative PC1 is shown in blue and is designated as B compartments). (D to F) Differential heatmaps are shown for the q arm of chromosome 18 for the indicated conditions. The color maps are displayed on the same scale for each comparison. Red is used to designate enrichment in the first condition (senescent or quiescent cells), and blue depletion.

  • Fig. 2 Quantitative comparison of short-range versus long-range contacts.

    (A) Contact probability (Ps) was calculated as a function of genomic distance for interactions within individual chromosome arms across the whole genome. (B) The ratio of short-range (≤2 Mb) versus long-range (≥2 Mb) interactions (SVL) was calculated for each chromosome arm across the whole genome. The SVL is significantly higher in senescent cells compared to proliferating or quiescent cells (***P < 0.001).

  • Fig. 3 Switching of TADs between A and B compartments.

    (A) Genome Browser view showing an example of a TAD that switches from an A compartment (red, positive PC1 signal) in proliferating and quiescent cells to a B compartment (blue, negative PC1 signal) in senescent cells. (B) Example of TADs switching from a B compartment in proliferating and quiescent cells to an A compartment in senescent cells. (C) Genome-wide summary of TAD switching using proliferating cells as a reference point. Venn diagrams show the number of TADs switching between proliferating, quiescent, and senescent conditions. Left: A-to-B compartment switches. Right: B-to-A compartment switches. (D) Same as (C) but displaying the number of genes that switch compartments. G1 to G6 designate gene sets containing the genes within each Venn diagram compartment. (E) GSEA analysis of a microarray expression data set from proliferating and senescent cells (33) using our gene set G2 (A to B switch in both quiescent and senescent cells). Significant overrepresentation of genes down-regulated in senescent/quiescent cells is evident [<0.001 false discovery rate (FDR)]. (F) The same analysis with gene set G3 (A to B switch only in senescent cells) showed overrepresentation of genes down-regulated in senescent cells (<0.001 FDR). (G) The same analysis with gene set G6 (B to A switch only in senescent cells) showed a significant overrepresentation of genes up-regulated in senescent cells (0.003 FDR).

  • Fig. 4 Physical distances between individual loci within a single chromosome arm.

    (A) Four DNA-FISH probes (p1 to p4) were designed in the p arm of chromosome 4. Probes p1 and p3 are in nonadjacent A compartments, and p2 and p4 are in nonadjacent B compartments. Probes were chosen on the basis of strong A-A and B-B interactions in Hi-C data. (B) The schematic shows the 3D spatial relationship predicted by Hi-C for probes p2 and p4 (nonadjacent B compartments) and p3 (A compartment). (C) The distances between transcompartment (A-B) probes (p2-p3 and p3-p4) are significantly decreased in senescent cells (***P < 0.001). (D) Representative 3D DNA-FISH images of quiescent (upper panel) and senescent (lower panel) cells.

  • Fig. 5 Assessment of relative chromatin accessibility.

    (A) FAIRE experiments were performed on three separate occasions using cells in the indicated conditions as starting material. Yields of FAIRE extracted (soluble) DNA are shown as picogram per cell. Relative to input DNA, these values represent 12, 10, and 4%, respectively. Error bars show SDs (**P < 0.01; *P < 0.05). (B) DNase I sensitivity of intact nuclei was visualized using the comet assay.

  • Fig. 6 Compaction of chromosomes in senescent cells.

    (A) 3D chromosome painting of chromosomes 4 and 18 in quiescent and senescent cells. 3D renderings of Z stacks of images were generated with Imaris software. Representative nuclei are shown for each condition (quiescent, senescent) for chromosome 4 (top), chromosome 18 (middle), and both visualized in the same cells (bottom). (B) Chromosome volumes were calculated from the 3D renderings, and the resulting distributions of the volumes are shown as box plots. Chromosomes in senescent cells had a significantly smaller volume (***P < 0.001; **P < 0.01). (C) 3D modeling of chromosome 18 based on Hi-C contact probabilities and mean chromosome radii from chromosome painting as scaling factors. The colors designate A (red) and B (blue) compartment signals. (D) In the collapsing spring model, chromosome arms shrink in size as a consequence of an increased local compaction of the chromatin. Increased contact probability in TADs observed in senescent cells is consistent with a model in which the intra-TAD distances decrease more than the inter-TADs distances (d, intra-TAD distance; D, inter-TAD distance; q, quiescent; s, senescent). TADs are depicted here as spheres, shaded either red for A compartments or blue for B compartments.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/2/2/e1500882/DC1

    Table S1. Sequencing and iterative alignment statistics for Hi-C experiments.

