Fig. 5: REV3L is essential to protect and restart stalled replication forks. | Nature Communications

Fig. 5: REV3L is essential to protect and restart stalled replication forks.

From: MAD2L2 promotes replication fork protection and recovery in a shieldin-independent and REV3L-dependent manner

Fig. 5

a Quantification of survival assays in control (sgCTRL) and REV3L-depleted HeLa cells, untreated or treated with 0.5 μΜ MMC or 1 μΜ CIS. Bars represent the mean ± SD. Each dot represents one of three independent experiments. b Schematic and quantification of EdU incorporation intensity in control and REV3L-depleted HeLa cells. Cells were incubated with or without 5 μΜ MMC for 24 h, followed by EdU labeling for 2 h before harvesting. White bars represent the median of three independent experiments. c Schematic and quantification of fork progression assays in control and REV3L-depleted HeLa cells, untreated or treated with 4 mM HU for 1 h. A representative fiber experiment from two independent biological replicates is shown. d Schematic and quantification of fork degradation assays. Control and REV3L-depleted HeLa cells were treated with 4 mM HU ± 50 μΜ mirin for 4 h. Open circles, no treatment; closed circles, treatment. A representative fiber experiment from three independent biological replicates is shown. e Schematic and quantification of fork restart, fork stalling, and origin firing in control and REV3L-depleted HeLa cells treated with 4 mM HU for 1 h as indicated. Data are represented as mean ± SD. Each dot represents one of three independent experiments. f Quantification of the percentage of cells with REV3L/biotin PLA foci in control and MAD2L2-depleted HeLa cells, untreated or treated with 4 mM HU for 2 h. Bars represent the mean ± SD. Each dot represents one of three independent experiments. Statistical analysis in (a, e) was performed according to one-way ANOVA with Dunnett’s multiple comparisons test. Statistical analysis in (b–d) was performed according to two-tailed Mann-Whitney test. Pink bars in the fiber plots represent the mean. Additional replicates and combined fiber plots are provided in the Supplementary Information. Statistical analysis in (f) was performed according to two-way ANOVA with Tukey’s multiple comparisons test.

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