EMBO/soda-nel huggingface dataset:
The chromatin remodeler SRCAP promotes self-renewal of intestinal stem cells (2020)
Figure 6

Figure 6. Rest or Prdm16 deletion abrogates the stemness of ISCs.

(A) Representative images of intestines from 8-week-old Rest+/+ and Rest-/- mice (n=6 per group). Length of 120 villi and 120 crypts (20 villi and 20 crypts per mouse) were measured in right panel as means± S.D. ** P < 0.01 by two-tailed Student's t-test. Scale bar, 100 μm. Dotted lines indicate the boundary between crypts and villi. Arrows indicate the length of crypts or villi.

(B) 1×104 ISCs from indicated mice were collected for organoid formation. Scale bar, 200 μm. n=3 for each group. Organoid formation ratios was shown as means ± S.D. ** p<0.01 by two-tailed Student's t-test.

(C) Intestine tissues from 8-week-old Rest+/+ and Rest-/- mice (n=6 per group) were collected for immunohistochemistry staining. Typical images were shown in left panel and statistical ISC numbers from 120 crypts (20 crypts per mouse) were shown in right panel as means± S.D. ** P < 0.01 by two-tailed Student's t-test. Scale bar, 50 μm.

(D) Immunofluorescence staining of intestine tissues from Srcapfl/fl; Lgr5GFP-CreERT2 and Restfl/fl; Lgr5GFP-CreERT2 mice. Scale bar, 50 μm.

(E) Lgr5GFP-CreERT2;Rosa26lsl-lacZ (LRlacZ) were crossed with Rest-/- mice, followed by TAM administration for lineage tracing for intestinal whole-mount staining for β-gal (upper panel) . Mice were sacrificed after one week and typical sections were shown (middle panel). Scale bar, 200 μm. Numbers of traced crypt-villus units were calculated as means± S.D. ** P < 0.01 by two-tailed Student's t-test in right panel. 50 fields (10 fields per mouse) were calculated for each group (n=5) (lower panel).

(F) Rest+/+ and Rest-/- mice were treated with 10 Gy's radiation and sacrificed at the indicated time points. Intestinal tissues were collected for H&E staining. Scale bars, 100 μm. Numbers of intact crypts were shown in lower panel as means± S.D. ** P < 0.01 by two-tailed Student's t-test. 50 fields (10 fields per mouse) were taken for each group (n=5).

(G) Representative images of organoid formation from indicated ISCs were shown (upper panel). Scale bar, 200 μm. n=6 for each group. Organoid number per well was counted as means ± S.D. (lower panel). Lentivirus-carried sgPrdm16 and sgPpard were prepared in 293T cells and infected into ISCs derived from TAM-treated LRCas9 mice to obtain Prdm16-/- and Ppard-/- ISCs. 1×104 ISCs from indicated mice were collected for organoid formation. Scale bar, 200 μm. n=5 for each group.

Data information: All data are representative of five independent experiments.
Jag2b-Notch3/1b-mediated neuron-to-glia crosstalk controls retinal gliogenesis (2022)
Figure 1

Figure 1. Transient Notch activation during MG maturation

A Schematic showing Notch dynamics in MG generation. We photoconverted a mother progenitor cell (RPC, magenta in nuclei and GFP indicated notch activation) at 48 hpf using the 405-nm laser and examined the Notch activity within clones at 3 dpf.

B, C Representative images of mother RPC (B) and its final clone (C). Notch activity indicated by mNeonGreenCAAX (green); the area of retina is labeled with solid line, and each layer structure is labeled with dashed lines and marked with outer nuclear layer (ONL), inner nuclear layer (INL), inner plexiform layer (IPL), and ganglion cell layer (GCL). Scale Bars: 10 μm.

D Working model describing the dynamic of Notch activity during MG generation at lineage levels. MG: Müller glial cells; BP: bipolar cells; Notchhigh represents high Notch activity (green); Notchabsent represents no Notch activity.

E Representative images of Notch activity indicated by Tg(Tp1:mVenus)( green) and matured MG stained with GS (magenta) at different stages. Scale Bars: 20 μm.

F Quantitative plots showing the numbers of Tp1+ (left), GS+ (right) cells in the developing retinas at different stages. We collected retinas from 60 hpf (n = 14), 3 dpf (n = 12), 5 dpf (n = 12), and 7 dpf (n = 12, Mean ± SEM; * p<0.05, ***p<0.001, t-test).

