MeCP2 binding to methylated chromatin is aided by nucleosome linker DNA interactions

Abstract

Methyl-CpG-binding protein 2 (MeCP2) is an abundant nuclear protein which preferentially binds methylated cytosines via its methyl-CpG binding domain (MBD) and causes gene repression through the recruitment of additional factors. MeCP2 is highly abundant in neurons and is essential for proper neuronal function, with mutations in the protein leading to the neurological disorder Rett syndrome. How MeCP2 interacts with methylated DNA in the context of chromatin is not well understood.

I took an in vitro biochemical approach to investigate the interaction of MeCP2 with methylated nucleosomes. I reconstituted nucleosomes with defined DNA methylation sites at specific nucleosome locations and assayed binding of purified MeCP2 proteins. Surprisingly, methylation position on a nucleosome had little effect on MeCP2 binding. Indeed, MeCP2 could even bind to a single methylated CpG site in bent core nucleosomal DNA. However, this binding was dependant on a previously uncharacterised central region of MeCP2, which provides additional DNA binding activity. I characterised this central region as a novel DNA binding module, made up of several small DNA binding motifs, which are important for MeCP2 function on nucleosomes. This activity is essential to stabilise nucleosome DNA methylation binding and suggests a requirement for MeCP2 to engage with linker regions between the nucleosome core particles.

I found that the nucleosome surface itself, histone tails or histone post translational modifications had little effect on MeCP2 interaction. However, removing or occluding nucleosome linker DNA, through the N-terminal tail of histone H3 or C-terminal tail of linker histone H1, disrupted the MeCP2-nucleosome interaction. This again highlighted the importance of nucleosome linker DNA to MeCP2 binding.

Overall, this research infers a mechanism where this novel central DNA binding region in MeCP2 interacts with linker DNA on nucleosomes, aiding the subsequent specific binding of the MBD to methylated nucleosomal DNA. It suggests that the MBD in isolation, or constructs lacking the central region such as the recent clinically developed MeCP2 ‘mini-gene’, would be insufficient to allow binding to DNA methylation at all sites in the genome.