Investigation of the Role of Chromosome 7 in Human Cell Immortalisation and Cancer

Berrington, Mary (1999) Investigation of the Role of Chromosome 7 in Human Cell Immortalisation and Cancer. PhD thesis, University of Glasgow.

Abstract

High frequencies of loss of heterozygosity have been observed at the q31 region of chromosome 7 in a variety of tumour types, suggesting that a multi-tissue tumour suppressor gene maps to this region. Introduction of an intact copy of human chromosome 7 into non-tumourigenic cell lines that have lost 7q31 markers and belong to immortality complementation group D causes a proportion of hybrids to senesce. This indicates that the group D senescence gene maps to chromosome 7. Moreover, this gene may be the target of losses observed in tumours. To further investigate these effects I have used the microcell mediated monochromosome transfer technique to introduce chromosome 7 into the ovarian tumour cell line OVCAR5 and the breast tumour cell line MDA-MB-231. I had previously shown that both these cell lines are hemizygous for chromosome 7q and belong to senescence complementation group D as determined by mortalin staining and cell-cell fusion analysis. Following microcell transfer approximately 30% of hybrids of each cell type underwent a rapid growth arrest. Growth-arrested hybrids stained positive for senescence-associated endogenous ?-galactosidase activity and showed reversion of the mortality marker mortalin to its mortal distribution. In total 79 immortal hybrids, which have presumably deleted or otherwise inactivated the senescence gene on the introduced chromosome, were also generated. PCR-based microsatellite deletion analysis demonstrated that more than half of these had lost markers from the introduced chromosome, but no single region of loss was identified. To investigate whether the 7q senescence and tumour suppressor gene are one and the same, I compared the growth in soft agar and the tumourigenicity in nude mice of three immortal OVCARS/Chromosome 7 hybrids-specifically hybrids for which no microsatellite losses had been found-to that of the parent cell line. A reduced tumourigenicity of these hybrids would suggest that the senescence-inducing and tumour suppressive effects of the chromosome are separable. Although two of the hybrids did indeed exhibit reduced colony formation in soft agar and delayed tumour formation in nude mice, interpretation of these results was clouded by the high level of clonal variation found within the cell line as a whole. To circumvent this problem I investigated the soft agar growth and tumourigenicity of a panel of OVCAR5/Chromosome 7 hybrids that I had derived from the same single OVCAR5 cell clone. Five of the six hybrids generated showed a marked reduction in colony formation in soft agar compared to the parent cell clone. Three of these hybrids were then tested for tumourigenicity in nude mice: two displayed a significant delay in tumour formation. These results suggest that there is a gene on chromosome 7, independent of the senescence gene, that can suppress the tumourigenicity of the OVCAR5 cell line. The challenge now is to identify these genes and to assess their relative contributions to tumour suppression and tumourigenicity.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Genetics
• Date of Award: 1999
• Depositing User: Enlighten Team
• Unique ID: glathesis:1999-74528
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 27 Sep 2019 17:56
• Last Modified: 27 Sep 2019 17:56
• URI: https://theses.gla.ac.uk/id/eprint/74528