MBD2 is a critical component of a methyl cytosine-binding protein complex isolated from primary erythroid cells

The chicken embryonic beta-type globin gene, p, is a member of a small group of vertebrate genes whose developmentally regulated expression is mediated by DNA methylation. Previously, we have shown that a methyl cytosine-binding complex binds to the methylated p-globin gene in vitro. We have now chromatographically purified and characterized this complex from adult chicken primary erythroid cells. Four components of the MeCP1 transcriptional repression complex were identified: MBD2, RBAP48, HDAC2, and MTA1. These 4 proteins, as well as the zinc-finger protein p66 and the chromatin remodeling factor M, were found to coe-lute by gel-filtration analysis and pull-down assays. We conclude that these 6 proteins are components of the MeCPC. In adult erythrocytes, significant enrichment for MBD2 is seen at the inactive p-globin gene by chromatin immunoprecipitation assay, whereas no enrichment is observed at the active beta(A)-globin gene, demonstrating MBD2 binds to the methylated and transcriptionally silent p-globin gene in vivo. Knock-down of MBD2 resulted in up-regulation of a methylated p-gene construct in mouse erythroleukemic (MEL)-p cells. These results represent the first purification of a MeCP1-like complex from a primary cell source and provide support for a role for MBD2 in developmental gene regulation.