Analysis of CTCF Function in Neurodevelopment - Project Summary/Abstract Neurodevelopmental disorders (NDD) are a broad category of congenital neurological disease characterized by a wide range of cognitive and behavioral impairments. The etiology of NDD remains elusive, with genetic and epigenetic factors playing crucial roles in its pathogenesis. Recent advances in genomics have provided valuable insights into the role of dynamic chromatin reorganization in neurodevelopment and neurodevelopmental disorders. One key protein involved in chromatin organization is CTCF, which is known for its pivotal role in regulating gene expression by facilitating long-range chromatin interactions and global chromatin organization, and was recently identified as the causative gene for a rare NDD referred to as CTCF-related disorder (CRD). However, the precise mechanisms by which CTCF contributes to neural differentiation-dependent chromatin organization and NDD pathogenesis are poorly understood. The central aim of this project is to elucidate the function of CTCF in chromatin organization and transcriptional dynamics during neural differentiation by studying the molecular and cellular consequence of two CRD-associated CTCF mutations in human pluripotent stem cells (hPSCs), hPSC-derived neural progenitor cells (NPCs), and NPC-derived neural organoids. Preliminary data indicate that CTCF is a critical regulator of the dynamic transcriptional changes that occur during neural differentiation. Our research will employ a multi-pronged approach, utilizing both 2D and 3D models of human neural differentiation. We will use state-of-the-art chromatin conformation capture techniques such as Hi-ChIP and lamina-associated domain (LAD) mapping to examine the role of CTCF in shaping neural differentiation- dependent long-range chromatin interactions and nuclear architecture. Moreover, we will utilize neural organoids to investigate the impact of CTCF mutations on neuronal differentiation, maturation, and function. The outcomes of this research have significant implications for our understanding of CRD pathogenesis. By uncovering the role of CTCF in neural differentiation-dependent chromatin organization, we hope to identify potential mechanisms that contribute to the development of CRD and other neurodevelopmental disorders that stem from dysfunctional chromatin regulation. In summary, this pilot project grant application seeks funding to investigate the role of CTCF in neural differentiation and neurodevelopment. Through an integrated approach combining genomics, epigenomics, and functional analyses, this research aims to shed light on the intricate molecular mechanism underlying CRD and ultimately contribute to the development of targeted therapies and diagnostic tools for this complex neurodevelopmental disorder.
