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UBC Theses and Dissertations

Molecular functions of the tumour suppressor p33ING2 Chin, Mei Yieng


Since p33/A/G2 was cloned in 1998, there have been several studies indicating that p33ING2 is a tumour suppressor candidate as it regulates gene transcription, cell cycle arrest and apoptosis in various human carcinoma cell lines. These functions have been shown to be dependent on the activity of wild-type p53. p33ING1 b, which shares 58.9% homology with p33ING2, has been shown to play a role in cellular stress response to UV irradiation. Due to the structural similarities between p33ING1b and p33ING2, we hypothesized that p33ING2 enhances UVB-induced apoptosis. To test this hypothesis, we investigated the role of p33ING2 in UV-induced apoptosis and the repair of UV-damaged DNA. We report that overexpression of p33ING2 enhances UVB-induced apoptosis in wild-type p53 MMRU melanoma cells, but not in mutant p53 MeWo melanoma cells. We demonstrate that the enhancement of UVB-induced apoptosis by p33ING2 requires the presence of functional p53. Furthermore, we found that overexpression of p33ING2 upregulates the expression of endogenous Bax and downregulates the expression of Bcl-2, resulting in an increased Bax/Bcl-2 ratio. Moreover, we also found, that p33ING2 promotes Bax translocation to the mitochondria, alters the mitochondrial membrane potential, and induces cytochrome c release, thus activating caspases 9 and 3 upon UV irradiation. In addition, we show that under non-stressed conditions, p33ING2 upregulates Fas expression and activates caspase 8. These results indicate that p33ING2 cooperates with p53 to regulate apoptosis via activation of both the mitochondrial/intrinsic and death-receptor/extrinsic apoptotic pathways. To investigate if p33ING2 is also involved in DNA repair in melanoma cells, we transfected the p33ING2 expression vector into melanoma cells. We report that overexpression of p33ING2 enhances the repair of UV-damaged DNA, and that this process requires the activity of functional p53. We also show that p33ING2 is not directly recruited to UV-induced DNA lesions, both cyclobutane pyrimidine dimers (CPDs) and 6-4 photoproducts (6-4PPs), suggesting that p33ING2 is not a component of the repair core complexes. Furthermore, we found that p33ING2 enhances p53 acetylation and upregulates the expression of XPA, which is involved in DNA damage recognition. These observations suggest that p33ING2 cooperates with p53 in mediating nucleotide excision repair, and that XPA may be involved in this repair process. In conclusion, we have elucidated in this thesis, the role of p33ING2 in cellular stress response to UV irradiation, and the importance of this gene in the context of tumour suppression.

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