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. 2006 Oct 26;25(22):5317–5328. doi: 10.1038/sj.emboj.7601404

Figure 6.

Figure 6

MBD1-SUMO does not efficiently repress p53BP2 transcription. (A) Schematic representation of p53BP2 promoter. The regions amplified by PCR in ChIP (C) and RT–PCR (B) experiments are indicated. CpGs are shown as vertical bars. (B) Semiquantitative RT–PCRs detect p53BP2 transcription in cells transfected with either SETDB1 siRNA or with GFP-PIAS1 and Flag-SUMO1 (P1/F-S1) but not in cells transfected with mutant GFP-PIAS1 and Flag-SUMO1 (C350S/F-S1). The nonsumoylatable MBD12KA (2KA) rescues p53BP2 repression in GFP-PIAS1/ Flag-SUMO1-transfected cells. Neither the wtMBD1 proteins nor MBD1 with single lysine mutations (K450A and K489A) could efficiently rescue p53BP2 repression. GAPDH is a ubiquitously expressed control gene. The triangles above the RT–PCR lanes represent two-fold increase in the amount of cDNA used in the PCR reactions. M is a molecular weight DNA marker. (C) Chromatin IPs detect MBD1 and MBD1-SUMO1 bound to the p53BP2 promoter. SETDB1 and H3K9 methylation are reduced at the p53BP2 promoter in cells overexpressing GFP-PIAS1 (P1) and GFP-PIAS1 and Flag-SUMO1 (P1/F-S1) but not in cells overexpressing mutant GFP-PIAS1 and Flag-SUMO1 (PC350S/F-S1).