Fig 5

psyAShM transition exerts an allelic effect on TF binding affinity. A Estimates for four discordant AShM sites in PDC twin pairs. Each point represents the β0 value of the psyAShM sites showing the strongest evidence of AShM in unaffected or affected individuals (highlighted in red, p<1e−3) and a discordant AShM pattern in PDC twin pairs (BF>10). Error bars indicate 95% credible intervals. B eQTL patterns of the four psyAShM sites from the GTEx dataset. C Allelic effects of rs4558409 (G/T), rs708769 (T/C), rs10866916 (T/C), and rs2851443 (C/T) on promoter activity of the luciferase reporter in HEK293T and SK-N-SH cells are shown in columns with standard errors indicated by bars. The gray column represents cells transfected with pGL4.23-empty vector (Con), the orange column represents cells transfected with the reference allele-containing fragment cloned in pGL4.23, and the red column represents the alternative allele-containing fragment. D, E Effect of POU3F2 overexpression on endogenous PLLP expression (D) or rs4558409 allele-dependent promoter activity of the luciferase reporter (E) in HEK293T cells. Vec represents the pcDNA3.1 empty vector. F Sanger sequencing traces from ChIP-PCRs of POU3F2 and IgG at the rs4558409 (G/T) site in HEK293T cells with a heterozygous rs4558409 genotype. G EMSA and competition analysis. Assays were performed with the rs4558409 alternative T allele (red bold) as the hot probe, the rs4558409 alternative T allele or the reference G allele as the cold probe (left) and the alternative T allele without or with hydroxymethylated CpG (red bold) as another cold probe (right) with HEK293T nuclear extracts. Fold differences of molar excess of the cold probe compared to the hot probe and the relative intensity of the binding complex are shown underneath each panel. **p < 0.01, ***p < 0.001, ****p< 0.0001 or ns, nonsignificant from t test