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Title: Functional characterization of WRNIP1
Author: Rossi, Fabio
ISNI:       0000 0004 2742 3273
Awarding Body: Open University
Current Institution: Open University
Date of Award: 2012
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In order to guarantee cell survival, and the transmission of the correct genetic information to their offspring, organisms had to develop molecular pathways to deal with damaged genetic material and mechanisms to fix the damage. During their life, organisms are exposed to enormous amounts of DNA damage due both to mutagenic agents and normal environmental conditions. For instance, hydrolysis causes the release of up to 10000 purine bases from the DNA of a single cell per day. Therefore, DNA Damage Response (DDR) pathways comprise a variety of mechanisms including control and arrest of cell growth, tolerance of datpaged DNA, and DNA repair pathways. In this thesis, I focused on Wemer helicase interacting protein 1 (WRNIPl), and its role in the regulation of DDR. WRNIPl contains a single UBZ domain, which is also found in a group of proteins that are involved in DNA damage tolerance. Moreover WRNIP 1 is post-translationally modified with polyubiquitin chains and polyubiquitinylation is selectively induced after exposure to UV irradiation, suggesting that WRNIPl may have a role in the DNA damage response. Despite this evidence, the specific function of WRNIP 1 in mammalian cells remains unclear. To address this point, I have used different cell based viability assays and I have verified that the level of expression of WRNIP 1 negatively correlates with cell survival upon DNA damage. Significantly, down-regulation of WRNIPl results in a faster release from cell cycle check points, which is one of the main cellular responses to DNA damage. I have also found that these phenotypes are correlated with a WRNIP1-dependent negative regulation of DNA repair. Importantly, I have demonstrated that the effect of WRNIPl on these processes is dependent on the activity of the AAA + ATPase domain. Furthermore, my results on WRNIPl ubiquitinylation revealed that lysine 274 is specifically ubiquitinylated upon UV irradiation. Importantly, lysine 274 is a key residue for the Activity of the AAA+ ATPase, indicating that the ATPase activity of WRNIPl is impaired as part of the cellular response to UV damage. Taken together, these results suggest that WRNIPl functions as negative regulator of DNA repair, and that this function is specifically inhibited through ubiquitinylation when cells face UV-induced DNA damage.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available