title: Sequence homology and microhomology dominate chromosomal double-strand break repair in African trypanosomes.
abstract: Genetic diversity in fungi and mammals is generated through mitotic double-strand break-repair (DSBR), typically involving homologous recombination (HR) or non-homologous end joining (NHEJ). Microhomology-mediated joining appears to serve a subsidiary function. The African trypanosome, a divergent protozoan parasite, relies upon rearrangement of subtelomeric variant surface glycoprotein (VSG) genes to achieve antigenic variation. Evidence suggests an absence of NHEJ but chromosomal repair remains largely unexplored. We used a system based on I-SceI meganuclease and monitored temporally constrained DSBR at a specific chromosomal site in bloodstream form Trypanosoma brucei. In response to the lesion, adjacent single-stranded DNA was generated; the homologous strand-exchange factor, Rad51, accumulated into foci; a G(2)M checkpoint was activated and >50% of cells displayed successful repair. Quantitative analysis of DSBR pathways employed indicated that inter-chromosomal HR dominated. HR displayed a strong preference for the allelic template but also the capacity to interact with homologous sequence on heterologous chromosomes. Intra-chromosomal joining was predominantly, and possibly exclusively, microhomology mediated, a situation unique among organisms examined to date. These DSBR pathways available to T. brucei likely underlie patterns of antigenic variation and the evolution of the vast VSG gene family.
authors.affiliation: London School of Hygiene & Tropical Medicine, Keppel Street, London, WC1E 7HT, UK.
authors.forename: Lucy
authors.initials: L
authors.lastname: Glover
authors.forename: Richard
authors.initials: R
authors.lastname: McCulloch
authors.forename: David
authors.initials: D
authors.lastname: Horn
mesh_terms: D000818:Animals; D002453:Cell Cycle; D019457:Chromosome Breakage; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D016054:DNA, Protozoan / Q000737:chemistry; D007700:Kinetics; D015800:Protozoan Proteins / Q000378:metabolism; D051135:Rad51 Recombinase / Q000378:metabolism; D011995:Recombination, Genetic; D012689:Sequence Homology, Nucleic Acid; D014346:Trypanosoma brucei brucei / Q000235:genetics* / Q000378:metabolism
chemical_list: D016054:DNA, Protozoan; D015800:Protozoan Proteins; D051135:Rad51 Recombinase