We also suggest a possible involvement of RAD50 in subtelomeric regions stability. We demonstrate the occurrence of chromosomal translocations in the MRE11 and RAD50 deficient cells and described a MRE11-independent microhomology-mediated end-joining mechanism at the level of translocation breakpoints. In this manuscript, we present the conditional inactivation of the Leishmania infantum RAD50 gene that was only possible in MRE11 deficient cells and suggest the genetic background is crucial for RAD50 inactivation. Studies in other organisms have provided evidence that the DNA repair proteins MRE11 and RAD50 are involved in chromosomes organization, protection of chromosomes ends and therefore in the maintenance of genomic integrity.
However, maintaining a minimum level of genomic integrity is crucial for cell survival. The parasite Leishmania relies on gene rearrangements to survive stressful conditions. Moreover, we suggest the possible involvement of RAD50 in subtelomeric regions stability. This study demonstrates an MRE11-independent microhomology-mediated end-joining mechanism and a prominent role for MRE11 and RAD50 in the maintenance of genomic integrity. Sequencing data also showed a decreased coverage at subtelomeric locations in many chromosomes in the MRE11 -/- RAD50 -/- parasites. Sequencing of the junction between translocated chromosomes highlighted microhomology sequences at the level of breakpoint regions. Inactivation of the MRE11 and RAD50 genes led to a decreased frequency of homologous recombination and analysis of the null mutants by whole genome sequencing revealed several chromosomal translocations. Remarkably, inactivation of RAD50 was possible in a MRE11 null mutant that we had previously generated, providing good evidence that RAD50 may be dispensable in the absence of MRE11.
Attempts to generate RAD50 null mutants in a wild-type background failed and we provide evidence that this gene is essential. We hypothesized that maintenance of genomic integrity in Leishmania would imply a leading role of the MRE11 and RAD50 proteins considering their role in DNA repair, chromosomal organization and protection of chromosomes ends in other organisms. However, safeguarding a minimum level of genome integrity is important for cell survival. The parasite Leishmania often relies on gene rearrangements to survive stressful environments.
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