Fig. 4

Chromatin Immunoprecipitation (ChIP) analysis of Ikaros, NFKB1 (NF-kappa B p50), THAP11, and HCF-1 binding to M4-bearing promoters of the genes Skp2, CDC25A, and G9a. Formaldehyde crosslinked chromatin of Molt-IK-HA or Molt4-THAP11-HA cells were sheared by ultrasonication into an average length of 300 to 1000 bps to be subjected to immuno-precipitation assays using anti-NF-kappa B p50, anti-HCF-1, anti-HA/Ikaros, anti-HA/THAP11, or preimmune IgG sera (control). The TF-associated chromatin were analyzed via quantitative real time PCR (qRT-PCR) using primer sets flanking the suspected TF binding site and background binding control regions: Promoter (M4 motif), Exon (exon control), ±2000 bp (control regions downstream and upstream from the corresponding M4 cis-element). Enrichment of TF-bound DNA amount were normalized to corresponding amount in input. ChIP results (average of three technical replicates) for Ikaros (a), NFKB1 (NF-kappa B p50) (b), THAP11 (c) and HCF-1 (d) are shown