Abstract
dc:description.abstractCHD4 (chromodomain helicase DNA-binding protein 4) is a widely conserved ATP-dependent chromatin remodeler essential for early mammalian development. Although best known for its role in gene regulation as a core component of the NuRD complex, CHD4 is also found within the ChAHP complex, which contributes to silencing of transposable elements (TEs). CHD4 generally functions to increase nucleosome density, however the specific chromatin remodelling mechanisms employed by NuRD and ChAHP to regulate gene expression and repress repetitive elements have not been fully characterised. Here, I used auxin-inducible depletion of CHD4 in mouse embryonic stem cells (ESCs) together with multi-omic approaches to dissect its role in the regulation of chromatin accessibility and transcription factor (TF) binding, providing insights into its impact on distinct transcriptional outcomes. I demonstrated that CHD4 performs two essential functions within the NuRD complex. It restricts chromatin accessibility across broad genomic regions to prevent non-specific TF binding. At active enhancer and promoter regions CHD4 maintains chromatin accessibility, ensuring these regulatory sites remain open, promoting precise and controlled transcriptional regulation. This dual regulatory role is further reflected in its influence on TF binding, where CHD4 depletion leads to altered binding patterns of NANOG and SOX2. I provided evidence that changes in TF binding often precede changes in chromatin accessibility, indicating that CHD4 directly limits TF binding in addition to indirectly controlling it by modulating chromatin structure. These results provide mechanistic insights into how CHD4 orchestrates transcriptional regulation within the NuRD complex. Characterisation of CHD4 binding sites together with ADNP demonstrated that these two proteins share many binding sites that extend beyond the known role of ChAHP at B2 SINEs. ADNP and CHD4 are found at repetitive elements of various classes as well as at regulatory regions, including promoters and active enhancers. Using the dTAG depletion system for ADNP and the auxin depletion system for CHD4 followed by ATAC-seq and RNA-seq revealed that the ChAHP complex silences evolutionary young repetitive elements by reducing chromatin accessibility. Both ADNP and CHD4 are required for transcriptional repression of B2 SINEs and all three proteins-CHD4, ADNP and HP1-are required for restricting chromatin accessibility at B2 SINEs. A novel role for ADNP in maintaining enhancer accessibility by recruiting CHD4 at enhancer regions was discovered. Simultaneous presence of NuRD and ChAHP at these enhancer elements suggested a functional redundancy between the two complexes, where the NuRD complex compensates for the loss of ChAHP. I propose that it is this function at enhancers that leads to aberrant gene expression of genes during neural development. Moreover, the data revealed that ADNP binding was redistributed across the genome in the absence of CHD4 suggesting that CHD4 restricts ADNP binding. Additionally, the ADNP/CTCF competition model at B2 SINEs is not supported, instead, evidence indicates that CTCF binding is regulated by chromatin accessibility. These findings provide a detailed picture of genome wide regulation of gene and TEs expression by CHD4, offering insights into the distinct and overlapping roles of CHD4 within the NuRD and ChAHP complexes.
Degree
thesis:*- Name dc:type.qualificationname
- Doctor of Philosophy (PhD)
- Level dc:type.qualificationlevel
- Doctoral
- Grantor dc:publisher.institution
- University of Cambridge
- Year dc:date.issued
- 2024
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Koulle, Antria
- Advisor dc:contributor.advisor
-
- Hendrich, Brian
Subjects
dc:subject × 6Rights
dc:rightsIdentifiers
dc:identifier.*- DOI dc:identifier.doi
- https://doi.org/10.17863/CAM.120184
- OAI identifier oai:identifier
- oai:www.repository.cam.ac.uk:1810/387377