The role of MBD1, a methyl-CpG binding protein, plays in methylation -mediated transcriptional silencing and DNA repair
DNA promoter hypermethylation has been shown to be a functional mechanism of transcriptional repression. This epigenetic gene silencing is thought to involve the recruitment of chromatin remodeling factors, such as histone deacetylaces (HDACs), to methylated DNA via a family of proteins termed methyl-CpG binding proteins (MBDs). MBD1, a member of this family had been shown to exhibit transcriptional repressive activity, but no interacting protein partners had been identified.
Through the use of a yeast two-hybrid screen an interaction between MBD1 and the p150 subunit of CAF-1 was identified. This interaction was further characterized demonstrating that these two proteins also form a multi-protein complex with HP1α in co-immunoprecipitation assays. The MBD1/CAF-1 p150 interaction requires the methyl-CpG binding domain of MBD1 and the association occurs in the C-terminus of CAF-1 p150. Immunofluorescent confocal imaging revealed that both proteins co-localize to regions of dense heterochromatin in mouse cells, and over-expression of the C-terminus of CAF-1 p150 prevents the targeting of MBD1 in these cells without disrupting global heterochromatin structure. This interaction suggests a role for MBD1 and CAF-1 p150 in methylation-mediated transcriptional repression and the inheritance of epigenetically determined chromatin states.
In another yeast two-hybrid screen, thymine DNA glycosylase (TDG) was discovered as an interacting partner with MBD1. Employing immunoprecipitation studies, MBD1 was shown to associate with TDG in the cell, and this interaction requires the methyl-CpG binding domain region of MBD1 in yeast two-hybrid assays. TDG fails to co-localize with MBD1 to regions of densely methylated heterochromatin in mouse cells.
The results suggest that TDG does not exhibit significant transcriptional repressive activity in transient transcription assays but when partnered with MBD1 forms a potent repressive complex. The glycosylase may act to recruit MBD1 to regions of DNA damage to reestablish repressive heterochromatin at locations where higher order chromatin structure is required for the proper repair of the DNA lesion. This finding provides further evidence that the role of methyl-CpG binding proteins as transcriptional repressors may not be solely as silencers of hypermethylated genes.