EMBO/soda-nel huggingface dataset:
The ribosomal chaperone NACA recruits PHD2 to cotranslationally modify HIF-{alpha} (2022)
Figure 4

Figure 4. Evidence for disome formation on the Hif2a mRNA.

A,B Ribosome profiling results of sequences corresponding to the (A) proposed HIF-2α and (B) known XBP1u translational pause sites (Chen et al., 2020; Gameiro & Struhl, 2018; Werner et al., 2015), visualized using GWIPS-viz (Michel et al, 2014). Proposed positions of monosome and disome at the pause site, and positions of primers for reverse transcription and Real Time PCR are shown. * = termination codon. Red boxes highlight the (A) proposed and (B) known tripeptide sequences that occupy the E-P-A sites in the ribosome.

C-F Mouse kidney was lysed, treated with or without RNase I, and RNA was isolated. The RNA was reverse transcribed with the indicated primers for (C,E) Hifa and (D,F) Xbp1u mRNA, and then analyzed by Real Time PCR for disomes using the indicated primers. ∆CT = (∆CT = CT-RNase - CT+RNase). Biological replicates (from three males), with mean ± SD are shown. * P < 0.05, ** P < 0.01, by unpaired two-tailed t-test.
ADAM17 selectively activates the IL-6 trans-signaling/ERK MAPK axis in KRAS-addicted lung cancer (2019)
Figure 1

Figure 1. ADAM17 deficiency abrogates oncogenic Kras-induced LAC.

A Representative low (left) and high (right) power images of H&E-stained lung sections from KrasG12D, KrasG12D:Adam17ex/+ and KrasG12D:Adam17ex/ex mice at 6 weeks post Ad-Cre inhalation. Scale bars, 3mm (left) and 300μm (right).

B, C Quantification of lung parenchyma area occupied by tumor lesions (B), and tumor incidence (C), per whole mouse lung in the indicated genotypes (n = 6 per genotype). *P<0.05, ***P<0.001, Student's t-test.

D Representative images of TTF-1-stained lung sections from the indicated genotypes. Scale bar, 100μm.

E Quantification of TTF-1-positive cells/high power field (HPF) in lungs of the indicated genotypes (n = 6 per genotype). *P<0.05, **P<0.01, Student's t-test.

Exact P values are specified in Appendix Table S4.
ADAM17 selectively activates the IL-6 trans-signaling/ERK MAPK axis in KRAS-addicted lung cancer (2019)
Figure 2

Figure 2. Reduced tumor cell proliferation and inflammation in KrasG12D:Adam17ex/ex mouse lungs.

A Representative images of PCNA-stained lung sections from KrasWT, KrasG12D and KrasG12D:Adam17ex/ex mice at 6 weeks post Ad-Cre (for KrasG12D) or vehicle (for KrasWT) inhalation. Scale bar, 100μm.

B Quantification of PCNA-positive cells/high power field (HPF) in the indicated mouse lungs (n = 6 per genotype). **P<0.01, Student's t-test.

C qPCR expression analyses of cell cycle regulation genes (normalized against 18SrRNA) in lungs from the indicated mice (n = 6 per genotype). *P<0.05, Student's t-test.

D Representative immunoblots of KrasG12D and KrasG12D:Adam17ex/ex lung lysates with the indicated antibodies. Each lane is an individual mouse sample.

E Semi-quantitative densitometry of Myc protein levels (relative to Actin) in lung lysates from (D). n = 6 per genotype. **P<0.01, Student's t-test.

F, G Representative images (F) and quantification of CD45-positive cells/HPF (G) in CD45-stained lung sections from the indicated genotypes at 6 weeks post inhalations. In (F), scale bar, 100μm. In (G), n = 6 per genotype. **P<0.01, Student's t-test.

Exact P values are specified in Appendix Table S4.
ADAM17 selectively activates the IL-6 trans-signaling/ERK MAPK axis in KRAS-addicted lung cancer (2019)
Figure 3

Figure 3. Cellular expression pattern of ADAM17 in lungs of tumor-bearing mice.

A-E Representative immunofluorescence images of lung sections of KrasG12D mice (6 weeks post Ad-Cre inhalation) co-stained for ADAM17 and markers for club cells (CC10) (A), alveolar type II (surfactant protein-C, SP-C) (B) and type I (Podoplanin, Pdpn) (C) cells, total immune cells (CD45) (D) and endothelial cells (CD31) (E). DAPI nuclear staining is blue. Scale bars, 100μm. Arrowheads in merged images indicate dual-positive ADAM17-expressing cells.

