FIGURE 5.

DNA methylation of major satellite repeats was completely restored in the Suv39h dn ES cells expressing Suv39h1ΔN. A, genomic DNA isolated from the indicated cell lines was digested with the methylation-sensitive restriction enzyme MaeII. Southern blot analysis was conducted using a major satellite repeat probe. Immunohistochemical staining analysis for Dnmt3a (B) or Dnmt3b (C) (red) and HP1β (green) were conducted with WT or Suv39h dn ES cells or Suv39h dn ES cells complemented with either FLAG-tagged WT Suv39h1, H324L, or ΔN. DNA was counterstained with DAPI (blue). D, DAPI intensity and the intensity of the antigen at the same region were quantified using ImageJ based on the fluorescence imaging data obtained for B and C. The intensities of 30 cells were quantified. The value of each antigen was expressed as a ratio between the DAPI intensity and the intensity of the antigen being measured. E, Western blot analysis of endogenous Dnmt3a and -3b in the indicated cell lines. F and G, immunohistochemical staining analysis for the two independent cell lines indicated at low magnification. A combination of color and antigen was same as that observed for B and C.