Chromosomal recombination and repair in African trypanosomes. (360G-Wellcome-089172_Z_09_Z)

The African trypanosome, Trypanosoma brucei, is an important human and animal pathogen. In T. brucei, antigenic variation is essential for survival in the mammalian host and involves homologous recombination (HR) at subtelomeric loci, leading to the genetic reassortment of variant surface glycoprotein (VSG) genes. The molecular controls and mechanisms underlying this process remain only partially characterised. We recently used inducible expression of a meganuclease to spatially and temporally c ontrol the introduction of a single DNA double-strand-break (DSB) per T. brucei genome. Our studies revealed efficient HR-mediated repair of the DSB at a chromosome-internal locus but little repair at a transcriptionally repressed sub-telomeric locus. End-joining repair was also detected but this was exclusively microhomology-mediated. We now propose to employ the meganuclease system to dissect the pathways and control of DSB repair in T. brucei. We will determine which pathways are enacted by D SBs at different loci and which cell-cycle checkpoints are invoked in these settings. We will monitor the assembly of the repair machinery at the different DSBs and genetically dissect these pathways. The work will reveal the molecular mechanisms that are co-opted for immune evasion in this important pathogen.