G qPCR analysis of mVenus (left) and glula (GS, right) RNA expression in retinas of developing Tg(blbp:dsred) x Tg(Tp1:mVenus) fish (n = 3, Mean ± SEM; * p<0.05, ** p<0.01, t-test).
Jag2b-Notch3/1b-mediated neuron-to-glia crosstalk controls retinal gliogenesis (2022)
Figure 2

Figure 2. Genetic mutants confirms the effects of different Notch ligands on MG generation

A Representative images of Notch activation labeled by Tg(Tp1:mVenus)(green) and mature MGs stained with glutamate synthase (GS, magenta) at 4 dpf in Notch ligands (dla-/-/dlb-/-/dlc-/-/dld-/-/jag1b-/-/jag2b-/-) mutants. Scale Bars: 20 μm.

B, C Quantitative plots showing the numbers of Notch activation (Tp1+) cells (B) and mature MG (GS+) cells (C) in A at 4 dpf. In total, we collected 11 fish for wild-type fish; 8 fish for jag2b-/- mutants; 15 fish for dla-/- mutants; 19 fish for dld-/- mutants; 19 fish for dlc-/- mutants, 11 fish for dlb-/- mutants, 11 fish for jag1b-/- mutants (mean ± SEM; ***p<0.001, t-test).

D Summary table showing the premature stop codon and unknown polypeptides sides of each mutant.
Jag2b-Notch3/1b-mediated neuron-to-glia crosstalk controls retinal gliogenesis (2022)
Figure 4

Figure 4. Crx promoter-driven jag2b rescues MG loss in jab2b-/- mutants

A Upper: representative images showing that GS+ MGs (magenta) were rarely observed in 5-dpf jag2b-/- mutants expressing H2B-CFP (green) driven by the crx promoter. Bottom: representative images showing that GS+ MGs (magenta) were frequently rescued underneath PRs when jag2b expression (green) was driven by the crx promoter in 5-dpf jag2b-/- mutants. The area of retina is labeled with solid line, and each layer structure is labeled with dashed lines and marked with outer nuclear layer (ONL), inner nuclear layer (INL), inner plexiform layer (IPL), and ganglion cell layer (GCL). Scale bars: 20μm. The remaining MGs are indicated by white arrowheads, and the rescued MGs are indicated by yellow arrows.

B Quantification of GS+ MGs (magenta in A) with (green) or without (gray) CFP expression driven by the crx promoter. Ctrl indicates H2B-CFP expression driven by the crx promoter (n = 16 larvae); rescue indicates jag2b-H2B-CFP expression driven by the crx promoter (n = 21 larvae, mean ± SEM; * p<0.05, ns, no significant difference, pair-wised t-test).

C The proportions of jag2b-misexpressing PRs (Green) with underneath MGs (GS+, magenta in A) in 5-dpf jag2b-/- mutants and all MGs in retina. In total, we analyzed retinae of 16 control (Ctrl, jag2b-/- , crx::h2b-ecfp expression) and 21 rescue (jag2b-/-, crx::jag2b expression) larvae (Mean ± SEM; ***p<0.001, t-test).
Jag2b-Notch3/1b-mediated neuron-to-glia crosstalk controls retinal gliogenesis (2022)
Figure 5

Figure 5. Differentiating MGs were re-specified into bipolar neurons in jab2b-/- mutants

A Schematic showing the labeling and tracing of single differentiating MG using Tg(Tp1:gal4::uas:kaede). We photoconverted a differentiating MG exhibiting Müller-like morphology at 60 hpf (magenta) with the 405-nm laser and took clones at 4 dpf.

B Representative images showing the progeny of Müller-like Tp1+ differentiating MG precursors, which became MG in the wild type (jag2b+/+, n = 45), but BP in the jag2b-/- mutants (n = 63). The area of retina is labeled with solid line, and each layer structure is labeled with dashed lines and marked with outer nuclear layer (ONL), inner nuclear layer (INL), inner plexiform layer (IPL), and ganglion cell layer (GCL). Scale Bars: 10 μm.

C Pie plot (upper) and bar plot (below) showing the proportions of MGs and BPs in the wild-type retina (jag2b+/+, n = 6) and jag2b-/- mutants retina (n = 7, mean ± SEM; ***p<0.001, t-test).

D Representative image showing the GS+, but vsx1- expression in the progeny of Tp1+ Müller-like differentiating MGs at wild-type (jag2b+/+). Layer structure indicated in (A). Scale Bars: 20 μm.

