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. 2018 Dec 7;11(9):761–769. doi: 10.1093/jmcb/mjy074

Figure 2.

Figure 2

AFF3 binds to the methylated allele of XIST in female cells. (A) Schematic illustration of the location of the XIST CpG island/XIST DMR, the amplified genomic regions of the XIST DMR, and the amplified genomic region of the XIST DMR after bisulfite treatment. (B and C) AFF3 is recruited to the XIST DMR in HEK293T (B) and IMR-90 (C) cells used in ChIP assay. The nonexpressed β-globin gene (HEMO) served as a negative control. Error bars represent standard deviations. (D) AFF3 binds to the methylated allele of the XIST DMR. MeDIP assays were performed using AFF3 or AFF4 ChIP DNA in HEK293T cells. AFF4 ChIP DNA was used as a negative control for the MeDIP assay. Primer pair P2 was used to amplify the XIST DMR. The HEMO gene served as a negative control for ChIP-qPCR. Error bars represent standard deviations. (E) Bisulfite-sequencing analysis of whole-cell extract (WCE; input for AFF3 ChIP) and AFF3 ChIP in HEK293T at the XIST DMR. Methylated and unmethylated cytosines are designated by filled and unfilled circles, respectively. Each line indicates a unique DNA clone. In contrast to WCE, the AFF3 ChIP DNA is predominantly methylated at the XIST DMR.