Article Figures & Data
Figures
- FIGURE 1.
Low-dose radiation exposure induces significant loss of DNA methylation in the murine thymus. Levels of global genome DNA methylation in the thymus tissue of irradiated mice were measured by the cytosine extension assay on treatment of DNA with a methylation-sensitive restriction enzyme HpaII that cleaves CCGG sequences when internal cytosine residues are unmethylated on both strands. The enzyme leaves a 5′-guanine overhang after DNA cleavage that is used for subsequent single nucleotide extension with labeled [3H]dCTP. The extent of [3H]dCTP incorporation opposite the exposed guanine is directly proportional to the number of cleaved and thus unmethylated CpG sites and inversely proportional to the levels of methylation (i.e., the higher the methylation, the lower is the incorporation of [3H]dCTP). Columns, mean levels of [3H]dCTP (n = 6); bars, SE. a, P < 0.05, significantly different from control; b, P < 0.05, significantly different from female mice; c, P < 0.05, significantly different from male mice exposed to acute low-dose radiation, ANOVA. Black columns, females; gray columns, males.
- FIGURE 2.
Radiation-induced accumulation of γH2AX foci in murine thymus. Histone H2AX is phosphorylated at Ser139 immediately on induction of DNA damage, forming γH2AX foci, which revealed DNA double-strand breaks. A. Control female. B. Control male. C. Female, fractionated exposure. D. Male, fractionated exposure. E. Female, acute exposure. F. Male, acute exposure. G. Levels of γH2AX foci per cell. Columns, mean (n = 240); bars, SE. a, P < 0.05, significantly different from control. Black columns, females; gray columns, males.
- FIGURE 3.
Induction and persistence of fractionated radiation-induced γH2AX foci in murine thymus. A. Control female. B. Control male. C. Female, fractionated exposure, 0.5 Gy total dose, 4 hours after the last fraction. D. Male, fractionated exposure, 0.5 Gy total dose, 4 hours after the last fraction. E. Female, fractionated exposure, 0.5 Gy total dose, 24 hours after the last fraction. F. Male, fractionated exposure, 0.5 Gy total dose, 24 hours after the last fraction; G. Female exposed to a single fraction of 0.05 Gy, 4 hours after exposure. H. Male exposed to a single fraction of 0.05 Gy, 4 hours after exposure.
- FIGURE 4.
Radiation exposure alters the expression of DNA (cytosine-5) methyltransferases and methyl-binding proteins in thymus of female and male mice. Lysates from thymus tissue were subjected to immunoblotting using antibodies against DNMT1, DNMT3a, DNMT3b, MBD2, and MeCP2. Columns, mean protein levels relative to those of control animals; bars, SE. a, P < 0.05, significantly different from control; b, P < 0.05, significantly different from female mice, Student's t test. Black columns, females; gray columns, males. CT, control animals; FR, animals subjected to fractionated exposure; AC, acutely exposed animals. All sample loading was normalized to protein content. Representative Western blots from three independent experiments are shown; each lane represents a protein extract of a thymus of one animal.
- FIGURE 5.
Radiation-induced loss of histone H4-Lys20 trimethylation in thymus tissue. Acidic thymus tissue lysates enriched in histones were subjected to immunoblotting using anti–trimethyl histone H4-Lys20 antibodies. Columns, mean levels relative to those of control animals; bars, SD. a, P < 0.05, significantly different from control, Student's t test. Black columns, females; gray columns, males. All sample loading was normalized for protein content. Representative blots from two independent experiments are shown.
Volume 3, Issue 10
