Disruption of UHRF1 expression in chicken DT40 cells

Sun, Huaping and Lindsay, Howard and Taylor, Elaine (2017) Disruption of UHRF1 expression in chicken DT40 cells. PhD thesis, Lancaster University.

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Epigenetics regulate the gene expression while imposing no change in the underlying gene sequence constitution. The abnormal regulation of epigenetics is associated with the development and progression of cancer. Among the epigenetic regulators, UHRF1 (ubiquitin-like with PHD and ring finger domain containing protein 1) has attracted considerable attentions in cancer research in past years due to its universally increased expression in a wide range of different cancer cells, and its ability to facilitate the crosstalk between DNA methylation and histone modification, thereby driving the occurrence of cancer and ensuring the inheritance of accurate epigenomic information to descendent cells. Depletion of UHRF1 proteins in nuclear significantly blocked DNA replication in Xenopus egg extracts. However, the mechanism underlying the role of UHRF1 in DNA replication remains unclear. Considering the effects of UHRF1 depletion on DNA replication is independent of cell cycle checkpoints and transcription events in Xenopus egg extracts, it is important to extend the work into a higher eukaryote system. Therefore, we planned to generate UHRF1 knockout cell line in the most effective gene targeting system of DT40 cells to observe the effect of UHRF1 deletion on DNA replication and the cellular response to DNA damage. CRISPR/Cas9 technology and traditional gene targeting were applied to generate conditional UHRF1 knockout cell lines. It was found that knocking out the expression of both alleles of UHRF1 was lethal to cell viability and that there was a threshold of UHRF1 overexpression that can be tolerated by DT40 cells. Additionally, the ability of DT40 cells to tolerate DNA damage was positively related to the expression levels of UHRF1 proteins.

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Thesis (PhD)
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05 Dec 2017 09:46
Last Modified:
16 Jul 2024 05:40