Metabolismo de DNA em Trypanosoma cruzi: vias de reparo mitocondrial e formação de híbridos

Abstract

Trypanosoma cruzi is a member of the Kinetoplastida and as so, present a single and unique mitochondria, called kinetoplast, which possesses several distinct features in comparison with other eukaryotes mitochondria. Although several proteins of the DNA repair systems have already being described as present on the parasite mitochondria, there’s not much information about the DNA repair pathways involved on the kinetoplast DNA (kDNA) maintenance. In this work we investigate possible DNA repair pathways that could be involved on T. cruzi kDNA metabolism. To first investigate the repair proteins related to the kDNA metabolism we analyzed the effect of overexpressing the protein TcMYH, which is involved in base excision DNA repair in T. cruzi. As shown by qPCR, the overexpressor strain of TcMYH has more damages on the kDNA at the first moment, when compared to the wild type strain. Also, when we used a reagent designed to specifically cause damage on the mitochondrial DNA, we could observe that this mutant is more sensitive than the WT control strain, and that the overexpression of the protein generates more AP sites on the parasite organelle. These results suggest that the parasite is able to deal with oxidative damage that attacks the kDNA. One of the unique features of the kinetoplast is the presence of specific proteins called kinetoplast associated proteins (KAP). Although some works described the location of those proteins on the antipodal sites, regions of DNA metabolism on the kinetoplast, their function are not yet clear. In this work we study the function of the proteinTcKAP7, which has sequence homology with the Mitochondrial transcription factor A (TFAM). Our results demonstrate that the absence of TcKAP7 is related to a long-term sensitivity to UV radiation and cisplatin; and a resistance to high levels oxidative stress on the parasite. The same phenotype has been observed for Angomonas deanei mutants, another trypanossomatid and an insect parasite. In this work we also investigate the genetic exchange events in different T. cruzi strains. Our results demonstrate that T. cruzi is able to perform genetic exchange among individuals in a same population. Also, the variety of T. cruzi strains are able to perform genetic exchange among them. This exchange is also linked to the levels of TcRad51 on these strains. Together, these results demonstrate that this parasite is able to perform genetic exchange and that the homologous recombination could be linked to this process.