DNA recombination pathways and antigenic variation in Trypanosoma brucei

African trypanosomes are single cell parasites of mammals that have major medical, veterinary and economic importance in sub-Saharan Africa. They have been studied for over 100 years, and have revealed many novel aspects of biology that have shed light on fundamental processes in all organisms, including humans. Trypanosomes escape being killed by the mammal?s immune system using a strategy termed antigenic variation. A major component of antigenic variation is rearrangements of specific genes within the parasite?s genome through a reaction termed recombination. Recombination is used in all organisms to repair damage to their genome, and is therefore needed to maintain the integrity of this information source, which dictates all aspects of an organism?s growth, development and reproduction. We have shown that trypanosomes have harnessed this repair pathway for antigenic variation. Nevertheless, our understanding of how recombination acts in antigenic variation is unclear, as is the way that it contributes to other features of the genome and biology of trypanosomes. This project will examine the factors that contribute to trypanosome recombination and antigenic variation, including how they interact, and will characterise a novel form of recombination not previously described in this parasite. Addressing these questions will have fundamental implications for our understanding of recombination in all organisms, but may also reveal specific adaptations of this process in trypanosomes that may be exploited in the future for clinical applications.

DNA recombination is central to the biology of Trypanosoma brucei, including the remarkable chromosome plasticity observed in this parasite and the use of telomeres and subtelomeres to harbour critical genetic functions. In particular, recombination is central to an immune evasion strategy, termed antigenic variation, which consists of periodic changes in the composition of the T. brucei Variant Surface Glycoprotein (VSG) coat. It is important to understand how trypanosomes have exploited recombination pathways to generate such strain diversity and antigenic variability. Previous work has suggested that antigenic variation utilises a conserved pathway for DNA repair, called homologous recombination, catalysed mainly by the enzyme RAD51. However, other, as yet uncharacterised, recombination pathways can also contribute to VSG switching. In addition, some factors (MRE11, MSH2 and MLH1) with demonstrated roles in the catalysis or control of recombination do not contribute to VSG switching, suggesting that the reaction may have specific features we do not yet understand. This proposal seeks to answer two questions that will address these issues. First, the functions of RAD51-related genes recently identified in the T. brucei genome will be elucidated in terms of their roles in general recombination and antigenic variation. This will contribute to our broad understanding of recombination functions in eukaryotes, will determine whether or not these proteins have adopted specific functions in T. brucei, and will test further the catalytic and regulatory functions that influence T. brucei antigenic variation. Second, we have recently characterised, in nuclear extracts, a pronounced, RAD51-independent recombination activity that acts upon short stretches of imperfect sequence homology. The factors that catalyse this reaction, the mechanisms involved and the cellular functions it provides will be elucidated by this proposal.