SUV4-20h1 mediates transcriptional silencing of the γ-gene. (A) Expression of SUV4-20h1 and (B) CK2α was analyzed in K562 cells expressing specific shRNAs or scrambled control sequences (scr). (C) H4K20me3 levels at the γ-promoter in SUV4-20h1-kd and scr control cells, and (D) H4S1ph levels at the γ-promoter in CK2α-kd and scr control cells were measured by ChIP analysis. The error bars correspond to the SD. Each experiment was performed twice independently. Normal rabbit IgG served as the control. (E) Analysis of γ-globin gene expression in SUV4-20h1-kd and scr control cells, and (F) CK2α-kd and scr control cells. (G) DNA methylation at the human γ-gene in SUV4-20h1-kd, CK2α-kd and scr control cells. Each row shows the methylation status of individual CpG dinucleotides derived from sequence analysis of 25 representative (of at least 40) individual cloned polymerase chain reaction (PCR) products of the γ-genes after bisulfite modification from knockdown or scr control K562 cells. The differences between the 2 knockdown lines and the scrambled control were highly significant (P < .05). The numbers at the top represent the positions of the CpG dinucleotides relative to the transcriptional start site of the γ-gene. (H) ChIP of the H4R3me2s level and (I) PRMT5 binding at the γ-promoter in SUV4-20h1-kd and scr control cells. Normal rabbit IgG served as the control. The error bars correspond to the SD. Each experiment was performed 3 times independently.