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. 2021 Nov 29;31(10):1610–1621. doi: 10.1093/hmg/ddab347

Figure 3.

Figure 3

Allele-specific transcription factor binding at rs7247241. (A) rs7247241 is located in the PPP1R14A promoter region with CTCF binding across human prostate cell lines. (B) JASPAR prediction of rs7247241 flanking sequences with allele difference. (C) Western blot analysis of CTCF, PLAGL2 and PPP1R14A protein expressions after stably knocking down CTCF and PLAGL2 in PrEC cells. (D–F) mRNA correlation between a putative regulatory transcription factor and PPP1R14A in ICGC (D), TCGA (E) and Broad Cornell (F) prostate primary tumor samples. (G, H) Relative enrichment measured by CTCF and PLAGL2 ChIP-qPCR at rs7247241 locus in 22Rv1 (G) and RWPE-1 cells (H). Columns represent the means of a total of four independent experiments. Error bars represent the standard error of the mean. (I, J) rs7247241 allele dosages in ChIP-qPCR amplicons from 22Rv1 (I) and RWPE-1 cells (J). (K) rs7247241 allele dosages in CTCF ChIP-seq tags from rs7247241 heterozygote prostate cells. (L, M) Quantification of promoter activities between rs7247241 T and C alleles by reporter assays in 22Rv1 (L) and PC3 (M) cells. Columns represent the means of a total of eight independent experiments. Error bars represent the standard error of the mean. (N) rs7247241 allele dosages in MBD-seq reads in 22Rv1 cells. (O) Correlations between CTCF occupancies and methylation status at rs7247241 locus in 22 human cell lines from ENCODE database. Each dot represents a cell line, and the solid line indicates the linear regression coefficient, and the dashed lines depict 95% confidence interval.