CTCF interacts with and recruits the largest subunit of RNA polymerase II to CTCF target sites genome-wide

CTCF is a transcription factor with highly versatile functions ranging from gene activation and repression to the regulation of insulator function and imprinting. Although many of these functions rely on CTCF-DNA interactions, it is an emerging realization that CTCF-dependent molecular processes involve CTCF interactions with other proteins. In this study, we report the association of a subpopulation of CTCF with the RNA polymerase II (Pot II) protein complex. We identified the largest subunit of Pot II (LS Pot II) as a protein significantly colocalizing with CTCF in the nucleus and specifically interacting with CTCF in vivo and in vitro. The role of CTCF as a link between DNA and LS Pot II has been reinforced by the observation that the association of LS Pot II with CTCF target sites in vivo depends on intact CTCF binding sequences. Serial chromatin immunoprecipitation (ChIP) analysis revealed that both CTCF and LS Pot II were present at the P-globin insulator in proliferating HD3 cells but not in differentiated globin synthesizing HD3 cells. Further, a single wild-type CTCF target site (N-Myc-CTCF), but not the mutant site deficient for CTCF binding, was sufficient to activate the transcription from the promoterless reporter gene in stably transfected cells. Finally, a ChIP-on-ChIP hybridization assay using microarrays of a library of CTCF target sites revealed that many intergenic CTCF target sequences interacted with both CTCF and LS Pot II. We discuss the possible implications of our observations with respect to plausible mechanisms of transcriptional regulation via a CTCFmediated direct link of LS Pot II to the DNA.