FIGURE 6.

Phosphorylation of PHF8 influences its binding to the cyclin E promoter. A–E, HeLa cells were transfected with pCMV FLAG-PHF8, pCMV FLAG-PHF8-S844A, or empty vector as a control (C). The cells were harvested and subjected to ChIP assays. Briefly, the cells were treated with formaldehyde and sonicated. The sheared DNAs were immunoprecipitated (IP) with FLAG or H3K9me2 antibodies and then subjected to PCR with primers for the cyclin E promoter (A and C) and a quantitative PCR assay on the cyclin E promoter (B and D). E, the cell lysates were subjected to immunoblot (IB) analysis. F–H, HeLa cells were transfected with CDK2 siRNA. CDK2 knockdown efficiency was confirmed by immunoblot analysis (F). The cells were harvested and subjected to ChIP assays with primers for the cyclin E promoter as above (G and H).