Project No. 2332

PRIORITY PROJECT

Primary Supervisor

Prof Aidan Doherty – University of Sussex

Co-Supervisor(s)

Prof Neil Kad- University of Kent

Summary

DNA is constantly damaged by external factors (e.g. UV radiation) and internal factors (e.g. free radicals).

Since genome maintenance is crucial for survival, organisms expend large amounts of energy to interrogate, maintain and repair their genomes. In humans, defects in this process can lead to the formation of cancer or premature ageing.

When the cell’s replication machinery is copying its genome, it encounters DNA damage (lesions) and structures, which stall the fork leading to replication stress, a major source of genome instability. To restart the fork again, DNA primers may need to be synthesized. A key protein that undertakes this task in human cells is PrimPol, which operates as both a DNA primase and polymerase. PrimPol acts like an ‘ambulance’, running alongside the replication fork to rapidly restart the copying process once it has been stalled. However, its exact mode of action in DNA replication remains to be elucidated and has not been visualized before. The PhD student will investigate this key process by directly imaging PrimPol in action on DNA using cutting-edge imaging technologies available in Kent. Prof Kad (Kent) will help you to create special DNA substrates that mimic damage and fluorescently tag PrimPol, and other proteins partners, to determine how they interact to restart replication. In parallel, you will be supported by Prof Doherty (Sussex) who will enable you to perform advanced biochemical and biophysical studies to determine how PrimPol is able to generate / extend primers to restart replication.

By directly visualizing how these proteins operate on DNA, coupled with biochemical and structural studies, will enable the student to shed significant new light on one of the most important replication restart mechanisms. These studies will potentially have important implications for our understanding of replication stress pathways involved in cancer progression and drug resistance mechanisms too.