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Title: Sequence stability of the APC gene : the role of DNA repair mechanisms in colon carcinogenesis
Author: Turnbull, Charlotte Louise
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2007
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To study the role of instability in colorectal cancer, this project focused on investigating any inherent sequence-specific mutation frequency of the APC gene. I used an approach where clonal selective advantage imparted by APC mutation is avoided, and have utilised a number of cancer and non-cancer cell lines, and patient tissue samples. As deficient DNA repair has been implicated in a number of CRC conditions, I have also investigated the effect of defective DNA mismatch repair (MMR) and base exclusion repair (BER) on APC mutation frequency and spectrum. In addition, I have also begun to develop a novel technique for directly measuring BER activity in live cells. I show substantial elevation of mutation frequency in the APC gene compared to control gene sequences. This is the first demonstration of inherent sequence instability of APC in normal cells. Polymorphism and evolutionary data indicate that APC is likely to be subject to a high neutral mutation rate and purifying selection. Furthermore, detailed analysis of de novo APC mutations has not implicated the involvement of any alternative mutational mechanisms such as CpG island methylation. Further work is therefore required to understand the sequence instability observed in these normal cells. In the cell lines studied, mutation frequency at all gene sequences was increased in the defective BER background and in heterozygous MMR cells. Detectable APC gene mutation frequency was substantially reduced in cell lines null for MMR. In patient samples a striking effect of Germline MUTYH mutation on APC mutation spectrum was observed in tumour samples, and the consequence of the bi-allelic inactivation of MUTYH on somatic inactivating mutations in APC and K-ras was clear. The findings presented in this thesis provide new understanding of early molecular events during tumourigenesis and how they are affected by defects in DNA repair systems, in particular BER and MMR.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available