F Quantification of immunofluorescence staining from A-E (n = 5 mice per stain).

G Representative images of H&E-stained lung sections from KrasG12D recipient mice reconstituted with KrasG12D (KrasKras) or KrasG12D:Adam17ex/ex bone marrow (KrasKras:ex/ex). Scale bar, 3mm.

H Quantification of lung parenchyma area occupied by tumor lesions in the indicated bone marrow chimeras (n = 6 mice per group).

Exact P values are specified in Appendix Table S4.
ADAM17 selectively activates the IL-6 trans-signaling/ERK MAPK axis in KRAS-addicted lung cancer (2019)
Figure 4

Figure 4. Lung tumorigenesis in KrasG12D mice is associated with upstream modulation of ADAM17 Thr735 phosphorylation by p38 MAPK, and downstream ADAM17-mediated ERK1/2 MAPK signaling.

A Representative images of pADAM17-stained lung sections from KrasWT and KrasG12D (6 weeks post inhalation) mice. Scale bar, 100μm.

B Quantification of pADAM17-positive cells/high power field (HPF) in lungs of KrasWT and KrasG12D mice (n = 6 per genotype). **P<0.01, Student's t-test.

C Representative immunoblots of individual KrasWT and KrasG12D mouse lung lysates with the indicated antibodies.

D Semi-quantitative densitometry of pADAM17 in lung lysates (n = 6 per genotype). *P<0.05, Student's t-test.

E Representative immunoblots of individual KrasWT and KrasG12D lung lysates with the indicated antibodies.

F Semi-quantitative densitometry of pp38 and pERK1/2 MAPK protein levels in KrasWT and KrasG12D lung lysates (n = 6 per genotype). **P<0.01, Student's t-test.

G Representative images of pADAM17-stained lung sections from KrasG12D mice injected with a single dose of vehicle (control), SB203580 p38 MAPK or U0126 ERK1/2 inhibitors. Scale bar, 100μm.

H Quantification of pADAM17-positive cells/HPF in lungs of the indicated groups (n = 5 per group). **P<0.01, Student's t-test.

I Immunoblots with the indicated antibodies on lysates from A549 and NIH-H23 LAC cell lines treated with SB203580 for 24 hours.

J, K MTT viability (J) and ATP proliferation (K) assays of A549 and NCI-H23 LAC cells treated with SB203580. Data are from 3 independent experiments performed in triplicate. *P<0.05, **P<0.01, Student's t-test.

L Representative immunoblots of individual KrasG12D and KrasG12D:Adam17ex/ex lung lysates with the indicated antibodies. For the Actin blot, lanes 1-3 are reproduced in lanes 4-6 of the Actin blot in Appendix Fig S2B. For the pp38/p38 MAPK blots, lanes 1-3 are reproduced in lanes 4-6 of the corresponding blots in panel E.

M Representative images of pERK1/2-stained lung sections from the indicated genotypes. Scale bars, 100μm.

N Quantification of pERK1/2-positive cells/high power field (HPF) in lungs of the indicated genotypes (n = 6 per genotype). **P<0.01, Student's t-test.

Exact P values are specified in Appendix Table S4.
ADAM17 selectively activates the IL-6 trans-signaling/ERK MAPK axis in KRAS-addicted lung cancer (2019)
Figure 5

Figure 5. In KrasG12D-induced LAC, ADAM17-mediated processing of sIL-6R is selectively impaired, and specific targeting of sIL-6R reduces ERK MAPK activation.

A ELISA of sIL-6R and TNFα protein levels in serum from KrasG12D and KrasG12D:Adam17ex/ex mice at 6 weeks post Ad-Cre inhalation, along with control KrasWT mice at 6 weeks post vehicle inhalation (n = 6 per genotype). *P<0.05, **P<0.01, Student's t-test.

B Immunoblots of representative individual KrasWT, KrasG12D and KrasG12D:Adam17ex/ex lung lysates with the indicated antibodies.

C Representative immunofluorescence staining of KrasG12D mouse lung sections (n = 5 mice) for ADAM17 and either (panels from left to right) IL-6R, TGFα, Notch1, Nrg1 and EGF. Sections were counterstained with DAPI (blue). Scale bars, 100μm.

