ORCID

https://orcid.org/0000-0003-2339-2492

Date of Award

12-18-2024

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Molecular Genetics & Genomics)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

CTCF is a DNA-binding zinc finger protein that is responsible in large part for the 3-dimensional organization of the mammalian genome. Working in tandem with a number of protein partners, it functions to define looping anchors and coordinate DNA contacts within the 3-dimensional space of the nucleus. Heterozygous missense mutations with this protein cause a rare neurodevelopmental disorder known as CTCF-related disorder (CRD). Patients carrying these mutations present with a strikingly neurological spectrum of phenotypes including intellectual disability, microcephaly, autistic-like behaviors, and motor delay despite carrying the mutation in every cell in the body. Most of the previous in vivo modeling of CTCF dysfunction has been through conditional knockouts, despite the disease presenting in a heterozygous loss of function manner. In this thesis, I hypothesized that creating a knock-in mouse model of CRD would expand our knowledge not only of the disease state, but of CTCF’s important function in neurodevelopment. Here, I describe the use of the novel heterozygous CtcfR567W/+ mouse model to investigate the marked neurological consequences of CTCF dysfunction. First, through in vitro assays and investigation into the specific deficits of missense mutations found within CTCF, I hypothesized the functional consequences of specific missense mutations in CTCF, including examining the exact deficit induced by the CTCFR567W mutation. Next, through an in-depth battery of behavioral and motor assessments, I found that this mouse model displays motor, communication, and social deficits at both the juvenile and adult stages, successfully recapitulating phenotypes seen in humans. Moving onto the molecular level, I generated a large genomic database for the adult cortex of CtcfR567W/+ mice that includes RNA-sequencing, ChIP-sequencing, and HiC analyses. My transcriptional analysis found that the CtcfR567W/+ mutation specifically upregulated the expression of the longest genes in the genome, which are often neuron-specific genes. Importantly, this observation proved true in an independently generated homozygous CtcfR567W/R567W mouse model. Paired with this, I found that a subset of genes that work to define neuronal subtype are a particularly affected population in this set of upregulated genes. Analysis of CTCF binding in the cortex showcased that occupancy was decreased at or near long genes in a manner that is dependent on DNA methylation both in the mCG and mCA context. This data was also paired with a disorganization of the 3-dimensional genomic architecture at the level of loops while larger order TAD structure remained unaffected. Finally, immunofluorescent analysis of adult mouse brain structures led to the identification of improper hippocampal dentate gyrus morphology and loss of PV+ neuronal populations. These findings have broad implications for the study of CRD and for the role of CTCF specifically in the coordination of proper neurodevelopment.

Language

English (en)

Chair and Committee

Jason Yi

Committee Members

Andrew Yoo; Harrison Gabel; John Edwards; Tychele Turner

Available for download on Wednesday, December 16, 2026

Included in

Neurosciences Commons

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