Posttranslational modifications are of great importance in the mediation of transcriptional effects, necessary for signalling in cancer. A characteristic example of such modifications is acetylation of the p53 tumour suppressor, a transcription factor involved in several crucial cellular functions including cell-cycle arrest and apoptosis. p53 is stabilised under hypoxic and DNA damaging-conditions. However, only in the latter scenario is p53 fully capable of inducing the expression of its proapoptotic targets through acetylation. The hypoxia inducible factor 1 (HIF-1) transcription factor is stabilised at low oxygen levels to mediate a cellular adaptive response under these conditions, promoting cell survival. As these two opposing transcription factors share a common transcriptional regulator, p300/CBP, this study focused on deciphering the p300/CBP complex components under differential stress to determine its composition required for cellular responses elicited in response to DNA damage or hypoxia, in an effort to investigate a possible link between differential posttranslational modifications and the resulting cell fate. Hence, the aim of this study was to investigate the roles of p300/CBP components in dictating transcriptional regulation of both HIF-1 and p53 in hypoxic conditions. To carry out this study, the proapoptotic BID gene was the system used, as its promoter contains a p53 response element and a HIF-1 response element (HRE). The p300/CBP associated factors PCAF and Strap were appointed as potent candidates for posttranslational modifications under differential conditions, as they are stress-responsive cofactors. Under DNA damage, PCAF acetylates p53 at K320 and Strap augments p300 binding to p53, both of which amplify the p53 response. Evidence from this study demonstrates that under hypoxia-mimicking conditions PCAF-mediated p53 acetylation at K320 is reduced to a greater extent compared to p300/CBP acetylation at K382. The limited amounts of acetylated p53 at K320 are preferentially recruited to the promoter of the cell cycle arrest p21WAF-1/CIP-1 gene that appears to be unaffected by hypoxia, but fail to be recruited to the BID promoter, rendering p53 incapable of upregulating proapoptotic BID in hypoxic conditions. In addition, under the same conditions, PCAF was found to acetylate, and direct HIF-1 to a particular subset of its targets, leading to alterations in the net physiological effect. Moreover, the intrinsic acetyl transferase activity of PCAF was shown to increase the stability of HIF-1. An additional role was attributed to PCAF in relation to apoptosis, albeit from another angle. BID protein translocation to the cytoplasm in hypoxic conditions was facilitated by ectopically expressed PCAF.Strap was found to be preferentially recruited to the HRE of the BID promoter in hypoxic conditions, and to exert a transrepression effect that appeared to be p53-dependent. Strap also interacted with specific PCAF isoforms depending on the type of cellular stress. Contrary to PCAF, ectopically expressed Strap did not have any effect on BID subcellular distribution. This study has provided additional insight in the mechanisms by which cofactors are involved in cell fate, either by affecting activity and stability of HIF-1 and p53, or having a direct effect on Bcl-2 member subcellular distribution.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:492897 |
| Date | January 2008 |
| Creators | Xenaki, Georgia |
| Publisher | University of Manchester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://www.manchester.ac.uk/escholar/uk-ac-man-scw:152382 |
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