D Representative images of pERK1/2-stained lung sections from KrasG12D mice treated with either vehicle (control, Ctl) or anti-IL-6R antibodies 1F7 or 25F10 over 6 weeks post Ad-Cre inhalation. Scale bar, 100μm.

E Quantification of pERK1/2-positive cells/high power field (HPF) in lungs of the indicated groups (n = 5 per group). *P<0.05, Student's t-test.

F Representative immunoblots with the indicated antibodies on whole cell lysates from primary immune and epithelial cells isolated from the lungs of mice (n = 3) harbouring the oncogenic KrasG12D allele that were stimulated with either PBS (-) or 100ng/ml Hyper-IL-6 (+). Graphs depict semi-quantitative densitometry of pERK1/2 MAPK protein levels in stimulated immune and epithelial cells (n = 2 experiments).

Exact P values are specified in Appendix Table S4.
ADAM17 selectively activates the IL-6 trans-signaling/ERK MAPK axis in KRAS-addicted lung cancer (2019)
Figure 6

Figure 6. Enhanced activation of the ADAM17-sIL-6R-ERK1/2 MAPK axis promotes tumor growth in human KRAS mutant LAC.

A Representative images of pADAM17-stained lung sections from normal (cancer-free) control and LAC patients. Scale bar, 100μm.

B, C Quantification of pADAM17-positive cells (B and C) and pERK1/2 MAPK-positive cells (C) in (B) normal (cancer-free) control individuals (n = 8) and LAC patients (n = 18), and (C) LAC patients either KRAS wild-type (WT, n = 13) or mutant (MUT, n = 5). *P<0.05, **P<0.01, Student's t-test.

D, E Linear regression analyses of pADAM17 with pERK1/2 (D) and pp38 (E) MAPK staining in serial lung sections from LAC KRAS wild-type and mutant patients. r, Pearson correlation coefficient.

F Immunoblots of lysates from normal human bronchial epithelial cells (HBEC) and LAC cell lines A549, NIH-H23 and NIH-H2228 with the indicated antibodies. Shown are lysates from 2 separate passages per cell line. Pro, proADAM17; Mat, mature ADAM17.

G Densitometric semi-quantification of immunoblots from (F) showing the relative expression levels of pADAM17 (relative to Actin), pro and mature forms of ADAM17 (relative to Actin), and pERK1/2 and pp38 MAPK (relative to their total counterparts) protein levels.

H ELISA of sIL-6R protein levels in culture supernatants from matching cells in (F) cultured over 24 hours. Data are from 4 independent experiments. **P<0.01, Student's t-test.

I, J MTT viability (I) and ATP proliferation (J) assays of A549 cells transduced with nontargeted control sgRNA (Ctl) and ADAM17 sgRNA (knockout, KO). Data are from 3 independent experiments performed in triplicate. *P<0.05, **P<0.01, Student's t-test.

K ELISA of sIL-6R protein levels in culture supernatants from matching A549 cells (I and J) cultured over 24 hours. Data are from 3 independent experiments. ***P<0.001, Student's t-test.

L Tumor volume (mm3) of KO and Ctl A549 xenografts assessed every 3 days starting at one week post injection of 106 cells into NSG mice (n = 5 per group). *P<0.05, Student's t-test.

M, N Weight (grams) (M) and representative images (N) of tumor xenografts from (L). Scale bar, 1cm. In (M), n = 5 per group. **P<0.01, Student's t-test.

O ELISA of human sIL-6R protein levels in serum of the indicated mice (n = 5 per group). Levels of sIL-6R are below detection in serum from KO xenograft mice. ND, not detected.

P Representative images of pADAM17 and pERK1/2 MAPK immunohistochemical staining on KO and Ctl A549 xenograft tumors. Scale bars, 100μm.

Q Immunoblots of lysates from individual A549 KO and Ctl xenograft tumors with the indicated antibodies.

Exact P values are specified in Appendix Table S4.
ADAM17 selectively activates the IL-6 trans-signaling/ERK MAPK axis in KRAS-addicted lung cancer (2019)
Figure 7

Figure 7. Anti-cancer activity of the specific ADAM17 prodomain inhibitor, A17pro, in preclinical genetically engineered and PDX KRAS mutant LAC models.

A Representative images of H&E-stained lung sections from KrasG12D mice treated with either vehicle (control) or the ADAM17 prodomain inhibitor, A17pro (1mg/kg, 3 times a week), for 6 weeks starting on the day after Cre inhalation. Scale bars, 3mm.

