Fig. 1

Senescent cells accumulate elevated levels of H4K20me3 in vitro and in vivo. a Quantification of SA β-galactosidase-positive (SA β-gal+) IMR90 cells 3–12 days after infection with either empty vector control (CON) or H-RASG12V virus. b Cells from a were pulse labeled with 5-ethynyl-2′-deoxyuridine (EdU) and positive cells scored. c Western blot of indicated proteins in whole cell extracts of cells from a. d Western blot of H4K20me3 and histone H4 in whole cell extracts from c, normalized for total histone H4 content. e Western blot of histone H4 and indicated H4K20 modifications from whole cell extracts of proliferating (PRO), replicative senescent (RS), control-infected proliferating (CON) and H-RASG12V-infected senescent (OIS) IMR90 cells, normalized for total histone H4 content. f Western blot of H4K20me3 and histone H4 from whole cell extracts of proliferating (PRO), RS and quiescent (QUI) IMR90 cells, normalized for total histone H4 content; (s) and (l) denote short and long autoradiographic exposures, respectively. Experiments in a–f are representative of at least five similar experiments. g Western blot of SUV420H2 and GAPDH from whole cell extracts of PRO, RS, CON, and OIS cells. h Immunofluorescent images of H4K20me3 staining in CON and OIS cells 12 days after infection. i Quantitative image analysis of H4K20me3 immunofluorescence in CON and OIS cells (181 CON and 129 OIS cells were scored). j Relative percentages of the different methylation states of H4K20 in PRO and RS cells as determined by quantitative mass spectrometry; error bars represent standard error of the mean. k Immunohistochemical images of human melanocytic nevus (N) and overlaying epidermis (E) stained with antibodies against Melan-A and H4K20me3. The arrow indicates a non-nevus epidermal melanocyte. Data are representative of at least ten different human nevi