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    Investigating the structure and architecture of the NuA4 subcomplex TINTIN

    Aciyan, Emir (2021) Investigating the structure and architecture of the NuA4 subcomplex TINTIN. PhD thesis, Birkbeck, University of London.

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    The regulation of gene expression is essential for the correct function of virtually all cellular processes and misregulation has a range of pathological consequences, including Alzheimer’s disease and cancer. The NuA4 coactivator complex is an essential eukaryotic histone acetyltransferase, that promotes transcriptional activation and DNA repair. The complex targets the N-terminal tail of H4 and H2A, recruited to specific loci by DNA-binding transcriptional activators. NuA4 is highly modular, with several subcomplexes that can exist independently, or shared with other coactivator complexes such as SAGA, allowing for targeted acetylation of specific gene loci. An investigation of the structure and function of the NuA4 subcomplex TINTIN, is presented herein, a trimeric module comprised of the subunits Eaf3, Eaf5 and Eaf7. TINTIN has been implicated in promoting transcription elongation, via a proposed interaction with RNA polymerase II and regulating nucleosome stability. TINTIN has been recombinantly expressed, purified and subjected to crystallography trials, which failed, despite multiple truncated TINTIN constructs that produced complexes of higher stability and purity. Nevertheless, these constructs provided insights into TINTIN architecture and identified key determinants for complex stability and major differences with the orthologous complex found in humans. A combination of SEC-MALS, native mass spectrometry and small-angle X-ray scattering analysis, provide low-resolution information, regarding the overall topology and oligomeric state of the complex. TINTIN exists predominantly in its heterotrimeric state, though the interaction between Eaf5 and the rest of the complex is relatively labile. The complex adopts an extended conformation, with some indications of partial flexibility/disorder. Additional factors may be required to stabilise TINTIN, such as subunits of the parent NuA4 complex and/or the RNA Pol II CTD, which pull-down experiments presented herein, suggest directly interacts with the complex.


    Item Type: Thesis
    Copyright Holders: The copyright of this thesis rests with the author, who asserts his/her right to be known as such according to the Copyright Designs and Patents Act 1988. No dealing with the thesis contrary to the copyright or moral rights of the author is permitted.
    Depositing User: Acquisitions And Metadata
    Date Deposited: 07 Sep 2021 11:59
    Last Modified: 01 Jul 2024 06:26


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