ASXL3 Links Chromatin Biology to Neurodevelopment Disorders

ABSTRACT/PROJECT SUMMARY Neurogenetic studies have identified pathogenic variants in genes that encode chromatin components, highlighting chromatin’s importance in corticogenesis. Dynamic regulation of histone modifications is critical for the transcriptional plasticity required during cellular differentiation. One such modification is the mono- ubiquitination of histone H2A (H2AUb1), a conserved, traditionally repressive histone mark. H2AUb1 is ligated by the Polycomb Repressive Complex 1 (PRC1) and removed by the Polycomb Repressive Deubiquitinase (PR- DUB) complex. PR-DUB is composed of the ubiquitin hydrolase BAP1 and one of three ASXL-family members (ASXL1-3), all of which target BAP1 hydrolase activity to discrete genomic loci. However, each ASXL protein is associated with a clinically distinct neurodevelopmental disorder, suggesting non-redundant functions in the regulation of H2AUb1 occupancy and transcriptional regulation. We identified de novo dominant truncating variants in ASXL3 as the genetic basis of both Bainbridge Ropers Syndrome (BRS) and autism spectrum disorder (ASD). Primary BRS patient cells display a dysregulation of H2AUb1 that is mirrored by mouse and in vitro disease models, denoting it as a key molecular pathology. The goal of this project is to uncover nonredundant ASXL3-dependent H2AUb1 functions in neurodevelopment, which will elucidate important functions in development and neuropathology. Using neural progenitor cells differentiated from human pluripotent stem cells (hPSCs) harboring clinically relevant ASXL variants, I will elucidate loci of differential H2AUb1 occupancy and transcriptional regulation important for ASXL3 pathology (Aim 1). To identify the developmental mechanism of BRS, I will use a combination of traditional immunohistochemistry and single-cell RNA sequencing analysis of dorsal forebrain organoids (Aim 2). This proposal will elucidate how H2AUb1 occupancy patterns are regulated by ASXL3 during corticogenesis. Such knowledge will uncover an important aspect of epigenetic regulation that is poorly understood. Additionally, knowing how these mechanisms unfold will give us targets with which to modulate them, the first step in developing therapeutic strategies for ASXL3- specific pathologies such as ASD and BRS.