    Table S2. Resampling statistics for Hi-C experiments.

    Table S3. Overlap of switching TADs with ENCODE regulatory elements.

    Table S4. Compartment switching gene sets corresponding to Fig. 3D.

    Table S5. GSEA summary statistics for compartment switching gene sets.

    Table S6. Overlap of compartment switching genes with gene sets of senescence-associated phenotypes.

    Table S7. FISH probes used for 3D DNA-FISH experiments.

    Table S8. Summary statistics for 3D DNA-FISH experiments.

    Table S9. Sanger sequencing validation of quiescent and senescent Hi-C libraries.

    Fig. S1. Hi-C interaction matrices for the q arm of chromosome 2.

    Fig. S2. Hi-C interaction matrices for the q arm of chromosome 3.

    Fig. S3. Hi-C interaction matrices for the q arm of chromosome 4.

    Fig. S4. Hi-C interaction matrices for the p arm of chromosome 4.

    Fig. S5. Genomic feature analysis of contact probability.

    Fig. S6. Comparison of first and second Hi-C experiments.

    Fig. S7. Characteristics of TADs and A and B compartments.

    Fig. S8. Representative genes that switch compartments.

    Fig. S9. Physical distances between individual loci within a single chromosome arm.

    Fig. S10. Quantification of comet assay images.

    Fig. S11. Measurement of chromosome arm volumes.

    Fig. S12. Measurement of centromere and telomere volumes in senescent cells.

    Fig. S13. Comparison of Hi-C data between replicative senescence and oncogene-induced senescence.

    Fig. S14. High-resolution comparison of Hi-C data between replicative senescence and oncogene-induced senescence.

    Movie S1. Rotating movie of the 3D Hi-C model for chromosome 18 in quiescent (left structure) and senescent cells (right structure).

    Movie S2. Rotating movie of the 3D Hi-C model for chromosome 4 quiescent (left structure) and senescent cells (right structure).

  • Supplementary Materials

    This PDF file includes:

    • Legends for tables S1 to S9
    • Fig. S1. Hi-C interaction matrices for the q arm of chromosome 2.
    • Fig. S2. Hi-C interaction matrices for the q arm of chromosome 3.
    • Fig. S3. Hi-C interaction matrices for the q arm of chromosome 4.
    • Fig. S4. Hi-C interaction matrices for the p arm of chromosome 4.
    • Fig. S5. Genomic feature analysis of contact probability.
    • Fig. S6. Comparison of first and second Hi-C experiments.
    • Fig. S7. Characteristics of TADs and A and B compartments.
    • Fig. S8. Representative genes that switch compartments.
    • Fig. S9. Physical distances between individual loci within a single chromosome
      arm.
    • Fig. S10. Quantification of comet assay images.
    • Fig. S11. Measurement of chromosome arm volumes.
    • Fig. S12. Measurement of centromere and telomere volumes in senescent cells.
    • Fig. S13. Comparison of Hi-C data between replicative senescence and oncogene-induced senescence.
    • Fig. S14. High-resolution comparison of Hi-C data between replicative
      senescence and oncogene-induced senescence.
    • Legends for movies S1 and S2

    Download PDF

    Other Supplementary Material for this manuscript includes the following:

    • Table S1 (Microsoft Excel format). Sequencing and iterative alignment statistics
      for Hi-C experiments.
    • Table S2 (Microsoft Excel format). Resampling statistics for Hi-C experiments.
    • Table S3 (Microsoft Excel format). Overlap of switching TADs with ENCODE
      regulatory elements.
    • Table S4 (Microsoft Excel format). Compartment switching gene sets
      corresponding to Fig. 3D.
    • Table S5 (Microsoft Excel format). GSEA summary statistics for compartment
      switching gene sets.
    • Table S6 (Microsoft Excel format). Overlap of compartment switching genes with
      gene sets of senescence-associated phenotypes.
    • Table S7 (Microsoft Excel format). FISH probes used for 3D DNA-FISH
      experiments.
    • Table S8 (Microsoft Excel format). Summary statistics for 3D DNA-FISH
      experiments.
    • Table S9 (Microsoft Excel format). Sanger sequencing validation of quiescent and
      senescent Hi-C libraries.
    • Movie S1 (.mp4 format). Rotating movie of the 3D Hi-C model for chromosome
      18 in quiescent (left structure) and senescent cells (right structure).
    • Movie S2 (.mp4 format). Rotating movie of the 3D Hi-C model for chromosome
      4 quiescent (left structure) and senescent cells (right structure).

    Files in this Data Supplement:

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