Structural Basis of MeCP2 Distribution on Non-CpG Methylated and Hydroxymethylated DNA. Issue 10 (19th May 2017)

• Record Type: Journal Article
• Authors: Sperlazza, M. Jeannette; Bilinovich, Stephanie M.; Sinanan, Leander M.; Javier, Fatima R.; Williams, David C.
• Is Part Of: Journal of molecular biology. Volume 429:Issue 10(2017)
• Page Start: 1581
• Page End: 1594
• Publication Date: 2017-05-19
• Subjects: MeCP2 methyl-CpG binding protein 2 -- mCG 5-methylcytosines in CpG -- mCH non-CG methylated sites -- hmC hydroxymethylcytosine -- mCA methylated CpA -- hmCA hydroxymethylated CpA -- MBD methyl-CpG-binding domain -- ITC isothermal titration calorimetry -- CSP chemical shift perturbation -- HSQC heteronuclear single-quantum correlation -- BMRB Biological Magnetic Resonance Data Bank -- CPMG Carr-Purcell-Meiboom-Gill -- BDNF brain-derived neurotrophic factor; epigenetics -- Rett syndrome -- methyl–binding domain -- protein–DNA interaction -- relaxation dispersion; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2017.04.009
• Languages: English
• ISSNs: 0022-2836

Abstract

Abstract: The Rett-syndrome-associated methyl-CpG-binding protein 2 (MeCP2) selectively binds methylated DNA to regulate transcription during the development of mature neurons. Like other members of the methyl-CpG-binding domain (MBD) family, MeCP2 functions through the recognition of symmetrical 5-methylcytosines in CpG (mCG) dinucleotides. Advances in base-level resolution epigenetic mapping techniques have revealed, however, that MeCP2 can bind asymmetrically methylated and hydroxymethylated CpA dinucleotides and that this alternative binding selectivity modifies gene expression in the developing mammalian brain. The structural determinants of binding to methylated CpA (mCA) and hydroxymethylated DNA have not been previously investigated. Here, we employ isothermal titration calorimetry and NMR spectroscopy to characterize MeCP2 binding to methylated and hydroxymethylated mCG and mCA DNA, examine the effects of Rett-syndrome-associated missense mutations, and make comparisons to the related and evolutionarily most ancient protein, MBD2. These analyses reveal that MeCP2 binds mCA with high affinity in a strand-specific and orientation-dependent manner. In contrast, MBD2 does not show high affinity or methyl-specific binding to mCA. The Rett-associated missense mutations (T158M, R106W, and P101S) destabilize the MeCP2 MBD and disrupt the recognition of mCG and mCA equally. Finally, hydroxymethylation of a high-affinity mCA site does not alter the binding properties, whereas hemi-hydroxylation of the equivalent cytosine in an mCG site decreases affinity and specificity. Based on these findings, we suggest that MeCP2 recognition of methylated/hydroxymethylated CpA dinucleotides functions as an epigenetic switch redistributing MeCP2 among mCG and mCA loci. Graphical Abstract: Highlights: MeCP2 binding of mCA is high-affinity, strand-specific, orientation-dependent, and unique among the MBD-containing proteins. MeCP2 bound to methylated DNA displays dynamic motion at the protein–DNA interface that extends out the core α-helix. Hydroxymethylation of cytosine shifts the distribution of MeCP2 proteins from mCG to mCA loci.