Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.726397
Title: The roles of p53, p21, and RB in regulation of proliferation and apoptosis in hepatocytes
Author: Sheahan, Sharon
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2003
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Abstract:
Precise control of proliferation and apoptosis is essential for the prevention of cancer. Pathways involving p53, p21, and RB are central to the regulation of these processes, and are dysregulated in almost all human cancers. This thesis describes an investigation into the effects of inactivating components of the p53-p21-RB pathway in order to better understand the mechanisms underlying its role in preventing cancer. Hepatocytes isolated from transgenic mice bearing mutations in p53, p21, Rb, and combinations of all three, were employed to provide insight into the pathways controlling hepatocyte proliferation and apoptosis. In the case of the Rb mutation, Cre/LoxP conditional gene targeting was used to overcome the problem of embryonic lethality of Rb knock-out. In vitro Rb gene deletion mediated by adenovirus-Cre was successfully developed and characterised. This revealed that adenovirus-Cre-mediated gene deletion is extremely efficient in hepatocytes and that adenoviral infection had no affect on cell viability or growth. Expression of Cre, however, stimulated DNA synthesis while expression of LacZ adversely affected viability at higher MOI. These findings underline the need for careful characterisation of gene deletion systems employing both adenovirus and Cre recombinase. Analysis of hepatocytes deficient in p53, Rb, and p21, revealed severe defects in proliferation accompanied by an increase in ploidy, and nuclear and mitotic abnormalities. Analysis of hepatocytes deficient in combinations of all three genes provided insight into the level of interdependency between these genes, suggesting that there are two major pathways regulating hepatocyte proliferation: a p21-RB pathway that operates independently of p53, and a p53-p21 pathway that operates independently of RB. The p53-p21-RB pathway is critically important in regulating cellular responses to stress. The role of p53, p21, and RB in regulation of hepatocyte responses to DNA damage was investigated. It was found that each of these proteins is required for the maintenance, but not initiation, of UV-induced arrest. Both p53- dependent and -independent pathways of UV-induced apoptosis exist, and loss of Rb or p21 increased susceptibility to p53-independent, but not p53-dependent, apoptosis. In conclusion, the work presented in this thesis demonstrates that adenovirus-Cremediated gene deletion is extremely efficient in hepatocytes. It also shows that p53, p21, and RB are critical in regulating proliferation and apoptosis both in the absence and presence of exogenous stress, underlining the importance of these proteins in regulation of cell growth and hence presumably in prevention of cancer.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.726397  DOI: Not available
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