E The percentage of the progeny of Müller-like Tp1+ differentiating MG precursors (red+) expression GS (gray), vsx1 (green), or both negative (magenta) in wild-type (jag2b+/+) fish (n = 14, Mean ± SEM; ***p<0.001, t-test).

F Representative image showing vsx1+, but GS- expression in the progeny of Tp1+ Müller-like differentiating MGs in jag2b-/- mutants. Layer structure indicated in (A). Scale Bars: 20 μm.

G The percentage of the progeny of Müller-like Tp1+ differentiating MG precursors (red+) expressing GS (gray), vsx1 (green), or both negative (magenta) in jag2b-/- mutants (n = 29, mean ± SEM; ***p<0.001, t-test).
Jag2b-Notch3/1b-mediated neuron-to-glia crosstalk controls retinal gliogenesis (2022)
Figure 6

Figure 6. Notch1a and Notch3 are required for Jag2b-dependent MGs generation

A Representative images of Notch activation labeled by Tg(Tp1:mVenus)( green) and mature MGs stained with glutamate synthase (GS, magenta) at 4 dpf in Notch receptors (notch1a-/-/notch1b-/-/notch3-/-) mutants. Scale Bars: 20 μm.

B Quantitative plots showing the numbers of cells with active Notch (Tp1+) and mature MG (GS+) cells at 4 dpf. In total, we collected 4 fish for wild-type fish; 7 fish for notch1a-/- mutants; 5 fish for notch1b-/- mutants; 18 fish for notch3-/- mutants (Mean ± SEM; ** p<0.01; ***p<0.001, t-test).

C Representative images showing that GS+ MGs (magenta) were rarely observed when H2B-CFP expression (green) was driven by the gfap promoter in 5-dpf jag2b-/- mutants. The area between the upper dashed and solid lines is the outer nuclear layer (ONL). The area in the middle two dash lines is the inner plexiform layer (IPL). INL, inner nuclear layer; GCL, ganglion cell layer. Scale bars: 20μm.

D Representative images showing that GS+ MGs (magenta) were frequently rescued underneath cells with notch3 expression (green) driven by the gfap promoter in 5-dpf jag2b-/- mutants. Layer structure is similar as (C). Scale bars: 20μm.

E The cell number of GS+ MGs (magenta) with or without CFP expression driven by gfap promoter. Ctrl: H2B-CFP expression (green) driven by the gfap promoter (n = 5 fish); rescue: notch1b (n = 13 fish) and notch3 (n = 17 fish) NICD-H2B-CFP expression (green) driven by the gfap promoter (Mean ± SEM; * p<0.05, ***p<0.001, ns, no significant difference, pair-wised t-test).

F The proportions of notch1b and notch3 NICD-misexpressing cells (green) with underneath MGs (GS+, magenta) in 5-dpf jag2b-/- mutants, indicating that gfap promoter-driven notch1b (n = 13 fish) or notch3 (n = 17 fish) NICD expression could rescue MG loss in jag2b-/- mutants (Mean ± SEM; ***p<0.001, t-test).
Arginine methylation of SIRT7 couples glucose sensing with mitochondria biogenesis (2018)
Figure 1

Figure 1. SIRT7 is methylated at arginine 388

A. Arginine methylase inhibitor decreases SIRT7 methylation. HEK293T cells expressing Flag-tagged SIRT7 were treated with increasing doses of AdOx as indicated for 24 hrs. After immunoprecipitated with Flag-beads, SIRT7 was analyzed by western blot and detected by an antibody against mono-methylarginine (α-me1). Relative methylation ratio (Ratio) was calculated by normalizing arginine methylation level against the Flag-SIRT7 protein. WCL, whole cell lysate.

B. R388 of SIRT7 is evolutionarily conserved. Amino acid sequences corresponding to R388 of human SIRT7 were aligned.

C. PRMT inhibitor reduces arginine methylation of wildtype SIRT7, but not its RK/RF mutants. HEK293T cells expressing wildtype SIRT7 or its RK/RF mutants were treated with or without AdOx for 24 hrs. Arginine methylation of immunopurified SIRT7 was determined by western blot.

D. SIRT7 is mono- and asymmetrically di-methylated at R388. Flag-tagged SIRT7 and its RK/RF mutants were expressed in HEK293T cells. Immunopurified SIRT7 was blotted with antibodies against mono-methylarginine (α-me1), asymmetric di-methylarginine (α-me2a), and symmetric di-methylarginine (α-me2s), respectively.

