title: DNA polymerase beta from Trypanosoma cruzi is involved in kinetoplast DNA replication and repair of oxidative lesions.
abstract: Specific DNA repair pathways from Trypanosoma cruzi are believed to protect genomic DNA and kinetoplast DNA (kDNA) from mutations. Particular pathways are supposed to operate in order to repair nucleotides oxidized by reactive oxygen species (ROS) during parasite infection, being 7,8-dihydro-8-oxoguanine (8oxoG) a frequent and highly mutagenic base alteration. If unrepaired, 8oxoG can lead to cytotoxic base transversions during DNA replication. In mammals, DNA polymerase beta (Polβ) is mainly involved in base excision repair (BER) of oxidative damage. However its biological role in T. cruzi is still unknown. We show, by immunofluorescence localization, that T. cruzi DNA polymerase beta (Tcpolβ) is restricted to the antipodal sites of kDNA in replicative epimastigote and amastigote developmental stages, being strictly localized to kDNA antipodal sites between G1/S and early G2 phase in replicative epimastigotes. Nevertheless, this polymerase was detected inside the mitochondrial matrix of trypomastigote forms, which are not able to replicate in culture. Parasites over expressing Tcpolβ showed reduced levels of 8oxoG in kDNA and an increased survival after treatment with hydrogen peroxide when compared to control cells. However, this resistance was lost after treating Tcpolβ overexpressors with methoxiamine, a potent BER inhibitor. Curiously, a presumed DNA repair focus containing Tcpolβ was identified in the vicinity of kDNA of cultured wild type epimastigotes after treatment with hydrogen peroxide. Taken together our data suggest participation of Tcpolβ during kDNA replication and repair of oxidative DNA damage induced by genotoxic stress in this organelle.
authors.affiliation: Department of Biochemistry and Immunology, Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, Brazil.
authors.forename: Bruno Luiz Fonseca
authors.initials: BL
authors.lastname: Schamber-Reis
authors.forename: Sheila
authors.initials: S
authors.lastname: Nardelli
authors.forename: Carlos Gustavo
authors.initials: CG
authors.lastname: Régis-Silva
authors.forename: Priscila Carneiro
authors.initials: PC
authors.lastname: Campos
authors.forename: Paula Gonçalves
authors.initials: PG
authors.lastname: Cerqueira
authors.forename: Sabrina Almeida
authors.initials: SA
authors.lastname: Lima
authors.forename: Glória Regina
authors.initials: GR
authors.lastname: Franco
authors.forename: Andrea Mara
authors.initials: AM
authors.lastname: Macedo
authors.forename: Sergio Danilo Junho
authors.initials: SD
authors.lastname: Pena
authors.forename: Christophe
authors.initials: C
authors.lastname: Cazaux
authors.forename: Jean-Sébastien
authors.initials: JS
authors.lastname: Hoffmann
authors.forename: Maria Cristina Machado
authors.initials: MC
authors.lastname: Motta
authors.forename: Sergio
authors.initials: S
authors.lastname: Schenkman
authors.forename: Santuza Maria Ribeiro
authors.initials: SM
authors.lastname: Teixeira
authors.forename: Carlos Renato
authors.initials: CR
authors.lastname: Machado
mesh_terms: D019951:DNA Polymerase beta / Q000378:metabolism*; D004260:DNA Repair; D004261:DNA Replication; D018105:DNA, Kinetoplast / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000737:chemistry / Q000201:enzymology; D018384:Oxidative Stress; D014349:Trypanosoma cruzi / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics
chemical_list: D018105:DNA, Kinetoplast; D019951:DNA Polymerase beta