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Title: Understanding the role of BRCA1 and the mRNA splicing machinery in the maintenance of genomic stability
Author: Vohhodina, Jekaterina
Awarding Body: Queen's University Belfast
Current Institution: Queen's University Belfast
Date of Award: 2016
Availability of Full Text:
Full text unavailable from EThOS. Thesis embargoed until 01 Dec 2021
Abstract:
In order to withstand millions of DNA lesions per day, a sophisticated safeguard and repair system, known as the DNA Damage Response (DDR) system, has evolved to maintain genomic stability within our cells. BRCA1 is one of the key members of the DDR pathway responsible for regulation of transcription, cell cycle arrest and repair of DNA double strand breaks. During the last few years, BRCA1-mediated coordination of RNA processing has been defined as a new layer of complexity in the DDR mechanism. Specifically, a novel BRCA1 interacting RNA splicing complex has recently been identified by our group with the BCLAF1 splicing factor playing a key role in this complex. Other members of this complex are primarily mRNA processing elements, however, their role in the DDR is poorly understood. Here, we demonstrate the importance of a novel, BRCA1 associated DNA damage response (DDR) protein THRAP3 (Thyroid Hormone Receptor Associated Protein 3/TRAP150), which is involved in the same mRNA splicing complex. Moreover, we show that BCLAF1 is able to partially compensate for the absence of THRAP3 and vice-versa, with double knockdown of both splicing factors leading to severe cellular sensitivity to DNA damage and DNA repair deficiency. We also demonstrate that this sensitivity to DNA damaging agents can be at least partially explained by the involvement of THRAP3 and BCLAF1 in the processing of ATM kinase encoding transcripts as well as of a number of other DDR related transcripts. Specifically, we found that BCLAF1 promotes the splicing of these transcripts, whilst THRAP3 promotes their export to the cytoplasm, both tightly linked processes required for transcript stability and protein expression. Taken together, these results suggest that THRAP3 and BCLAF1 mutant tumours may be promising targets for DNA damaging chemotherapy. As discussed, the main role of BRCA1 in the maintenance of genomic stability is protection of cells from DNA double strand breaks. However, recent studies have also demonstrated a role for BRCA1 in the repair of R-loops and Topoisomerase ll-DNA/protein adducts. Here we show that depletion of BRCA1/2 results in cellular sensitivity to 5-azadC and Cu(OP)2, drugs which cause DNA-protein crosslinks. Thus, we suggest that BRCA2 and BRCA1 are involved in the elimination of DNA-protein crosslinks via the FA/ICL and HR mediated DNA repair pathways as the absence of BRCA1/2 results in inhibited DNA replication and reduced DNA end resection. 5-azadC is licensed for the treatment of hematological diseases, however it has not been used as a therapeutic agent for BRCA1/2-mutant tumours. Additionally, we found that BRCA1 and BRCA2 deficient cells are exquisitely sensitive to combination therapy with 5-azadC and PARPi, suggesting synergistic synthetic lethality with this combination. Given that both of these drugs have relatively limited toxicity profiles, this suggests that this combination may prove effective in the treatment of BRCA1/2 deficient tumours, such as those that arise in BRCA1/2 mutation carriers. In conclusion, our results emphasize the significant role of BRCA1 in various aspects of DNA damage repair, through the regulation of both mRNA splicing and export as well as DNA protein crosslink repair. Together these two novel functions of BRCA1 are likely to contribute to BRCATs ability to maintain genomic stability and suppress tumour development.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.713460  DOI: Not available
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