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Title: Development of novel mouse models to study the p53 tumour suppressor
Author: Crawford, Kerryanne
ISNI:       0000 0004 2732 331X
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2011
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TP53 is the most commonly mutated gene in all human cancers [Vogelstein, 1990] and co- ordinates many of the cellular responses to DNA damage. It is vitally important therefore that we gain a better understanding of the function and regulation of p53. Since a major consequence of p53 mutation is cancer, a disease of complex multicellular organisms, the best system to study the real complexities of p53 function and regulation is in vivo. Accordingly we have proposed to use gene targeting in ES cells to create genetically altered strains of mice to enable a range of studies of p53 regulation and function. Given the importance of p53 in human disease processes, it is perhaps surprising that there have been relatively few p53 reporter mouse strains described. Heretofore few p53 reporter mouse strains have been described and these have been based upon un-targeted, often artificial, repeated p53 response elements in constructs that integrate at random [Komarova et aI., 1997, Gottlieb et aI., 1997, Vasey et aI., 2008, Briat and Vassaux, 2008]. Such random integration is known to be associated with loss of fidelity resulting from positional effects and thus it is unlikely that any of these provide authentic reporting of p53 activity. The ideal solution might be to target a p53 responsive gene using a knock-in approach, preferably in a gene that was exclusively activated by p53, eliciting a specific response program (cell cycle arrest/senescence/apoptosis) and which also does not display any haploinsufficiency. This project was based on such a strategy. In order to study p53 activation and consequence(s) in the gastrointestinal tract we aim to generate novel transgenic mice which would contain fluorescent reporters in the downstream target genes of p53 activation in both of p53s main responses to DNA damage (apoptosis and cell cycle arrest). The resulting dual apoptosis/cell cycle arrest p53-reporter mice would allow a unique opportunity to visualise not only that p53 was activated but what responses p53 had induced. Furthermore, as visual technology advances these mice, or second generation mice with newer fluorescent proteins with improved bioimaging properties (such as increased tissue penetration and brightness), could also be used in real-time analyses of p53 responses to DNA damage or other stimuli, as well as for long term monitoring of gene-regulation during cancer development. In addition, we aim to address fundamental questions regarding the regulation of p53 again using such gene targeting technology for in vivo studies. Importantly it is not yet known whether the p53-mediated up-regulation of its essential negative regulator Mdm2 is absolutely required for viability. Therefore we intend to make two transgenic mouse strains which lack either the Mdm2 P1 (constitutive) promoter or the p53 RE sites within the Mdm2 P2 (inducible) promoter. Studies of these mice are likely to have a profound impact on the field of cancer biology and normal development.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral