Single-molecule dissection of the interaction between a methyl-DNA binding protein and chromatin.

Abstract

The methyl-CpG binding protein 2 (MeCP2) is a prominent DNA methylation reader that is essential for proper gene expression programming and normal functioning of nerve cells. Mutations in MeCP2 cause Rett syndrome, a severe neurological disorder mainly affecting females and characterized by psychiatric and motor regression at 6-18 months. Despite decades of efforts, there are currently no approved therapies that directly target MeCP2 loss of function. One primary reason for the lack of MeCP2-targeted interventions is an inadequate understanding of how MeCP2 reads methylated DNA within eukaryotic chromatin. To address this knowledge gap, we developed an experimental platform that combines optical tweezers and single-molecule fluorescence microscopy to directly visualize the dynamic behavior of MeCP2 on chromatin. This assay enabled us to discover unexpectedly that nucleosomes serve as higher affinity substrates for MeCP2 than CpG methylated DNA, challenging the canonical view of MeCP2 as purely a methyl-CpG DNA reader. Instead, MeCP2 recognizes methylated DNA sites only after exhausting all available nucleosome sites. We further showed the high-affinity MeCP2-nucleosome interaction is dependent on MeCP2's disordered C-terminal domain. In addition, we demonstrated counterintuitively that MeCP2 and linker histone H1—two highly abundant nuclear proteins—can occupy the same nucleosome sites. Taken together, our results indicate that nucleosomes heavily regulate the availability and function of MeCP2 in the nucleus. Moreover, this work paves the way for mechanistic interrogation of Rett syndrome mutations and their impact on MeCP2 function at chromatin.