E. R388 site-specific methylation antibody recognizes R388-methylated peptide, but not the unmodified peptide. The unmodified peptide corresponding to R388, R388-monomethylated (R388me1) peptide, R388-asymmetrically di-methylated (R388me2a) peptide were blotted with α-meSIRT7(R388) antibody.

F. R388-methylated peptide, but not the unmodified peptide, blocks α-meSIRT7(R388) antibody. After incubation with unmodified peptide or R388-methylated peptide (R388me2a), α-meSIRT7(R388) antibody was used to detect immunopurified SIRT7.

G. RK and RF mutations disrupt SIRT7 methylation. Flag-tagged wildtype SIRT7 and its RK/RF mutants were immunoprecipitated and blotted with α-meSIRT7(R388) antibody.

H. SIRT7 is methylated at R388. HEK293T cells ectopically expressing Flag-tagged SIRT7 were treated with increasing concentrations of AdOx for 24 hrs as indicated. SIRT7 protein was pulled down by Flag-beads. Arginine methylation of SIRT7 was detected with a site-specific antibody against R388 methylation [α-meSIRT7(R388)].

I-J. AdOx treatment decreases R388 methylation of SIRT7. MEF and L02 cells were treated with AdOx for 24 hrs. Methylation of immunoprecipitated endogenous SIRT7 was determined by western blot (I). The band intensity of western blots was measured to quantify relative R388 methylation ratio (J) (mean ± SD, n=3 experimental replicates, ***p < 0.001, unpaired two-tailed t-test).

K-L. Quantification of R388-methylated endogenous SIRT7 in HEK293T and L02 cells. R388-methylated endogenous SIRT7 was captured from HEK293T and L02 cells by using R388 site-specific methylation antibody (K). Input, output, and immunoprecipitate were analyzed by western blot to quantify the percentage of R388-methylated protein (L) (mean±SD, n=3 experimental replicates).
Arginine methylation of SIRT7 couples glucose sensing with mitochondria biogenesis (2018)
Figure 2

Figure 2. PRMT6 directly methylates SIRT7 at R388

A. SIRT7 binds to PRMT1, PRMT3, and PRMT6. GFP-tagged PRMTs (PRMT1-9) were expressed in HEK293T cells. PRMT protein was immunoprecipitated with GFP antibody and analyzed by western blot.

B. Co-expression of PRMT6, but not PRMT1 or PRMT3, increases R388 methylation of SIRT7. Flag-tagged SIRT7 and GFP-tagged PRMT1, PRMT3, or PRMT6 were co-expressed in HEK293T cells. R388 methylation of immunopurified SIRT7 was analyzed by western blot.

C. Wildtype PRMT6, but not its catalytic-deficient mutant, upregulates SIRT7 R388 methylation. Wildtype PRMT6 or its V86K/D88A mutant (DM) was co-expressed with Flag-tagged SIRT7 in HEK293T cells. R388 methylation of immunopurified SIRT7 was detected by western blot.

D-E. PRMT6 physically interacts with endogenous SIRT7. Endogenous SIRT7 (D) or PRMT6 (E) was pulled down using corresponding antibody from HEK293T, MEF, and primary mouse hepatocytes, respectively. Input and immunoprecipitates were analyzed by western blot.

F. SIRT7 associates with PRMT6 in vitro. Recombinant GST-PRMT6 were incubated with HEK293T lysates. After GST pulldown, the interaction of recombinant protein with endogenous SIRT7 was analyzed by western blot. The arrows indicate GST and GST-PRMT6 protein, respectively.

G. PRMT6 inhibitor reduces arginine methylation of wildtype SIRT7, but not RK/RF mutants. Wildtype SIRT7 or its RK/RF mutants were stably expressed in MEF cells. After treating cells with or without PRMT6-specific chemical inhibitor (PRMT6i) for 24 hrs, SIRT7 was immunoprecipitated and blotted with R388-specific methylation antibody.

H. Prmt6 depletion decreases R388 methylation of endogenous Sirt7. Scramble control cells (scr) or MEF cells stably expressing two different shRNAs (#1 and #2) against Prmt6 was established. R388 methylation of immunopurified endogenous Sirt7 was determined by western blot.

