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. 2021 Jun 10;12:3520. doi: 10.1038/s41467-021-23809-2

Fig. 8. A role of H4K20 methylation at stalled forks in LSH-deficient cells.

Fig. 8

a Representative images (left panel) and quantification of PLA signals (H4K20me2: EdU) from U2OS cells transfected with the indicated siRNA (siCtrl or siLSH) after exposure to 4 mM HU for 5 h. The scale bar represents 10 µm. Quantification of PLA signals (H4K20me2: EdU) is based on EdU-positive cells. Representative of n = 3 independent experiments. Data are represented as median ± 95% CI (n = 200 cells). ****p < 0.0001 by two-tailed Mann–Whitney test. b Quantification of EdU signals from U2OS cells transfected with the indicated siRNA after exposure to 4 mM HU for 5 h. Representative of n = 3 independent experiments. Data are represented as median ± 95% CI (n = 200 cells). ns = not significant by two-tailed Mann–Whitney test. c Quantification of PLA signals (H4K20me2: EdU) from U2OS cells transfected with the indicated siRNA (siCtrl or simH2A) after exposure to 4 mM HU for 5 h. Quantification of PLA signals (H4K20me2: EdU) is based on EdU-positive cells. Representative of n = 3 independent experiments. Data are represented as median ± 95% CI (n = 200 cells). ****p < 0.0001 by two-tailed Mann–Whitney test. d ChIP-qPCR analysis for detection of H4K20me2 enrichment at CFSs in mock-treated (DMSO) or aphidicolin-treated (APH) U2OS cells transfected with shLSH or shCtrl. Data are represented as mean ± SD (n = 3 independent experiments). *p < 0.05, **p < 0.01, ***p < 0.001 by Student’s two-tailed t test. e Quantification of PLA signals (H4K20me2: EdU) from U2OS cells transfected with the indicated siRNA (siCtrl or siLSH or siLSH together with siKMT5) after exposure to 4 mM HU for 5 h. Quantification of PLA signals (H4K20me2: EdU) is based on EdU-positive cells. Representative of n = 3 independent experiments. Data are represented as median ± 95% CI (n = 200 cells). ****p < 0.0001 by two-tailed Mann–Whitney test. f Quantification of PLA signals (53BP1: EdU) from U2OS cells transfected with the indicated siRNA (siCtrl or siLSH or siLSH together with siKMT5) after exposure to 4 mM HU for 5 h. Quantification of PLA signals (53BP1: EdU) is based on EdU-positive cells. Representative of n = 3 independent experiments. Data are represented as median ± 95% CI (n = 200 cells). ****p < 0.0001 by two-tailed Mann–Whitney test. g Quantification of PLA signals (BRCA1: EdU) from U2OS cells transfected with the indicated siRNA (siCtrl or siLSH or siLSH together with siKMT5) after exposure to 4 mM HU for 5 h. Quantification of PLA signals (BRCA1: EdU) is based on EdU-positive cells. Representative of n = 3 independent experiments. Data are represented as median ± 95% CI (n = 200 cells). ****p < 0.0001 by two-tailed Mann–Whitney test.