The CTCF protein is a highly conserved zinc finger protein that is implicated in many aspects of gene regulation and nuclear organization. Its functions include the ability to act as a repressor of genes, including the c-myc oncogene. In this paper, we show that the CTCF protein can be posttranslationally modified by the small ubiquitin-like protein SUMO. CTCF is SUMOylated both in vivo and in vitro, and we identify two major sites of SUMOylation in the protein. The posttranslational modification of CTCF by the SUMO proteins does not affect its ability to bind to DNA in vitro. SUMOylation of CTCF contributes to the repressive function of CTCF on the c-myc P2 promoter. We also found that CTCF and the repressive Polycomb protein, Pc2, are colocalized to nuclear Polycomb bodies. The Pc2 protein may act as a SUMO E3 ligase for CTCF, strongly enhancing its modification by SUMO 2 and 3. These studies expand the repertoire of posttranslational modifications of CTCF and suggest roles for such modifications in its regulation of epigenetic states.

BackgroundThe CTCF insulator protein is a highly conserved zinc finger protein that has been implicated in many aspects of gene regulation and nuclear organization. The protein has been hypothesized to organize the human genome by forming DNA loops.ResultsIn this paper, we report biochemical evidence to support the role for CTCF in forming DNA loops. We have measured DNA bending by CTCF at the chicken HS4 β-globin FII insulator element in vitro and have observed a unique DNA structure with aberrant electrophoretic mobility which we believe to be a DNA loop. CTCF is able to form this unusual DNA structure at two other binding sites: the c-myc P2 promoter and the chicken F1 lysozyme gene silencer. We also demonstrate that the length though not the sequence of the DNA downstream of the binding site is important for the ability of CTCF to form this unusual DNA structure. We hypothesize that a single CTCF protein molecule is able to act as a "looper" possibly through the use of several of its zinc fingers.ConclusionsCTCF is able to form an unusual DNA structure through the zinc finger domain of the protein. This unusual DNA structure is formed in a directional manner by the CTCF protein. The findings described in this paper suggest mechanisms by which CTCF is able to form DNA loops, organize the mammalian genome and function as an insulator protein.