MeCP2 binds cooperatively to its substrate and competes with histone H1 for chromatin binding sites.

Sporadic mutations in the hMeCP2 gene, coding for a protein that preferentially binds symmetrically methylated CpGs, result in the severe neurological disorder Rett syndrome (RTT). In the present work, employing a wide range of experimental approaches, we shed new light on the many levels of MeCP2 interaction with DNA and chromatin. We show that strong methylation-independent as well as methylation-dependent binding by MeCP2 is influenced by DNA length.

Unique physical properties and interactions of the domains of methylated DNA binding protein 2.

Methylated DNA binding protein 2 (MeCP2) is a methyl CpG binding protein whose key role is the recognition of epigenetic information encoded in DNA methylation patterns. Mutation or misregulation of MeCP2 function leads to Rett syndrome as well as a variety of other autism spectrum disorders. Here, we have analyzed in detail the properties of six individually expressed human MeCP2 domains spanning the entire protein with emphasis on their interactions with each other, with DNA, and with nucleosomal arrays.

Rett syndrome-causing mutations in human MeCP2 result in diverse structural changes that impact folding and DNA interactions.

Most cases of Rett syndrome (RTT) are caused by mutations in the methylated DNA-binding protein, MeCP2. Here, we have shown that frequent RTT-causing missense mutations (R106W, R133C, F155S, T158M) located in the methylated DNA-binding domain (MBD) of MeCP2 have profound and diverse effects on its structure, stability, and DNA-binding properties. Fluorescence spectroscopy, which reports on the single tryptophan in the MBD, indicated that this residue is strongly protected from the aqueous environment in the wild type but is more exposed in the R133C and F155S mutations.

MeCP2-chromatin interactions include the formation of chromatosome-like structures and are altered in mutations causing Rett syndrome.

hMeCP2 (human methylated DNA-binding protein 2), mutations of which cause most cases of Rett syndrome (RTT), is involved in the transmission of repressive epigenetic signals encoded by DNA methylation. The present work focuses on the modifications of chromatin architecture induced by MeCP2 and the effects of RTT-causing mutants. hMeCP2 binds to nucleosomes close to the linker DNA entry-exit site and protects approximately 11 bp of linker DNA from micrococcal nuclease.

Multiple modes of interaction between the methylated DNA binding protein MeCP2 and chromatin.

Mutations of the methylated DNA binding protein MeCP2, a multifunctional protein that is thought to transmit epigenetic information encoded as methylated CpG dinucleotides to the transcriptional machinery, give rise to the debilitating neurodevelopmental disease Rett syndrome (RTT). In this in vitro study, the methylation-dependent and -independent interactions of wild-type and mutant human MeCP2 with defined DNA and chromatin substrates were investigated. A combination of electrophoretic mobility shift assays and visualization by electron microscopy made it possible to understand the different conformational changes underlying the gel shifts.

Organization of interphase chromatin.

The organization of interphase chromatin spans many topics, ranging in scale from the molecular level to the whole nucleus, and its study requires a concomitant range of experimental approaches. In this review, we examine these approaches, the results they have generated, and the interfaces between them. The greatest challenge appears to be the integration of information on whole nuclei obtained by light microscopy with data on nucleosome-nucleosome interactions and chromatin higher-order structures, obtained in vitro using biophysical characterization, atomic force microscopy, and electron microscopy.

Chromatin compaction by human MeCP2. Assembly of novel secondary chromatin structures in the absence of DNA methylation.

MeCP2 is a transcriptional repressor that contains an N-terminal methylated DNA-binding domain, a central transcription regulation domain, and a C-terminal domain of unknown function. Whereas current models of MeCP2 function evoke localized recruitment of histone deacetylases to specific methylated regions of the genome, it is unclear whether MeCP2 requires DNA methylation to bind to chromatin or whether MeCP2 binding influences chromatin structure in the absence of other proteins. To address these issues, we have characterized the complexes formed between MeCP2 and biochemically defined nucleosomal arrays.