Figure 4

SET7/9-mediated YY1 methylation regulates YY1 genomic association.

(A,B) Genomic distribution of YY1 binding sites. ChIP-seq was performed in HeLa cells using anti-YY1 antibody, followed by peak finding (A) and motif analysis (B) using HOMER. (C) YY1 binding detected by ChIP-seq was shown for p53, RAD1 and ABL1 genes, as indicated. (D) HeLa cells were transfected with control vector or vector expressing SET7/9, followed by ChIP with anti-YY1 antibody and q-PCR with primers specifically targeting promoter regions of selected genes as indicated. ChIP signals were presented as percentage of inputs (±s.e.m., **P < 0.01, ***P < 0.001). Experiments were repeated three times and representative data was shown. (E) HeLa cells were transfected with vectors expressing Flag-tagged YY1(wt), YY1(K173R) or YY1(K411R), followed by ChIP with anti-Flag antibody and q-PCR with primers specifically targeting promoter regions of selected genes as indicated. ChIP signals were presented as percentage of inputs (±s.e.m., **P < 0.01, ***P < 0.001). Experiments were repeated three times and representative data was shown. (F) HeLa cells stably expressing Flag-tagged YY1(wt) or YY1(K173R) were subjected to affinity purification with Flag M2 agarose. The resultant proteins were separated by SDS-PAGE gel, followed by silver staining. Four bands (1–4) specifically present in YY1(wt) sample were cut and subjected to mass spectrometry (MS) analysis.