Fig. 8

Transient enrichment of H3K27me3 near the nuclear lamina is lost when EZH1/2 is blocked. Confocal and dSTORM super-resolution images of immunofluorescence staining at E11.5 XX and XY gonads cultured for 48 h with either DMSO (control) or 10 μm GSK126. a Confocal images (×80) showing efficacy of H3K27me3 depletion by GSK126, eGFP (germ cells; green) and H3K27me3 (red). 10 μm scale bars. The reduction in H3K27me3 in GSK126-treated samples is quantified in Supp. Figure 3. b Super-resolution dSTORM images. Left panels show merged wide-field (×160) images: eGFP (germ cells: green) and H3K27me3 (red). White dotted boxes indicate super-resolved germ cells. 10 μm scale bars. Right panels show dSTORM super-resolution images of H3K27me3 (grayscale) control and GSK126-treated germ cells. 1 μm scale bars. Representative images chosen from 3 to 5 super-resolved images in three biological replicates for each time point. c Quantification of H3K27me3 localization in E11.5 XX and XY gonads cultured for 48 h with either DMSO (control) or 10 μm GSK126. ×80 confocal immunofluorescent images were analyzed by ImageJ Cell Counter. Percentages of cells with uniform nuclear localization (UNL; black bars) and peripheral nuclear localization (PNL; gray bars) are shown in the stacked histogram. Data represent 3–6 biological replicates for each sex, for each treatment group. (XX/XY cells counted: vehicle control n = 159/99; GSK126 n = 180/115)