Figure 4.

DOT1L-mediated H3K79 dimethylation is affected by Sly knockdown. (a) Immunofluorescence detection of H3K79 dimethylation (H3K79me2) in stage X testicular sections from WT and Sly-KD mice. Scale bar indicates 20 μm. Pictures were taken using the same image capture parameters (see also Supplementary Figure 6). (b) Schematic showing H3K79me2 level quantified by immunofluorescence in Sly-KD and WT step 10–12 elongating spermatids. The graph represents mean ±S.E.M. The difference between the two genotypes is statistically significant (t-test on six samples per genotype, P=0.0002). (c) ChIP-qPCR experiments on elongating/condensing spermatids from WT and Sly-KD mice, using antibody against H3K79me2. The Y-axis represents the mean enrichment (% IP/input) ±S.E.M. normalized to a negative control region (NC) located at ~170 kb from a TSS, and to the corresponding WT value. A significant decrease of H3K79me2 level at gene TSS was found in Sly-KD compared to WT samples when considering all tested genes (t-test, P<0.05, n= 3–4 samples per genotype). (d) Immunofluorescence detection of H3K79me2 (red) in WT epididymal spermatozoa. Hoechst (blue) was used to stain nuclei. Scale bar indicates 5 μm