Fig. 5. Recovery of nascent DNA degradation upon RS-1 treatment.

a Nascent DNA degradation analysis in U2OS cells transfected with the indicated siRNA after exposure to HU in the presence or absence of 10 µM RS-1. The IdU/CldU ratio calculations are done as outlined for Fig. 2a. Representative of n = 3 independent experiments. Data are represented as median ± 95% CI (n = 250 DNA fibers). ****p < 0.0001 by two-tailed Mann–Whitney test. b Immunoblot analysis of chromatin fractions for the detection of RAD51 protein. U2OS cells were transfected with the indicated siRNA and exposed to 4 mM HU for 5 h in the presence or absence of 10 µM RS-1. c Nascent DNA degradation analysis in LSH-deficient lymphocytes (KO) or control lymphocytes (Ctrl) after exposure to HU in the presence or absence of 10 µM RS-1. The IdU/CldU ratio calculations are done as outlined for Fig. 2a. Representative of n = 3 biologically independent experiments. Data are represented as median ± 95% CI (n = 250 DNA fibers). ****p < 0.0001 by two-tailed Mann–Whitney test. d Immunoblot analysis of chromatin fractions for the detection of γH2AX. U2OS cells transfected with the indicated siRNA were analyzed by immunoblotting after exposure to 2 mM HU for 24 h in the presence or absence of 10 µM RS-1. e DNA damage was evaluated by neutral comet assay in U2OS cells transfected with the indicated shRNA after exposure to 2 mM HU for 24 h in the presence or absence of 10 µM RS-1. Left: representative images. Right: A scatter plot of tail length from comet assay. Representative of n = 3 independent experiments. Data are represented as mean ± SD (n = 50 cells). ****p < 0.0001 by Student’s two-tailed t test. f Chromosome aberrations to metaphase chromosome from LPS stimulated LSH-depleted lymphocytes (LSH-KO) in the presence or absence of 10 µM RS-1 was determined by Giemsa staining. Representative of n = 3 biologically independent experiments. Data are represented as mean ± SD (n = 54 cells). **p < 0.01 by Student’s two-tailed t test.