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Title: Identification of differentially expressed transcription factors and dissecting the role of DREAM complex associated components in cellular senescence
Author: Kumari, Ruchi
ISNI:       0000 0004 7661 1465
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2019
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Cellular senescence is a stable cell cycle arrest that normal cells undergo in response to a variety of intrinsic and extrinsic stimuli. Being implicated in ageing and age-related diseases including cancer it is of great importance to elucidate the signalling pathways involved in regulating the senescent state. The p53/p21WAF1 and p16INK4A/pRB tumour suppressor pathways have clearly been implicated in senescence, but the critical downstream targets of these pathways are unclear. Transcription factors (TFs) regulate gene expression at different stages of embryonic development and are key to the establishment and maintenance of specific cell fates. My primary goal is to identify TFs that act downstream of these pathways. To identify the TFs that act downstream of the p53/p21WAF1 and p16INK4A/pRB pathways, the previously identified list of differentially expressed transcripts were overlaid with known sequence-specific DNA binding factors. The list was then refined by examining what happens to their expression, when senescence was bypassed and if the change in expression upon senescence is inversely correlated with expression in cancer. This identified 10 upregulated and 74 downregulated TFs. Their ability to directly bypass senescence was examined in the conditionally immortalized fibroblasts by lentivirus mediated RNA silencing or ectopic expression. The MuvB complex (LIN9, LIN37, LIN52, LIN54 and RBBP4) associates with the RB-like proteins, DP1 and E2F4 to form a repressive complex, DREAM, that induces quiescence. When cells re-enter the cell cycle, MuvB dissociates from DREAM and sequentially recruits MYBL2 (B-MYB) and FOXM1 to promote cell cycle progression. Reconstitution of B-MYB-MuvB-FOXM1 i.e. MMB-FOXM1 complex demonstrated that it can bypass senescence under very stringent conditions and that LIN52, FOXM1 and B-MYB were the crucial components. Moreover, this required non-phosphorylated LIN52, suggesting a role for phosphorylated LIN52 and the DREAM complex in inducing senescence, which is not very widely studied. Further reconstitution experiments using a cocktail of TFs targeting the up- and down-regulated factors has revealed the presence of synergy indicating that there are other key TFs which remain to be identified. This study has enabled us to identify TFs that play a causal role in senescence. This opens the door to identifying their downstream targets and lays the foundation for a better understanding of the pathways underlying cellular senescence and its therapeutic intervention in cancer and age-related diseases.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available