Award details

Understanding the molecular dynamics of NHEJ-mediated synapsis

Reference	BB/F013795/1
Principal Investigator / Supervisor	Professor Aidan Doherty
Co-Investigators / Co-Supervisors
Institution	University of Sussex
Department	Genome and Damage Stability Centre
Funding type	Research
Value (£)	818,247
Status	Completed
Type	Research Grant
Start date	01/02/2008
End date	31/01/2012
Duration	48 months

Abstract

In mammalian cells, repair of DNA double strand breaks by non-homologous end-joining (NHEJ) is critical for genome stability. Although the end-bridging and ligation steps of NHEJ have been reconstituted in vitro, little is known about the end processing reactions that occur prior to ligation. Recently, we have identified functionally homologous end-bridging and ligation activities in prokarya. Consistent with its homology to polymerases and nucleases, we have shown that DNA ligase D from Mycobacterium tuberculosis (Mt-Lig) possesses a unique variety of nucleotidyltransferase activities, including gap-filling polymerase, terminal transferase and primase, and is also a 3' to 5' exonuclease. These activities allow the Mt-Ku and Mt-Lig proteins to join incompatible DSB ends in vitro, as well as to reconstitute NHEJ in vivo in yeast. These results demonstrate that prokaryotic Ku and ligase form a bona fide NHEJ system that encodes all the recognition, processing and ligation activities required for DSB repair. The main aim of this proposal is to understand, at the molecular level, how the bacterial NHEJ complexes bring the ends of a break together to form the synaptic complex, which is an essential process prior to the processing and ligation steps of repair. We will employ biochemical, biophysical and crystallographic approaches to 'visualise' the steps that lead to end-synapsis. This research will also provide a conceptual framework for delineating how breaks are brought together in eukaryotic cells.

Summary

Our cells contain DNA, the so called 'genetic blueprint of life' which encodes the information for our genes. DNA has a simple repeating structure composed of two complementary strands of DNA which form a double-helix structure. The integrity of DNA is constantly being challenged by various DNA-damaging agents. These agents include high energy UV and X-ray radiation from the sun, chemicals both man-made and environmental and even the oxygen we breathe can attack and damage DNA. In recent years it has been realised that our own cells produce a large number of proteins responsible for repairing this DNA damage which, if left unchecked, would lead to the development of conditions such as cancer. These protein 'machines' can cut out and replace aberrant DNA mutations/structures and splice together broken DNA strands. One such protein is called DNA ligase, which interacts with a partner protein, Ku. These proteins are able to detect physical breaks in the DNA helices, bring the ends back together and reseal these breaks thus restore the double-strand integrity of the DNA. The focus of this work is to understand how these proteins can bridge the break and make the DNA ends come back together prior to the sealing step.

Committee	Closed Committee - Biomolecular Sciences (BMS)
Research Topics	Microbiology, Structural Biology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

I accept the terms and conditions of use (opens in new window)

export PDF file

back to list new search