MeCP2 binds to nucleosome free (linker DNA) regions and to H3K9/H3K27 methylated nucleosomes in the brain

Abstract

Methyl-CpG-binding protein 2 (MeCP2) is a chromatin-binding protein that mediates transcriptional regulation, and is highly abundant in brain. The nature of its binding to reconstituted templates has been well characterized in vitro. However, its interactions with native chromatin are less understood. Here we show that MeCP2 displays a distinct distribution within fractionated chromatin from various tissues and cell types. Artificially induced global changes in DNA methylation by 3-aminobenzamide or 5-aza-2′-deoxycytidine, do not significantly affect the distribution or amount of MeCP2 in HeLa S3 or 3T3 cells. Most MeCP2 in brain is chromatin-bound and localized within highly nuclease-accessible regions. We also show that, while in most tissues and cell lines, MeCP2 forms stable complexes with nucleosome, in brain, a fraction of it is loosely bound to chromatin, likely to nucleosome-depleted regions. Finally, we provide evidence for novel associations of MeCP2 with mononucleosomes containing histone H2A.X, H3K9me2 and H3K27me3 in different chromatin fractions from brain cortex and in vitro. We postulate that the functional compartmentalization and tissue-specific distribution of MeCP2 within different chromatin types may be directed by its association with nucleosomes containing specific histone variants, and post-translational modifications.

Phosphorylation of Distinct Sites in MeCP2 Modifies Cofactor Associations and the Dynamics of Transcriptional Regulation

“…3C). This is in agreement with recently published results showing that the majority of MeCP2 from brain and liver is also found in the S1 fraction (37). There were no apparent differences in the distribution of total and phosphorylated MeCP2e1-FLAG between the fractions, as all MeCP2 forms were predominantly found in the active S1 fraction along with the known MeCP2 interacting protein Sin3A (25), while the MeCP2-interacting protein HP1␣ (1) was present in both the S1 and S2 fractions (Fig.…”

Rett syndrome: a complex disorder with simple roots

“…Finally, in vitro crosslinking experiments suggest that contact occurs between MeCP2 and nucleosomal histone H3 (REF. 68), and it has also been reported that MeCP2 can bind to an isolated N-terminal histone H3 tail 69 . Whether binding to histones represents a mechanism for recruiting MeCP2 to chromatin in vivo has yet to be assessed.…”

The characterization of macroH2A beyond vertebrates supports an ancestral origin and conserved role for histone variants in chromatin

“…In this very coarse fractionation, 71,72 the SI (supernatant) fraction recovered immediately upon nuclease digestion contains digested DNA and nucleosomes from chromatin regions readily accessible to the nuclease (i.e., euchromatic regions). The SI fraction was subsequently fractionated further using a 5-20% sucrose gradient in 25 mM NaCl, 10 mM Tris-HCl (pH 7.5), 0.2 mM EDTA buffer run for 21 h at 96,000 £ g at 4 C. The SE (supernatant) fraction, obtained after hypotonic lysis of the pelleted nuclei, is highly enriched in facultative heterochromatin.…”