Award details

The role of NHEJ repair in bacteria and bacteriophage

Reference	BB/D522746/1
Principal Investigator / Supervisor	Professor Aidan Doherty
Co-Investigators / Co-Supervisors	Dr Robert Pitcher
Institution	University of Sussex
Department	Genome and Damage Stability Centre
Funding type	Research
Value (£)	529,853
Status	Completed
Type	Research Grant
Start date	01/11/2005
End date	31/10/2009
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 nucleotidyl transferase activities, including gap filling polymerase, terminal transferase and primase, and is also a 3¿ to 5¿ exonuclease. These activities allowed 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 the prokaryotic Ku and ligase from 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 identify the cellular role of NHEJ repair. We will characterise the processing and repair of double-strand breaks by bacterial non-homologous end joining (NHEJ) proteins both in vitro and in vivo. We will also investigate how these repair activities are co-ordinated and regulated at both the molecular and cellular level. Recently, Ku proteins have been discovered in bacteriophage. We will also characterise the activity and cellular function of this NHEJ factor.

Summary

All 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. The role of ligases in the cell is to repair breaks in the backbone of the DNA. These enzymes are able to detect physical breaks in the DNA helices and to reseal these breaks and restore the double-strand integrity of the DNA. The focus of this work is to understand how ligase proteins perform their repair function in the cell.

Committee	Closed Committee - Biochemistry & Cell Biology (BCB)
Research Topics	Microbiology
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

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