B Quantification of lung parenchyma area occupied by tumor lesions in the indicated groups (n = 6 per group) from (A). **P<0.01, Student's t-test.

C, E, G Representative images of lung sections from the indicated mice from (A) stained with antibodies against TTF-1 (C), PCNA (E) and pERK1/2 (G). Scale bars, 100μm.

D, F, H Quantification of positive cells/high power field (HPF) in the lungs of mice from (C), (E) and (G) stained for TTF-1 (D), PCNA (F) and pERK1/2 (H). n = 6 per group. *P<0.05, **P<0.01, Student's t-test.

I ELISA of serum sIL-6R protein levels in the indicated mice (n = 6 per group) from (A). *P<0.05, Student's t-test.

J Tumor volume (mm3) of a KRAS mutant PDX treated with vehicle control (Ctl) or A17pro (1mg/kg) every second day. Treatment started when tumors reached a volume of 100-150mm3 after injection of 2 × 106 cells in NSG mice. n = 5 per group. *P<0.05, Student's t-test.

K, L Weight (grams) (K) and representative images (L) of tumor PDXs from (J). In (K), n = 5 per group. *P<0.05, Student's t-test. Scale bar, 1cm.

M ELISA of human sIL-6R protein levels in serum of the indicated mice (n = 5 per group). *P<0.05, Student's t-test.

N Immunoblots with the indicated antibodies of representative lysates from individual PDX tumors treated with vehicle control or A17pro.

O Representative image of pADAM17 immunohistochemical staining on an untreated KRAS mutant PDX tumor. Scale bar, 100μm.

Exact P values are specified in Appendix Table S4.
Listeriolysin O allows Listeria monocytogenes replication in macrophage vacuoles (2007)
figf1

a, Mice were infected for 21 days and liver granulomas analysed by transmission electron microscopy (TEM). Shown are spacious vacuoles (SLAPs) containing multiple bacteria. Magnification, ×5,200. Region 1 and 2 (white boxes) are enlarged in the lower panels. b, SCID mice were infected as in a and liver sections stained for LAMP1 (green), bacteria (red) and DNA (blue). Shown is a LAMP1+ SLAP. c, The percentage of bacteria in LAMP1+ compartments was quantified and characterized as either spacious or tight-fitting. Mean ± s.e.m. for three mice examined. The image in a (from ref. 2) and the tissue sections were provided by E. Unanue.
Listeriolysin O allows Listeria monocytogenes replication in macrophage vacuoles (2007)
figf2

a, RAW 264.7 macrophages infected for 4 or 8 h were analysed by TEM. Shown are spacious vacuoles (SLAPs) containing multiple bacteria. The arrow indicates septum of dividing bacteria. Scale bars, 0.5 &mgr;m. b, The arrow indicates a LAMP1+ SLAP colocalizing with green fluorescent protein (GFP)–LC3 in cells infected for 4 h. d, The arrowhead indicates a LAMP1+ SLAP devoid of cathepsin D in cells infected for 4 h. Scale bars, 5 &mgr;m. c, e, The percentage of GFP–LC3+ (c) or cathepsin D+ (e) SLAPs was quantified, and compared to GFP–LC3 or cathepsin D colocalization with cytosolic (Cyto.) bacteria (actin+ or LAMP1–) or PFA-killed bacteria in phagosomes (LAMP1+). Mean ± s.e.m. for three independent experiments. P values for conditions significantly different from PFA-killed bacteria are shown. f, GFP–LC3-transfected macrophages were infected with wild-type or &Dgr;actA bacteria. Where indicated, chloramphenicol (CM) was added to the media at 3 h post infection. The number of bacteria per SLAP was quantified. Brackets indicate significant differences, and corresponding P values are shown. g, Macrophages were infected with wild-type bacteria for 7 h, pulsed with BrdU for 1 h, and stained for LAMP1 (red), bacteria (blue) and BrdU (green). Magnified images and the arrow indicate SLAPs containing actively replicating bacteria (BrdU+). h, The percentage of BrdU+ bacteria in SLAPs, compared to cytosolic and PFA-killed bacteria, was quantified as in c.
Listeriolysin O allows Listeria monocytogenes replication in macrophage vacuoles (2007)
figf3