I. Treatment with PRMT6 inhibitor reduces R388 methylation of endogenous SIRT7 in a dose-dependent manner. MEF and HEK293T cells were treated with increasing doses of PRMT6i for 24 hrs as indicated. R388 methylation of immunoprecipitated endogenous SIRT7 was determined by western blot.

J-K. Prmt6 depletion reduces the percentage of R388-methylated endogenous SIRT7 in MEF cells. R388-methylated endogenous SIRT7 was captured from scramble control cells (scr) and Prmt6-knockdown MEF cells by using R388 site-specific methylation antibody (J). Input, output, and immunoprecipitate were analyzed by western blot to quantify the percentage of R388-methylated protein (K) (mean±SD, n=3 experimental replicates).

L. Recombinant PRMT6 directly methylates SIRT7 at R388 in vitro. Bacterially expressed PRMT6 and SIRT7 proteins were incubated with SAM or PRMT6i as indicated. Heat-inactivated PRMT6 was included as negative control R388 methylation of recombinant SIRT7 protein was determined by western blot.
Arginine methylation of SIRT7 couples glucose sensing with mitochondria biogenesis (2018)
Figure 3

Fig 3. R388 methylation of SIRT7 suppresses its H3K18 deacetylase activity

A. RF, but not RK mutant, suppresses H3K18 deacetylase activity of SIRT7. Flag-SIRT7 and its RK/RF mutants were expressed in HEK293T and MEF cells, respectively. Cell lysates were subjected to western blot to detect histone H3K18 acetylation. H3K27ac and H3K56ac were included as negative control. The level of each histone acetylation marker was normalized against histone H3 (Ratio).

B. R388 methylation of SIRT7 decreases its H3K18 deacetylase activity. HEK293T cells expressing wildtype SIRT7 or its RK/RF mutants were treated with or without AdOx for 24 hrs. The SIRT7 enzyme was immunopurified with Flag-beads and further incubated with extracted chromatin in vitro. R388 methylation of immunopurified SIRT7 and H3K18ac of in vitro deacetylated chromatin were determined by western blot.

C. RF, but not RK mutant, shows reduced H3K18 deacetylase activity in vitro. Chromatin was extracted from HEK293T cells and incubated with recombinant GST-tagged SIRT7 (wildtype and RK/RF mutants) in vitro for 2 hrs. H3K18ac was determined by western blot.

D. Wildtype PRMT6, but not its methylase-dead mutant, decreases the H3K18 deacetylase activity of SIRT7. Flag-tagged SIRT7 was co-expressed with wildtype PRMT6 or its catalytic-inactive mutant (DM) in HEK293T cells. The H3K18 deacetylase activity of immunopurified SIRT7 was assayed in vitro.

E. PRMT6 deficiency promotes the H3K18 deacetylase activity of SIRT7. Control sgRNA or two independent sgRNAs (#1 and #2) against PRMT6 was introduced into HEK293T cells. Flag-tagged SIRT7 was ectopically expressed in these cells. The histone deacetylase activity of immunoprecipitated SIRT7 was assayed in vitro using chromatin as a substrate. R388 methylation of immunopurified SIRT7 and H3K18 acetylation level of in vitro deacetylated chromatin was determined by western blot.

F-G. Methylated recombinant SIRT7 shows reduced H3K18 deacetylase activity. In vitro methylated SIRT7 from Fig 2L was subjected to in vitro deacetylation assay. H3K18 acetylation level of in vitro deacetylated chromatin was determined by western blot (F). Relative H3K18ac level was quantified from three independent experiments (G) (mean±SD, n=3 experimental replicates, **p<0.01, n.s.=not significant, unpaired two-tailed t test).
Arginine methylation of SIRT7 couples glucose sensing with mitochondria biogenesis (2018)
Figure 4

Figure 4. R388 methylation of SIRT7 promotes mitochondria biogenesis

A-D Wildtype SIRT7, but not its RF mutant, restores mitochondria mass, ATP level, and cellular respiration in Sirt7-deficient MEF cells. Wildtype SIRT7 or its RF mutant was re-expressed in Sirt7-knockdown MEF cells (A). Mitochondria mass was determined by Mito Tracker Red staining (B). Cellular ATP was quantified and normalized to cell number (C). Respiration flux was determined as indicated (D). (mean ± SD, n=3 experimental replicates, *p<0.05; **p<0.01; n.s.=not significant, unpaired two-tailed t test).

