Understanding how replication-coupled DNA repair protects against ageing and cancer

Supervisor:  Prof Peter McHugh

Our cellular DNA is constantly being assaulted by reactive metabolites and their by-products. Many of the resulting DNA lesions can be repaired throughout the cell cycle. However certain forms of damage such as DNA interstrand cross-links (ICLs) and DNA protein crosslinks (DPCs) are selectively recognised and repaired during DNA replication. ICLs and DPCs block DNA replication and if unrepaired, or misrepaired, produce devastating health conditions. This is exemplified by the inherited syndrome Fanconi anaemia, where patients suffer from bone marrow failure, developmental defects and ultimately a massively increased risk of leukaemia and solid tumours, often in childhood.

A key effector of the Fanconi anaemia DNA repair pathway is XPF/FANCQ protein. XPF is the active subunit of a dimeric endonuclease (XPF-ERCC1), capable of processing replication fork structures. Despite the clear link between XPF, replication-coupled DNA repair and human disease, very little is known about the mechanism of the recruitment of XPF to damage-arrested forks, and how XPF interacts with and processes these forks. The proposed project takes advantage of a newly developed a platform to study the dynamics of proteins such as XPF at replication forks using CRISPR-Cas9-based endogenous-gene tagging strategies coupled to super-resolution microscopy and ChIP-seq genomics approaches. This will allow us to define the spatio-temporal dynamics, and genetic requirements for the recruitment of XPF to damaged forks in living cells, and to identify new key players in this process. In addition, we possess unique capabilities for synthesising structures that mimic damaged DNA replication forks that will be used to reconstitute the key cellular reactions in vitro. Together with collaborators, we will also use structural and biophysical approaches to define replication fork repair at previously unachievable levels of detail.

Training opportunities

In addition to standard molecular/cellular biology approaches, the studentship will provide training in a wide range of cutting-edge scientific methods including: genome-editing (CRISPR-Cas9), super-resolution microscopy and imaging methods, protein purification and analysis, including mass-spectrometry, genomics, as well as biochemical and biophysical studies of DNA-protein interactions; there is also an opportunity to learn structural biology methodologies through our collaborations.

References 

  1. Abdullah UB, McGouran JF, Ptchelkine, D, El-Sagheer AH, Brown T and McHugh PJRPA activates XPF-ERCC1 to initiate the processing of DNA interstrand crosslinks.EMBO J 2017; 36(14):2047-2060
  2. Wang AT et al. Human SNM1A and XPF-ERCC1 collaborate to initiate DNA interstrand cross-link repair. Genes and Development 2011; 25 (17):1859-70

For further information please contact: Prof Peter J. McHugh