a, GFP–LC3-transfected macrophages were infected for 4 h, and the percentage of infected cells exhibiting SLAPs was quantified. Mean ± s.e.m. for three independent experiments. P values for strains with significant differences from wild-type levels are shown. b, The arrow indicates a GFP–LC3+ SLAP with internal LLO expression in cells infected for 4 h. Scale bar, 5 &mgr;m. c, Arrowheads indicate GFP–LC3+ SLAPs devoid of Lysotracker Red in cells infected for 4 h. d, The pH of FITC-labelled bacteria in SLAPs, PFA-killed bacteria or zymosan particles was determined by ratiometric imaging. P values compared to bacteria in SLAPs are shown. e, The arrow indicates a LAMP1+ SLAP colocalizing with v-ATPase staining in cells infected for 4 h. Scale bar, 5 &mgr;m. f, The percentage of v-ATPase+ SLAPs was quantified as in Fig. 2c. Mean ± s.e.m. for three independent experiments. P values for conditions significantly different from PFA-killed bacteria are shown.
Listeriolysin O allows Listeria monocytogenes replication in macrophage vacuoles (2007)
figf4

a, Arrows indicate LAMP1– wild-type bacteria with actin ‘comet tails’ in cells infected for 6 h. The boxed region is magnified in the bottom left panels. The merged image for this region is shown at the bottom right. Scale bar, 5 &mgr;m. b, Macrophages were infected with IPTG-induced iLLO bacteria. Arrowheads indicate actin– iLLO bacteria within LAMP1+ vacuoles. c, Macrophages were infected with wild-type (triangles) or IPTG-induced iLLO (circles) bacteria, and the percentage of LAMP1+ bacteria was quantified. Mean ± s.e.m. for three independent experiments. d, Macrophages were infected as in c with or without IPTG induction. Intracellular bacterial replication was determined using a gentamicin-protection assay. Shown is fold replication compared to 2 h post infection. Clear triangles, wild type; filled triangles, wild type + IPTG; clear circles, iLLO; filled circles, iLLO + IPTG. Mean ± s.e.m. or range for three (wild type, wild type + IPTG, iLLO + IPTG) or two (iLLO–IPTG) independent experiments, respectively. e, Wild-type or Atg5-/- MEFs were infected with iLLO bacteria, and intracellular bacterial replication was determined as in d. Mean ± s.e.m. for three independent experiments. Clear circles, wild-type MEFs; filled circles, wild-type MEFs + IPTG; clear squares, Atg5–/– MEFs; filled squares, Atg5–/– MEFs + IPTG. f, Wild-type (circles) or Atg5-/- (squares) MEFs were infected as in e with IPTG induction. The percentage of LAMP1+ bacteria was quantified as in c. Mean ± s.e.m. for three independent experiments. g, Model of the different fates of L. monocytogenes in host cells. High LLO activity allows bacterial escape from phagosomes. Under conditions where LLO activity is not sufficient to drive escape (low LLO), autophagy maintains bacteria within non-degradative vacuoles (SLAPs) that allow slow bacterial growth. Bacteria can also be degraded in phagolysosomes.
Telomeric epigenetic response mediated by Gadd45a regulates stem cell aging and lifespan (2018)
Figure 1

Figure 1. Gadd45a deletion improves organ maintenance and prolongs the lifespan of G3Terc-/- mice.

A,B Western blotting analysis of Gadd45a protein expression in mouse intestine crypt lysates from WT and G3Terc-/- mice (A). Quantification of Gadd45a protein expression, β-actin protein level was used as the loading control (n=7 mice per genotype), two independent experiments were carried out (B).

C Quantification of Gadd45a-ChIP values as percentage of total telomeric DNA and ALU elements in WT and G3Terc-/- MEF cells (n=3 for each genotype).

D Dot-bot assay following Gadd45a-ChIP in WT and G3Terc-/- MEF cells.

E Kaplan-Meier survival curves of the mice with indicated genotypes (WT, n=44; Gadd45a-/-, n=48; G3Terc-/-, n=37; G3-dKO, n=41). P value comparing the survival of G3Terc-/- with G3-dKO mice is calculated with the log-rank test.

F The average body weight of mice with indicated genotypes at indicated time points (G3-dKO Female (n=19) vs G3Terc-/- Female (n=17), P=0.004; G3-dKO Male (n=14) vs G3Terc-/- Male (n=14), P=0.0038). P values are calculated with log-rank test.

G,H Whole mount images of colon samples from 7-month old mice with indicated genotypes (G). Quantification on density of colon crypts (H) (n=5 mice per group).

