Analysis of protein domains and Rett syndrome mutations indicate that multiple regions influence chromatin-binding dynamics of the chromatin-associated protein MECP2 in vivo

The methyl-CpG-binding protein 2 (MECP2) serves both organizational and transcriptional functions in the nucleus, with two well-characterized domains integrally related to these functions. The recognition of methylated CpG dinucleotides is accomplished by the methyl-binding domain (MBD), and the transcriptional repression domain (TRD) facilitates protein-protein interactions with chromatin remodeling proteins. For each known function of MECP2, chromatin binding is a crucial activity. Here, we apply photobleaching strategies within the nucleus using domain-deleted MECP2 proteins as well as naturally occurring point mutations identified in individuals with the neurodevelopmental disorder Rett syndrome (RTT). These studies reveal that MECP2 is transiently associated with chromatin in vivo and confirm a central role for the MBD in directing the protein to heterochromatin. In addition, we report for the first time that the small region between the MBD and the TRD, known as the interdomain region (ID), stabilizes chromatin binding by MECP2 independently of the MBD. The TRD of MECP2 also contributes towards chromatin binding, whereas the N- and C-termini do not. Some common RTT missense and nonsense mutations significantly affect binding kinetics, suggesting that alterations in chromatin binding can result in protein dysfunction and hence a disease phenotype.