The p53 research area represents an ever expanding field in which new effectors and new biological functions contribute to broaden the cellular activities in which p53 is involved in both physiological and pathological conditions. Among these biological functions, p53 is acknowledged to be a key factor in controlling genomic stability, apoptosis, metabolism, response to stress and DNA damage. Thus, the search for new p53-target genes is crucial as these could shed new light into the mechanisms through which p53 controls cell integrity and response to damage, leading to the identification new perspective therapeutic targets. The aim of the present work was to complete the characterization of DRAGO (DRug Activated Gene Overexpressed)IKIAA0247, a highly conserved gene. Data previously obtained in the lab supported the hypothesis of a potential tumour-suppressive role for DRAGO since: the gene was demonstrated to be up-regulated in vitro in response to treatment with chemotherapeutic compounds; p53 and p73 were shown to be involved in gene expression regulation; DRAGO ectopic overexpression caused cell death in host cell lines; stage III ovarian cancers displayed significantly reduced levels ofDRAGO expression compared to stage I samples. However DRAGO knockout mice had a normal phenotype and did not develop spontaneous tumours. The experiments described in this thesis work confirmed the tumour-suppressor role of DRAGO by demonstrating that in p53-1- or p53+1- mice, the deletion of DRAGO alleles significantly accelerated tumour development and shortened lifespan compared to p53-1- or p53 +1- mice bearing wild-type DRAGO alleles. DRAGO was also proved to be responsive to DNA damaging treatments in vivo displaying a tissue-specific expression pattern, similarly to other p53-downstream targets. In vitro experiments showed that DRAGO expression is regulated both at transcriptional level - through p53, p73, and promoter-methylation mechanisms - and at post-transcriptional levels by miRNAs. The generation of GFP fusion protein confirmed that DRAGO expression induced cell death but neither apoptosis or senescence seemed to be involved in DRAGO-mediated cell toxicity. Overall these data contributed to the characterization of a new p53-downstream gene and confirmed its tumour suppressive properties. As regards the biological fi1nction of the gene, preliminary experiments performed on mouse macrophages seemed to suggest a possible role ofDRAGO in the immune system functionality, even though further work is required to define how the gene mediates its tumour protective functions in this immune cell subpopulation
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:607447 |
| Date | January 2014 |
| Creators | Rusconi, Paolo |
| Publisher | Open University |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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