Role of CTCF binding sites in the Igf2/H19 imprinting control region.
A approximately 2.4-kb imprinting control region (ICR) regulates somatic monoallelic expression of the Igf2 and H19 genes. This is achieved through DNA methylation-dependent chromatin insulator and promoter silencing activities on the maternal and paternal chromosomes, respectively. In somatic cells, the hypomethylated maternally inherited ICR binds the insulator protein CTCF at four sites and blocks activity of the proximal Igf2 promoter by insulating it from its distal enhancers. CTCF binding is thought to play a direct role in inhibiting methylation of the ICR in female germ cells and in somatic cells and, therefore, in establishing and maintaining imprinting of the Igf2/H19 region. Here, we report on the effects of eliminating ICR CTCF binding by severely mutating all four sites in mice. We found that in the female and male germ lines, the mutant ICR remained hypomethylated and hypermethylated, respectively, showing that the CTCF binding sites are dispensable for imprinting establishment. Postfertilization, the maternal mutant ICR acquired methylation, which could be explained by loss of methylation inhibition, which is normally provided by CTCF binding. Adjacent regions in cis-the H19 promoter and gene-also acquired methylation, accompanied by downregulation of H19. This could be the result of a silencing effect of the methylated maternal ICR.
Szabó
Piroska E
PE
Division of Biology, Beckman Research Institute of the City of Hope, 1450 East Duarte Rd., Duarte, CA 91010-3011, USA. pszabo@coh.org
Tang
Shih-Huey E
SH
Silva
Francisco J
FJ
Tsark
Walter M K
WM
Mann
Jeffrey R
JR
D000818:Animals; D001665:Binding Sites; D000076246:CCCTC-Binding Factor; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology; D018392:Genomic Imprinting; D007335:Insulin-Like Growth Factor II / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D009154:Mutation; D011485:Protein Binding; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated / Q000096:biosynthesis* / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism*
D000076246:CCCTC-Binding Factor; C000619698:Ctcf protein, mouse; D004268:DNA-Binding Proteins; C094335:H19 long non-coding RNA; D062085:RNA, Long Noncoding; D022661:RNA, Untranslated; D012097:Repressor Proteins; D007335:Insulin-Like Growth Factor II