Distinct Phenotypes Caused by Mutation of <i>MSH2</i> in Trypanosome Insect and Mammalian Life Cycle Forms Are Associated with Parasite Adaptation to Oxidative Stress

Background DNA repair mechanisms are crucial for maintenance of the genome in all organisms, including parasites where successful infection is dependent both on genomic stability and sequence variation. MSH2 is an early acting, central component of the Mismatch Repair (MMR) pathway, which is responsible for the recognition and correction of base mismatches that occur during DNA replication and recombination. In addition, recent evidence suggests that MSH2 might also play an important, but poorly understood, role in responding to oxidative damage in both African and American trypanosomes. Methodology/Principal Findings To investigate the involvement of MMR in the oxidative stress response, null mutants of MSH2 were generated in Trypanosoma brucei procyclic forms and in Trypanosoma cruzi epimastigote forms. Unexpectedly, the MSH2 null mutants showed increased resistance to H2O2 exposure when compared with wild type cells, a phenotype distinct from the previously observed increased sensitivity of T. brucei bloodstream forms MSH2 mutants. Complementation studies indicated that the increased oxidative resistance of procyclic T. brucei was due to adaptation to MSH2 loss. In both parasites, loss of MSH2 was shown to result in increased tolerance to alkylation by MNNG and increased accumulation of 8-oxo-guanine in the nuclear and mitochondrial genomes, indicating impaired MMR. In T. cruzi, loss of MSH2 also increases the parasite capacity to survive within host macrophages. Conclusions/Significance Taken together, these results indicate MSH2 displays conserved, dual roles in MMR and in the response to oxidative stress. Loss of the latter function results in life cycle dependent differences in phenotypic outcomes in T. brucei MSH2 mutants, most likely because of the greater burden of oxidative stress in the insect stage of the parasite.研究背景 脱氧核糖核酸（DNA）修复机制对所有生物体的基因组维持均至关重要，而寄生虫的成功感染同时依赖基因组稳定性与序列变异。MSH2是错配修复（Mismatch Repair, MMR）通路的早期作用核心组分，该通路负责识别并校正DNA复制与重组过程中出现的碱基错配。此外，近期研究显示，MSH2在非洲锥虫与美洲锥虫的氧化损伤应答中也可能发挥重要但机制尚未明确的作用。 方法学/主要研究结果 为探究MMR在氧化应激应答中的参与作用，研究人员分别在布氏锥虫（Trypanosoma brucei）前循环型虫体与克氏锥虫（Trypanosoma cruzi）上鞭毛体虫体中构建了MSH2基因敲除突变株。令人意外的是，与野生型细胞相比，MSH2敲除突变株对过氧化氢（H₂O₂）暴露的抗性显著提升，这一表型与此前报道的布氏锥虫血流型MSH2突变株敏感性升高的结果存在显著差异。互补实验证实，布氏锥虫前循环型虫体的氧化抗性增强源于其对MSH2缺失状态的适应。在两种锥虫中，MSH2的缺失均会导致其对MNNG（N-甲基-N'-硝基-N-亚硝基胍）烷基化损伤的耐受性增强，同时细胞核与线粒体基因组中8-氧代鸟嘌呤（8-oxo-guanine）的积累量升高，提示MMR功能受损。在克氏锥虫中，MSH2的缺失还可增强寄生虫在宿主巨噬细胞内的存活能力。 结论与意义 综合上述研究结果可知，MSH2在MMR与氧化应激应答中均具有保守的双重功能。MSH2缺失导致的后者功能丧失，会使布氏锥虫MSH2突变株的表型出现生命周期依赖性差异，这大概率是因为该寄生虫的昆虫阶段面临更高的氧化应激负荷。