It is thought that the H19 imprinting control region (ICR) directs the silencing 
of the maternally inherited Igf2 allele through a CTCF-dependent chromatin 
insulator. The ICR has been shown to interact physically with a silencer region 
in Igf2, differentially methylated region (DMR)1, but the role of CTCF in this 
chromatin loop and whether it restricts the physical access of distal enhancers 
to Igf2 is not known. We performed systematic chromosome conformation capture 
analyses in the Igf2/H19 region over >160 kb, identifying sequences that 
interact physically with the distal enhancers and the ICR. We found that, on the 
paternal chromosome, enhancers interact with the Igf2 promoters but that, on the 
maternal allele, this is prevented by CTCF binding within the H19 ICR. CTCF 
binding in the maternal ICR regulates its interaction with matrix attachment 
region (MAR)3 and DMR1 at Igf2, thus forming a tight loop around the maternal 
Igf2 locus, which may contribute to its silencing. Mutation of CTCF binding 
sites in the H19 ICR leads to loss of CTCF binding and de novo methylation of a 
CTCF target site within Igf2 DMR1, showing that CTCF can coordinate regional 
epigenetic marks. This systematic chromosome conformation capture analysis of an 
imprinting cluster reveals that CTCF has a critical role in the epigenetic 
regulation of higher-order chromatin structure and gene silencing over 
considerable distances in the genome.