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Natalie M. Hill, Lyndsay Davies, Dave Spiller, Mike White, Ian Grierson, Luminita Paraoan; Positive Feedback Between p53 and PERP During Apoptosis Induction in Uveal Melanoma Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):513.
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We have previously shown that the expression of PERP (p53 apoptosis effector related to PMP-22), an apoptosis-specific effector of p53, triggers the death of uveal melanoma cells by apoptosis. The purpose of this study was to investigate the effect of increased PERP expression on the expression and subcellular localisation of its own upstream transcriptional regulator, p53.
Fluorescent fusion protein constructs of PERP, p53 and MDM2, were used to transfect the human uveal melanoma cell line MEL202. Western blotting was used to detect the expression of green fluorescent protein-PERP fusion (GFP-PERP), p53 and MDM2. The subcellular localization of PERP, p53 and MDM2 fluorescent fusion proteins in transfected cells was analysed by confocal microscopy.
Significantly increased levels of p53 protein were observed in MEL202 cells expressing GFP-PERP compared with control cells, which was also accompanied by an increase in the levels of p53 negative regulator MDM2. Expression of GFP-PERP resulted in nuclear localization of MDM2-yellow fluorescent protein (MDM2-YFP) in 96.8% of co-transfected cells. In contrast, MDM2-YFP expression alone or in combination with GFP expression had additional diffuse cytoplasmic localization. In the absence of GFP-PERP expression, p53 fused to red fluorescent protein (p53-RFP) was localized primarily in the cytoplasm with low expression in the nucleus. However, following GFP-PERP expression, p53-RFP was localized predominantly in the nucleus in a significantly higher proportion of co-transfected cells.
Our results show that PERP protein levels influence the protein levels of its own transcriptional regulator, p53. Additionally, PERP expression causes nuclear localization of p53 that is critical for its transcriptional function. Together, these results propose a novel role for PERP in enhancing p53 levels and reveal a potential new target for exploitation in the development of new therapeutic agents aimed at increasing the endogenous p53 protein pool in neoplastic cells.
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