E-F. Wildtype SIRT7, but not its RF mutant, downregulates SIRT7 target genes involved in mitochondria biogenesis. Wildtype SIRT7 and its RF mutant were stably expressed in MEF cells (E). SIRT7 target gene expression was determined and presented as a heatmap on a log2 scale (F).

G-J Inhibition of Prmt6 decreases mitochondria mass, ATP level and the respiration flux of control cells, but not Sirt7-knockdown MEF cells. Control cells and stable Sirt7-knockdown MEF cells were treated with or without 5 µM PRMT6i for 24 hrs. The H3K18ac level at the promoter of SIRT7 target genes (Gfm2, Mrps31, Mrps33, and Mrpl40) was determined by ChIP-qPCR, Rabbit IgG was used as a negative control (G). Mitochondria mass was determined by Mito Tracker Red staining (H). Cellular ATP was quantified and normalized to cell number (I). Respiration flux was determined as indicated (J). ( Data shown are mean±SD, n=3 experimental replicates, *p<0.05; **p<0.01; ***p < 0.001; n.s.=not significant, unpaired two-tailed t test)
Arginine methylation of SIRT7 couples glucose sensing with mitochondria biogenesis (2018)
Figure 6

Figure 6. SIRT7 methylation connects glucose sensing with mitochondria biogenesis

A-B Glucose administration leads to R388 hypermethylation of Sirt7 in mouse hepatocyte. After fasting for 16 hrs, mice were injected with glucose. Hepatic tissue was harvested in a time-dependent manner as indicated (n=3 for each time point). SIRT7 protein was immunoprecipitated from whole liver extracts. R388 methylation of Sirt7 was determined by western blot (A) and immunohistochemical staining (B). Scale bar in the upper panel, 500 µm; scale bar in the lower panel, 100 µm.

C-F AICAR and PRMT6 inhibitor decrease R388 methylation and epigenetically downregulate SIRT7-target genes. MEF cells stably expressing Flag-SIRT7 were incubated with or without PRMT6i for 12 hrs, followed by treatment with 1 mM AICAR for 12 hrs. R388 methylation of SIRT7 was determined by western blot (C). H3K18ac level at SIRT7-target gene promoters were determined by ChIP-qPCR. Normal rabbit IgG was used as a negative control (D). mRNA expression was determined by qPCR analysis (E). Mitochondria mass was determined by Mito Tracker Red staining (F). (Data shown are mean±SD, n=3 experimental replicates, **p<0.01; ***p < 0.001, unpaired two-tailed t test)

G. The model illustrating regulation of SIRT7 by arginine methylation. R388 methylation inhibits the H3K18 deacetylase activity of SIRT7 to maintain mitochondria homeostasis in an AMPK-dependent manner.

Tables and their legends

Table EV1. List of oligonucleotides used in this study

Sequences for oligonucleotides used in this study are listed.

Expanded View Figure legends
HMGB1 coordinates SASP-related chromatin folding and RNA homeostasis on the path to senescence (2021)
Figure 1

Figure 1 Senescence entry by human cells is mostly transcriptionally driven.

Proliferating and senescent IMR90 and HUVECs assayed for β-galactosidase activity (left), cell cycle profiling via FACS (middle) and proliferation via MTT assays (right).

Heatmaps showing changes in gene expression levels upon senescence (log2FC) of genes encoding selected chromatin-associated factors. For each gene shown, statistically significant expression changes were recorded in at least one cell type.

Western blots showing changing protein levels on the path to senescence in IMR90 and HUVECs. Passage 6 cells represent the proliferating state, and passages 21 and 34 the senescent state for HUVECs and IMR90, respectively.

Super-resolution (gSTED) imaging of HMGB1 distribution in proliferating HUVEC nuclei counterstained with DAPI. Bar: 2μm.

Representative immunofluorescence images of IMR90 and HUVECs (left) showing reduced HMGB1 levels in senescent nuclei; bean plots quantify this reduction (right; N is the number of cells analyzed per each condition/cell type). Bars: 5 μm. *: P<0.01; Wilcoxon-Mann-Whitney test.

Scatter plots showing correlation between RNA-seq (transcription) and Ribo-seq (translation; log2) in proliferating and senescent IMR90. Pearson's correlation values (ρ) and the number of genes in each plot (N) are also shown.

As in panel F, but correlating RNA-seq and whole-cell proteome changes.

Heatmap showing GO terms/pathways associated with the gene subgroups from panel G (colour-coded the same way). The number of genes in each subgroup (N) are indicated.