Figure 7. Sp1 binding sites within CpG island in LEDGF promoter were enriched with SUV39H1 and dimethylated histones in LECs. (A) Western analysis of SUV39H1, H3K9me2, and H3K9me3 in the extract isolated from LECs exposed to UVB radiation. Right panel shows the relative densitometry of protein bands. (B) ChIP analyses revealed recruitment of histone methyltransferase SUV39H1 and enrichment of H3K9me2 at CpG island containing Sp1 sites. Chromatin immunoprecipitation analysis was performed using chromatin extract from LECs exposed to multiple doses of UVB radiation as shown in the scheme (Fig. 1E) with specific antibodies, as indicated. Top panel shows LEDGF promoter and location of PCR primers designed for the study. DNA fragments present in the immunoprecipitates were amplified with primers that specifically recognize a fragment of the LEDGF promoter containing Sp1 binding sites (-175 to +27). As a negative control, primers designed beyond the Sp1 sites (-2499 to -2277) or a mock ChIP with control IgG was used. PCR products were separated on 2.5% agarose gels and stained with ethidium bromide.