Structure:Function Correlation in the Human DNA Repair Factor CtIP

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This proposal will build upon our recently published work (Wilkinson at al., eLife) to provide a step change in our understanding of the stucture:function relationships in the important DNA break repair factor CtIP. This protein plays a critical role in the mechanism and regulation of the human resectosome as a hub protein that integrates signalling information and co-ordinates broken DNA with many other repair factors. Despite intense interest in CtIP and DNA break repair more widely, we have a very poor understanding of its function at the molecular level.

Our major objective is to identify and map the interactions that are made between CtIP and other protein partners within the resectosome, and to better understand how CtIP interacts with broken DNA, including though the identification of its unknown DNA binding site. We hypothesise that these interactions may be perturbed by regulatory post-translational modifications or disease-state mutations, and that these changes can impact on DNA break repair pathways. We will test this directly using a range on biochemical and cell biological assays. Therefore, the development of tools to define CtIP structure:function relationships will provide many novel molecular level insights into double-stranded DNA break repair mechanism, regulation and dysfunction in the disease state.

A variety of different stakeholders will benefit from our research into CtIP and its role in homologous recombination which we briefly summarise under four headings below. Further details, including how we will deliver these objectives through specific activities, are available in the Pathways to Impact document.

Academic Impact Objectives: We will gain fundamental new insights into DNA break repair by the homologous recombination (HR) pathway. This is directly relevant to the fields of DNA break repair, cell cycle regulation, mitosis, meiosis, telomere biology, genetic instability, molecular ageing, recombination, gene editing, macromolecular interactions, integrated structural biology and DNA:protein interactions. Our work will also involve technique development in mass spectrometry and single molecule methods.

UK Skills Base and Training Objectives: Our interdisciplinary proposal represents an outstanding training opportunity for the PDRA Wilkinson and other staff members in the collaborating groups. Novel developments in structural-MS will benefit the UK biosciences community more widely.

Biotechnology and Medicine Objectives: Our work is highly relevant to improving molecular level understanding of cancer and human genetic diseases including the dwarfism disorders Seckel and Jawad syndrome that are caused by mutation in CtIP. This protein is also a potential target as a cancer therapeutic based on the concept of synthetic lethality between degenerate DNA repair pathways. Finally, enhancement and/or modulation of the EJ/HR DSB repair pathway choice is an important element of improving modern gene editing techniques to a level where they might be routinely used therapeutically.

Education and Public Engagement Objectives: We will inform and inspire the public and explain why our work on DNA repair is important to them and should be funded.