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Identification of regulatory pathways controlling histone H3.3 deposition in the central nervous system (CNS)

Liang, Qingyi; (2019) Identification of regulatory pathways controlling histone H3.3 deposition in the central nervous system (CNS). Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Studies have implicated the importance of the histone H3 variant H3.3 in chromatin remodelling and transcriptional regulation in the central nervous system (CNS). Gain-of-function mutations affecting H3.3 and its loading machinery have been recently identified in glioblastoma multiforme (GBM) and other tumours. Moreover, H3.3 accumulates on chromatin during ageing and plays significant roles in ageing-related signalling pathways, as recently demonstrated in the context of Caenorhabditis elegans. Given H3.3 involvement in human neoplasia and ageing, there is a need to better understand how H3.3 loading is regulated in cells and how pharmacological intervention could be used to either augment or suppress its incorporation. Loss-of-function mutations of H3.3 chaperone DAXX (death domain–associated protein 6) lead to chromatin instability and alternative lengthening of telomeres (ALT), correlating with a variety of cancers including GBM and pancreatic neuroendocrine tumours (pNETs). Studying DAXX interactome would improve our knowledge on its novel cellular functions involved in normal biological processes as well as tumourigenesis. During my PhD project, I developed an H3.3 visualisation platform based on Halo-Tag technology in Neuro2a cells (a neuroblastoma cell line). The Halo-Tag visualisation assay was developed via leading-edge robotised automation, rendering it suitable for high-content/drug screening. I also established a FLAG-Daxx affinity purification system in mouse neural progenitor cells (NPCs). Mass spectrometry of FLAG-Daxx affinity purification revealed two novel Daxx-interacting partners, Nr2c2 (nuclear receptor subfamily 2 group C member 2; Tr4, testicular orphan nuclear receptor 4) and Cpsf2 (cleavage and polyadenylation specificity factor 2), which were verified by protein immunoblotting. Overall, this study provides: i) a new tool for discovery science efforts aimed at identifying novel actionable regulators of H3.3 deposition in cells in the context of neoplastic disease and ageing, and ii) a basis for further investigation of mechanisms regulating histone variant loading in the nervous system.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Identification of regulatory pathways controlling histone H3.3 deposition in the central nervous system (CNS)
Event: UCL (University College London)
Language: English
Additional information: Copyright © The Author 2019. Original content in this thesis is licensed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0) Licence (https://creativecommons.org/licenses/by/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request.
UCL classification: UCL > Provost and Vice Provost Offices
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Cancer Institute
URI: https://discovery.ucl.ac.uk/id/eprint/10072501
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