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Title: Nucleases and histone acetyltransferases in DNA repair and immune diversity
Author: Sheppard, E.
Awarding Body: University of Exeter
Current Institution: University of Exeter
Date of Award: 2019
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DNA repair mechanisms are essential for genome maintenance and adaptive immunity. A careful balance must be achieved whereby highly accurate and efficient canonical repair protects the genome from accumulating mutations that lead to aging and cancer, and yet mutation and error-prone non-canonical repair is required for generating immune diversity. Immune diversity is achieved within a tightly regulated environment in which mutator proteins are directed to the antibody locus to introduce a swathe of DNA damage. This produces high affinity antibodies that recognise an infinite number of invading pathogens. This process of secondary antibody diversification is dependent on both active transcription and DNA repair. Downstream of histone signalling, DNA repair nucleases are recruited to remove the damaged bases. The structure of damaged regions in the DNA can have very different conformations depending on whether the source of the damage is endogenous or exogenous. Specific DNA nucleases recognise particular DNA substrates and generate DNA intermediates that are repaired in conjunction with polymerases and ligases. Despite their multitude and importance to DNA repair, very few nucleases have been characterised, while the activities of some studied nucleases remain controversial. Conventional techniques for studying DNA nucleases have several disadvantages; they are hazardous, laborious, time-consuming, and capture nuclease activity in a discontinuous manner. Recognising a need for a safer, faster alternative, a fluorescence-based method has been developed for the study of DNA nucleases, nickases and polymerases. Key histone modifications that are known to orchestrate canonical DNA repair have since been discovered to regulate non-canonical repair at the antibody locus. The Kat5 histone lysine acetyltransferase functions highly upstream of DNA repair and promotes active transcription, yet a role for Kat5 in secondary antibody diversification has not yet been established. Using chemical inhibitors to prevent the catalytic activities of Kat5, and the genetic method of an inducible degron system for rapid and reversible downregulation of Kat5, a role for Kat5 in secondary antibody diversification is recognised, and the research contributes to our current understanding of the DNA repair signal transduction pathway.
Supervisor: Chahwan, R. ; Harmer, N. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available