Epigenetic and Neuronal Mechanisms of MECP2 Reactivation to rescue Rett Syndrome

PROJECT SUMMARY/ABSTRACT Rett syndrome (RTT) is a debilitating neurodevelopmental disorder without cure. RTT is characterized by cognitive impairment, seizures, and loss of intentional motor skills, and is caused by heterozygous mutations of the X-linked methyl-CpG-binding protein 2 (MeCP2) gene on the X chromosome. As female mammalian cells

undergo X chromosome inactivation (XCI), female patients with RTT are mosaic in terms of their MeCP2 expression; around half their cells express the wild-type allele and the other half express the loss-of-function mutant allele. Thus, reactivation of the silenced, functional MeCP2 allele may serve as a therapeutic treatment

for these patients with RTT. Current efforts to reactivate MeCP2 have focused on approaches to globally reactivate the inactive X chromosome. However, prior studies have shown that global reactivation of the inactive X is associated with adverse side effects, such as the development of hematologic and gastrointestinal

malignancies. Recently, we demonstrated that targeted reactivation of MeCP2 could be achieved efficiently by using a combination of CRISPR-based epigenome editing tools to demethylate the MECP2 promoter and insulate the MECP2 locus through recruitment of CTCF in vitro. However, whether this approach is sufficient to

rescue RTT phenotypes in vivo and the molecular mechanisms underlying MeCP2 reactivation remain unknown. This proposal seeks to address these questions using a diverse set of Mecp2 reporter systems. In Aim 1, I will perform multiplexed DNA methylation editing of Mecp2 in vivo and in vitro to identify the epigenetic

mechanisms underlying Mecp2 reactivation and to quantify the extent of reactivation in the mouse brain. In Aim 2, I will assess the motor function, hippocampal circuit function, cognitive function, and survival of edited RTT mice to determine if DNA methylation editing rescues RTT associated phenotypes in vivo. Together, these aims

will help elucidate the mechanisms that allow for Mecp2 reactivation from the inactive X chromosome and guide epigenetic approaches to treat RTT and other X-linked disorders. The training plan presented in this proposal will allow me to benefit from expert mentors and collaborators across Columbia University Irving

Medical Center, preparing me with the scientific and clinical training necessary to become an independent physician scientist studying biology of neurological disease.