Cellular DNA is inherently unstable and is continually undergoing damage and degradation. Such damage can be caused by exposure to environmental carcinogens and also as a consequence of normal cellular metabolism, for example via the production of reactive oxygen species (ROS). My research is aimed at understanding the cellular response to DNA damage, with particular emphasis on the base excision repair (BER) and single strand break repair (SSBR) pathways.
Understanding how DNA repair is regulated at the level of single repair events and also within the cell as a whole can aid our understanding of inter-individual variation in cancer susceptibility and hopefully provide clues as to how repair in tumour cells may be attenuated to render cancer treatments more effective. With this aim in mind, there are a number of projects currently in progress in my lab.
Mechanistic studies of DNA repair
DNA repair pathways are multi-step processes involving a number of different enzymes and a network of protein-protein interactions. Through my North West Cancer Research Fund fellowship I have been able to establish a programme of research aimed at understanding how the BER and SSBR pathways are co-ordinated. Through the detailed characterisation of repair of model DNA substrates by purified recombinant enzymes and cell extracts, facilitated by techniques such as site-directed mutagenesis, siRNA and protein-DNA interaction assays, we aim to gain a new mechanistic insight into how damaged DNA is repaired in vivo.
We have recently applied these methods to the study of the bifunctional SSBR enzyme, polynucleotide kinase (PNK). We were able to show that its two activities, kinase and phosphatase, were inter-dependent - with the phosphatase activity taking precedence over the kinase activity. We are currently engaged in follow-up work to understand the consequences of this order of precedence on cellular DNA repair.
In recent years it has become increasingly evident that the enzyme poly(ADP-ribose)polymerase (PARP-1) exerts a significant effect on DNA repair efficiency through both its high affinity for repair intermediates and its physical and functional interaction with many other key DNA repair enzymes. A major focus of my research is therefore aimed at understanding the mechanism of PARP-1 involvement in DNA repair.
The cellular response to UVA exposure
In collaboration with Trevor McMillan's group we are currently have two projects underway investigating the effects of UVA exposure on human cells. Originally thought not to be a significant risk factor for skin cancer, UVA is now believed to contribute as much as 10-20% of the cancer-causing dose of sunlight. UVA induces DNA damage indirectly via photosensitiser-mediated production of ROS.
Of the two projects in progress, the first, funded by the Colt Foundation, is focused on the role of cellular detoxification and DNA repair pathways in protecting against the harmful effects of UVA exposure. The second, funded by the North West Cancer Research Fund, aims to understand the damage response to UVA treatment, especially when administered at different dose intensities.