I,J H&E-stained small intestine sections from 7-month old mice with indicated genotypes (I). Quantification of the number of small intestine basal crypts (J) (n=5-6 mice per group).

Data information: Scale bars represent 200 μm for Fig. 1G and 50 μm for Fig. 1I. Data are presented as mean±s.e.m.
Telomeric epigenetic response mediated by Gadd45a regulates stem cell aging and lifespan (2018)
Figure 2

Figure 2. Gadd45a deletion improves intestinal stem cell maintenance and function in G3Terc-/- mice.

A,B Olfm4-positive intestinal stem cells (indicated by arrows) in small intestine sections (A). Quantification of intestinal stem cell (Olfm4+) number per 100 μm (n=4-5 mice per group) of small intestine sections (B).

C,D Representative images of small intestine crypt culture (day 2). Arrows mark the surviving cysts (C). Quantification of numbers of cysts per culture well (D)(n=4-5 mice/genotype).

E,F In vitro crypt culture (day 8) from small intestine crypts isolated from mice with indicated genotypes. Arrows mark the crypt-like structure (E). The number of organoids per culture wells (n=4-5 mice per group). Two hundred crypts are seeded per well (F).

G,H Representative images of TUNEL staining in small intestine sections. Arrows indicate TUNEL positive cells (G). Quantification of TUNEL positive cells per crypt (n=5 mice per group) (H).

I,J Representative images of phosphorylated histone 3 (pH3) antibody staining in mouse small intestine. Arrows indicate pH3 positive cells (I). Quantification of pH3 positive crypt cells per 100 μm of small intestines (n=4-6 mice per group) (J).

Data information: Scale bars represent 50 μm for Fig. 2A, 200 μm for Fig. 2C, 25 μm for Fig. 2G, 60 μm for Fig. 2E and 100 μm for Fig. 2I. Data are presented as mean±s.e.m.
Telomeric epigenetic response mediated by Gadd45a regulates stem cell aging and lifespan (2018)
Figure 3

Figure 3. Gadd45a deletion attenuates telomere-dysfunction induced DDR in the basal crypts of G3Terc-/- mice.

A,B Representative images of immunostaining for p53 on small intestine sections. Arrows indicate p53-positive cells (A). Quantification of the percentage of p53 positive cells per crypt (n=5-6 mice per group) (B).

C,D Representative images of γH2AX antibody staining on small intestine paraffin sections from WT, Gadd45a-/-, G3Terc-/-, and G3-dKO mice (7 months old). Arrows indicate γH2AX positive nuclei (C). Quantification of the percentage of γH2AX positive cells per small intestine crypt (n=4-6 mice per group) (D).

E,F Representative images of small intestine crypt sections stained with antibody against pATM (Ser-1981)(E). Quantification of frequencies of pATM (Ser-1981) positive crypt cells (n=4-6 of 7-month old mice per genotype) (F).

G,H Representative image of the anaphase bridge (indicated with arrows) in the basal crypt of G3Terc-/- mice (G). The percentage of cells with anaphase bridges in pH3 positive basal crypt cell population (n=4-6 mice per genotype) (H).

I Heat map indicating the relative expression levels of DDR and cell cycle-related gene transcripts in MEFs from indicated genotypes (Data are extracted from RNAseq datasets).

Data information: All scale bars represent 25 μm. Data are presented as mean±s.e.m.
Telomeric epigenetic response mediated by Gadd45a regulates stem cell aging and lifespan (2018)
Figure 4

Figure 4. Gadd45a deletion restores subtelomeric DNA methylation and telomeric heterochromatin marks in G3Terc-/- mice.

A,B CpG island methylation status of subtelomeric regions on chromosome 1q. gDNAs from MEF with indicated genotypes were PCR amplified and analysed by bisulfite sequencing. Yellow and blue bars denote the frequencies of methylated and unmethylated CpG islands, respectively, at each position (A). Quantification of subtelomeric CpG island methylation (B).

C-E Telomeric quantification and dot-bot assay of tri-methyl H3K9 (H3K9me3)-ChIP in indicated genotypes (C, up and low panel). Telomeric quantification and dot-bot assay of acetylated H3K9 (H3K9ac)-ChIP in indicated genotypes (D, up and low panel). Telomeric quantification and dot-bot assay of HP1a-ChIP in indicated genotypes (E, up and low panel).

Data information: For Figs. 4C-E, experiments were conducted in duplicates, and 3 independent cell lines were used for each group. Data are presented as mean±s.e.m.