Neuronal non-CG methylation is an essential target for MeCP2 function

Abstract

<jats:title>SUMMARY</jats:title><jats:p>DNA methylation is implicated in neuronal biology via the protein MeCP2, mutation of which causes Rett syndrome. MeCP2 recruits the NCOR1/2 corepressor complexes to methylated cytosine in the CG dinucleotide, but also to non-CG methylation, which is abundant specifically in neuronal genomes. To test the biological significance of its dual binding specificity, we replaced the MeCP2 DNA binding domain with an orthologous domain whose specificity is restricted to mCG motifs. Knock-in mice expressing the domain-swap protein displayed severe Rett syndrome-like phenotypes, demonstrating that interaction with sites of non-CG methylation, specifically the mCAC trinucleotide, is critical for normal brain function. The results support the notion that the delayed onset of Rett syndrome is due to the late accumulation of both mCAC and its reader MeCP2. Intriguingly, genes dysregulated in both <jats:italic>Mecp2</jats:italic>-null and domain-swap mice are implicated in other neurological disorders, potentially highlighting targets of particular relevance to the Rett syndrome phenotype.</jats:p>

Evidence that direct inhibition of transcription factor binding is the prevailing mode of gene and repeat repression by DNA methylation

“…Importantly, these observations are compatible with other proposed functions of MBD proteins-in particular MeCP2-in gene regulation, such as impacting transcriptional elongation by methyl-CA binding 65,81,82 , alternative splicing 83,84 , microRNA processing 85 or protecting CA repeats from nucleosome invasion 86 . While MBD proteins can have a repressive function-in particular when recruited to certain sites or at transfected reporter plasmids [17][18][19][87][88][89][90] -our experiments argue against functional redundancy between the four tested MBD proteins as a reason for the absence of more severe transcriptional phenotypes, as hypothesized in previous loss-of-function studies of selected MBD proteins 19,36 .…”

Epigenomic complexity of the human brain revealed by single-cell DNA methylomes and 3D genome structures

“…Both CG- and CH-methylation (mCG and mCH, respectively) are highly dynamic during brain development and show remarkable cell type specificity ( 1, 5, 6 ). mCG and mCH are both essential for gene regulation and brain functions ( 7 ). In addition to DNA methylation, gene expression also requires proper conformation of chromatin folding (3D chromatin conformation), which is organized into active or repressive chromatin compartments, topologically associating domains (TADs) and chromatin loops ( 8 ).…”

Quantitative single cell 5hmC sequencing reveals non-canonical gene regulation by non-CG hydroxymethylation

“…Given that asymmetric CH sites are much more abundant (∼1.1 billion in mouse genome) than CG sites (∼22 million symmetric CpG dyads) 32 , even a small fraction of modified 5hmCH sites (0.2-0.6%) may lead to a considerable impact on epigenetic gene regulation. Although the exact mechanism downstream of 5hmCH is currently unclear, this non-canonical gene regulatory function of 5hmCH is reminiscent of the repressive role of 5mCH 33 and may possibly be mediated by recruiting transcriptional repressor MeCP2, as previously predicted by in vitro biochemical binding assays 12, 34 . Intriguingly, a recent study of maturing cerebellar Purkinje neurons also shows that accumulation of both 5mCH and 5hmCH (measured by bulk BS-seq and oxBS-seq assays) within gene body of a cohort of developmentally down-regulated genes is accompanied by elevated level of repressive histone modification H3K27me3 (deposited by Polycomb repression complexes) 35 , suggesting an alternative but non-mutually exclusive transcriptional inhibition mechanism mediated by genic 5mCH and 5hmCH.…”