Normal and Pathologic Functions of CTCF and Its Distinct Classes of DNA-targets

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CTCF is a highly conserved, multi-functional nuclear factor involved in many aspects of gene regulation ranging from gene activation or repression to enhancer blocking and hormone regulated silencing. CTCF-proteins in many species, including Drosophila, have similar size and contain central highly conserved 11 Zn-finger DNA-binding domain that mediates multiple sequence-specificity of DNA binding flanked by N- and C-terminal fragments of roughly equal size. By making inter- and intra-chromosomal loops through selective interactions between various DNA-bound CTCF molecules at different genomic targets, CTCF can act as a universal and versatile component of chromatin-insulators and boundaries for spreading of nucleosome modifications, of non-coding intergenic transcripts, and of bi- and/or mono- allelic regional DNA methylation. It can also function as a promoter-proximal repressor or activator. Depending on the context, CTCF may also couple DNA-looping with activity of transcriptional enhancers. Genome-wide in-vivo mapping of tens of thousands of DNA/CTCF-complexes showed that CTCF can position nucleosomes around nuclease hypersensitive sites that landmark insulators, enhancers, and other regulatory sequences. The featuring of so many functions suggested that CTCF has important interacting partners (ranging from B23/nucleophosmin to YY1 and cohesins) and that it is an essential factor. Indeed, CTCF knockout in mice is lethal at early stages of embryo development. However, expression of ectopic CTCF results in a profound negative regulation of proliferation, indicating that CTCF may be a tumor suppressor gene (TSG). Human CTCF maps within 16q22 region of recurrent LOH in many tumors. Accordingly, several functional point mutations in the 11ZF DBD of CTCF have been characterized in tumors selected for the loss of the second CTCF allele. Given the importance of CTCF for development, cell proliferation, etc., we not only analyzed genome wide CTCF targets for the first time (Kim TH, Abdullaev ZK, Smith AD, et al., Cell 2007, vol. 128, pp1231-1245) but also studied the in vivo roles of CTCF in adult tissues and during embryonic development. We depleted maternal stores of CTCF from growing mouse oocytes using transgenic RNAi technology, and identified hundreds of misregulated genes. Moreover, our analysis suggests that CTCF predominantly activates or derepresses transcription in oocytes. CTCF depletion causes meiotic defects in the egg, and mitotic defects in the embryo that are accompanied by defects in zygotic gene expression, and culminate in apoptosis. Maternal pronuclear transfer and CTCF mRNA microinjection experiments indicate that CTCF is a mammalian maternal effect gene, and that persistent transcriptional defects rather than persistent chromosomal defects perturb early embryonic development. This is the first study detailing a global and essential role for CTCF in mouse oocytes and preimplantation embryos (Wan LB, Pan H, Hannenhalli S, et.al., Development 2008 vol.135, pp. 2729-2738). We also detailed hTERT up-regulation by methylation of the first exon CTCF methylation sensitive repressive site. We showed that in many cancers as well as in the reporter system methylation of exon 1 CTCF site results in activation of hTERT gene.This observation was rather unexpected as usually DNA methylation is associated with gene inactivation (Renaud S, Loukinov D, Abdullaev Z, et. al., Nucleic Acids Res 2007 vol. 35, pp.1245-56). Finally, we discovered role of CTCF as repressor of all three promoters of human BORIS gene. We showed that blocking of CTCF in normal cells results in demethylation and derepression of BORIS promoters. We identified a number of CTCF sites in 5non-coding region of BORIS gene. Our results provide a basis for understanding of functional connection between a lessening of the strictness of BORIS silencing in somatic cells with haploinsufficiency of CTCF observed in cancers. (Renaud S, Pugacheva EM, Delgado MD, et.al. Nucleic Acids Res, 2007, vol. 35, pp.7372-7388).