Methylation signals at Rasgrf1
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Imprinted loci are excellent models for the study of DNA methylation and its effect on gene expression. Imprinted expression at these loci is controlled by parental specific methylation. At the paternally imprinted Rasgrf1 locus, expression in neonatal mouse brain is exclusively from the paternal allele, which is methylated at a Differentially Methylated Domain (DMD) 5 ' of a repetitive sequence (Repeats). Here we characterize the DNA sequences controlling DNA methylation at Rasgrf1 and identify the mechanism by which differential methylation regulates imprinted expression. We demonstrate that the repeats are required for methylation at the DMD during spermatogenesis but not during oogenesis providing the basis for parental specific methylation inheritance (Yoon, Herman et al. 2002). We demonstrate that it is the methylation at the DMD, which determines expression as aberrant acquisition of methylation reactivates the naturally silent maternal allele (Herman, Lu et al. 2003). We provide evidence that the DMD acts as a CTCF dependent methylation sensitive enhancer-blocking element. Methylation at the DMD excludes the binding of the enhancer blocker-binding protein CTCF hence allowing expression. The unmethylated DMD binds CTCF in vitro and in vivo and exhibits a functional enhancer blocking activity to silence expression (Yoon, Herman et al. 2005). Outside its endogeneous context, the repeats are sufficient for the establishment of parental specific methylation at the DMD upon transmission of an unmethylated transgene. The DMD and repeats are also sufficient for methylation reprogramming in the male germline. Reprogramming in the female germline however, seems to require negative signal(s) outside the DMD and the Repeats (Herman, Hu et al. 2005).