FIG. 5.

Allele-specific chromatin marks at the TERT minimal promoter in thyroid cancer cell lines. (A) ChIP with either H3K4me3 (activating chromatin mark) or H3K27me3 (silencing chromatin mark) antibodies. Binding was measured relative to 5% input chromatin, and binding at the TERT locus assessed by qPCR. H3K4me3 binding at the TERT promoter was most abundant in the homozygous mutant cell line FTC-133 (black bar), intermediate in the heterozygous mutant cell lines (gray bars), and least abundant in the homozygous wild-type cell lines (white bars with cancer cell line WRO denoted by stipple compared with normal cell line NThy-ori-3). H3K27me3 binding was reciprocal of the H3K4me3 binding. All error bars represent standard error within triplicates. (B) Sanger sequencing of the TERT promoter immunoprecipitated DNA of heterozygous TERT mutant thyroid cancer cell lines at the −124 C > T mutation compared with input genomic DNA before ChIP. At the −124 TERT position, the mutant allele is represented by the red curve (T), while the wild-type allele is represented by the blue curve (C). The input genomic DNA shows the presence of both alleles at the −124 position, while the H3K4me3 immunoprecipitated DNA shows only the red curve, indicative of the mutant allele. H3K27me3 immunoprecipitated DNA shows only the blue curve, indicative of the wild-type allele. qPCR, quantitative PCR.