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Mechanisms of resistance to DNA methylation damage in human cells

Hampson, Richard John; (1997) Mechanisms of resistance to DNA methylation damage in human cells. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Methylating agents such as N-methyl-N-nitrosourea (MNU) kill cells by introducing-O6-methylguanine (06-meGua) into DNA. Cells may avoid its cytotoxic effects by at least two mechanisms: increasing cellular levels of O6-methylguanine DNA methyltransferase (MGMT), an enzyme capable of removing the methyl group from 06-meGua, and DNA methylation damage tolerance, whereby O6-meGua persists in the DNA but is not cytotoxic. Such methylation tolerance can result from DNA mismatch repair defects; the lethality of 06-meGua is thought to be due to processing by the mismatch repair system. The work presented in this thesis is concerned with resistant variants of the methylation damage sensitive Burkitts Lymphoma cell line RajiMex-. I have shown that RajiF12, a methylation tolerant variant of RajiMex- defective in a mismatch recognition activity, has a selective genome instability; this is consistent with a defect in one of the components of the mismatch recognition complex. I have also generated a new panel of resistant RajiMex- variants using two distinct selection procedures (acute and chronic exposure to MNU). I have studied the biochemical changes underlying this methylation resistance. Of 15 cell lines characterised, three had acquired resistance through increasing MGMT expression. Six lines were methylation tolerant but phenotypes characteristic of mismatch repair defective cells could not be ascertained; in a further six lines properties associated with mismatch repair defects were detected. My results indicate that (a) under certain conditions MNU may have epimutagenic effects, here leading to re-expression of MGMT (b) methylation tolerance can arise through loss of function of more than one mismatch repair component and (c) tolerance can be acquired in absence of any detectable mismatch repair defects. In addition I describe the identification of frequent frameshift mutations characteristic of mismatch repair defects in a tolerant human colorectal adenocarcinoma cell line. I have also detected this phenotype in tumour cells capable of avoiding immune recognition.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Mechanisms of resistance to DNA methylation damage in human cells
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
URI: https://discovery.ucl.ac.uk/id/eprint/10099702
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