Desvendando o reparo de dna mitocondrial em Trypanosoma cruzi

• Ano de defesa: 2019
• Autor(a) principal: Wesley Roger Rodrigues Ferreira
• Tipo de documento: Dissertação
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Minas Gerais
• Palavras-chave em Português: Bioquímica e Imunologia; Doença de Chagas; Reparo do DNA; Mitocôndrias; Doxorrubicina; Trypanosoma cruzi; Reparo de DNA
• Link de acesso: https://hdl.handle.net/1843/78873

Trypanosoma cruzi, the etiologic agent of Chagas disease – one of the seventeen neglected tropical diseases –, is a member of the Kinetoplastida order and, as such, has a single, elongated mitochondria named kinetoplast. In this study we investigated the DNA repair pathways that are responsible to maintain the integrity of the kinetoplastid genome (kDNA) from T. cruzi. Although we have evidences about the conduction of DNA repair to some extent in the maintenance of the kDNA, this process and proteins involved in this metabolism are not yet described. In this work we used wild-type and mutant epimastigotes of T. cruzi clone CL Brener, namely (i) single knockout strain for TcRAD51 (a gene which encodes a protein involved in homologous recombination); (ii) a single knockout strain for TcCSB (a gene which encodes a protein involved in nucleotide excision repair); and (iii) a strain overexpressing TcCSB. After treatment with MMS, an agent capable of generating double strand breaks to the DNA molecule – a damage repaired by homologous recombination –, we verified that the TcRAD51 deficient strain was more sensitive to the treatment. In order to verify whether the difference observed is associated to kDNA repair, we further performed the quantification of DNA damage. After the treatment with MMS, we observed a difference in the kinetics of DNA repair between both strains. In addition, we verified that TcRAD51 single knockout is more sensitive to agents capable of generating double strand breaks by distinct mechanisms. Mitochondria-oriented doxorubicin assays – a drug capable of causing transcription and replication problems – demonstrated that, in T. cruzi kinetoplast, there are pathways related to these damages. Single knockout and overexpressing TcCSB cells, following exposure to this compound, demonstrated an involvement of TcCsb with kDNA repair metabolism. These results suggest that TcRad51 and TcCSB are involved in kDNA repair in T. cruzi, although the exact mechanisms by which these proteins in T. cruzi mitochondria have yet to be determined. The influence of TcRad51 in the two repair moments in the damage generated by MMS also suggests that the mitochondrial repair pathways may be distinct from that one conducted in the nucleus.