Figure 5. ChIP analysis to quantify the H3K27me3 and H3K4me3 marks on p16^INK4A exon 1 when EBNA3C is inactivated and re-activated in LCL 3CHT.

(A) Schematic of the human p16^INK4A-ARF locus showing the location of coding exons (boxes) and transcription start sites (horizontal arrows) – not drawn to scale. The vertical (A-D) arrows refer to the approximate locations of primer pairs used for qPCR analysis of precipitated chromatin (as described in Materials and Methods). (B) ChIP analysis of H3K27me3 distribution on exon 1 (site C) of p16^INK4A. The histogram shows a decline in H3K27me3 relative to a cycling LCL 3CHT (day 0). (C) Corresponding changes in p16^INK4A mRNA quantified by qRT-PCR. (D) ChIP analysis of H3K27me3 distribution across the p16^INK4A-ARF locus in LCL 3CHT proliferating in the presence of HT, after 30 days without HT and 20 days after re-adding HT. (E) ChIP analysis of H3K4me3 the p16^INK4A-ARF locus in LCL 3CHT proliferating in the presence of HT, after 30 days without HT and 20 days after re-adding HT.