The role of the MYST lysine acetyltransferase TIP60 in human cells and mice
Document TypePhD thesis
Access StatusThis item is embargoed and will be available on 2023-02-16.
© 2020 Johannes Wichmann
Histone acetylation affects the way DNA and associated proteins are packaged in the cell nucleus and regulate chromatin organisation and gene expression. Acetylation of core histones has been broadly correlated with initiating and maintaining open chromatin, poised or active gene transcription, DNA damage repair, as well as chromosome decondensation during mitosis and meiosis. The acetylation of lysine residues is catalysed by histone lysine acetyltransferases (KATs) and deacetylases (HDACs), which are tightly regulated. Dysregulation of KATs and aberrant lysine acetylation has been associated with tumorigenesis and negative prognoses in a wide range of cancer, presenting a new area of potential therapeutic targets. Potential acetylation targets of KATs catalysing the acetylation of histones have predominantly been studied in cell-free assays, where the enzymes show little substrate specificity. In contrast, histone acetyltransferases appear to acetylate surprisingly specific residues in whole cells. In this thesis I investigate the effects of acute deletion of the MYST lysine acetyltransferase TIP60 (KAT5) using inducible cre-recombinase and CRISPR/Cas9-mediated deletion in human cells and mouse cells, as well as mouse embryos and its potential role in cancer cells. I found that loss of TIP60 caused complete cell growth arrest in human and mouse cells. In the absence of TIP60 cells displayed cell cycle arrest in G1 and G2/M phase with increased endoreplication, accompanied by chromosomal segregation defects. Remarkably, the proliferation arrest caused by loss of TIP60 also occurred in the absence of the tumour suppressors p53, INK4A and ARF and therefore was independent of these. In contrast, cell survival was not affected. Growth arrest independent of major tumour suppressors flags TIP60 a potential target for novel cancer therapeutics. TIP60 was found to be essential for of H2AZ acetylation, particularly, lysine 7 acetylation. In contrast, global chromatin bound H2AZ levels were not reduced. H2A and H4 acetylation was reduced slightly in TIP60 depleted cells. Identifying H2AZ lysine 7 acetylation as a biomarker for TIP60 activity is a major step in developing TIP60 as a drug target. The mRNA levels of 6236 human and 8238 mouse genes, including many metabolic genes, were dependent on TIP60, supporting a role for TIP60 as a key transcriptional co-activator. Characterization of key mechanisms causing chromosomal aberrations and identifying approaches that can be used to reduce likelihood of genome instability due histone acetylation defects will pave the way for better detection of early changes in cancer and development of therapeutic applications in the future. This work represents important steps towards the development of histone lysine acetyltransferases as drug targets. A comprehensive analysis of histone acetylation activity of TIP60 shines light on the potential of its many proposed roles.
KeywordsTIP60, KAT5, HTATIP, MYST acetyltransferase, p53, INK4A-ARF, H2A.Z, acH2A.Z, histone acetylation
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