INGEBI   02650
INSTITUTO DE INVESTIGACIONES EN INGENIERIA GENETICA Y BIOLOGIA MOLECULAR "DR. HECTOR N TORRES"
Unidad Ejecutora - UE
artículos
Título:
Disrupted ADP-ribose metabolism with nuclear Poly (ADP-ribose) accumulation leads to different cell death pathways in presence of hydrogen peroxide in procyclic Trypanosoma brucei.
Autor/es:
MARIANA SCHLESINGER; SALOMÉ VILCHEZ LARREA; TEEMU HAIKARAINEN; MOHIT NARWAL; HARIKANTH VENKANNAGARI; MIRTHA M. FLAWIÁ; LARI LEHTIÖ; SILVIA H. FERNÁNDEZ VILLAMIL
Revista:
PARASITES AND VECTORS
Editorial:
BIOMED CENTRAL LTD
Referencias:
Lugar: Londres; Año: 2015 vol. 9 p. 1 - 15
ISSN:
1756-3305
Resumen:
Background: Poly(ADP-ribose) (PAR) metabolism participates in several biological processes such as DNA damagesignaling and repair, which is a thoroughly studied function. PAR is synthesized by Poly(ADP-ribose) polymerase(PARP) and hydrolyzed by Poly(ADP-ribose) glycohydrolase (PARG). In contrast to human and other highereukaryotes, Trypanosoma brucei contains only one PARP and PARG. Up to date, the function of these enzymes hasremained elusive in this parasite. The aim of this work is to unravel the role that PAR plays in genotoxic stressresponse.Methods: The optimal conditions for the activity of purified recombinant TbPARP were determined by using afluorometric activity assay followed by screening of PARP inhibitors. Sensitivity to a genotoxic agent, H2O2, wasassessed by counting motile parasites over the total number in a Neubauer chamber, in presence of a potent PARPinhibitor as well as in procyclic transgenic lines which either down-regulate PARP or PARG, or over-express PARP.Triplicates were carried out for each condition tested and data significance was assessed with two-way Anovafollowed by Bonferroni test. Finally, PAR influence was studied in cell death pathways by flow cytometry.Results: Abolition of a functional PARP either by using potent inhibitors present or in PARP-silenced parasites hadno effect on parasite growth in culture; however, PARP-inhibited and PARP down-regulated parasites presented anincreased resistance against H2O2 treatment when compared to their wild type counterparts. PARP over-expressingand PARG-silenced parasites displayed polymer accumulation in the nucleus and, as expected, showed diminishedresistance when exposed to the same genotoxic stimulus. Indeed, they suffered a necrotic death pathway, while anapoptosis-like mechanism was observed in control cultures. Surprisingly, PARP migrated to the nucleus andsynthesized PAR only after a genomic stress in wild type parasites while PARG occurred always in this organelle.Conclusions: PARP over-expressing and PARG-silenced cells presented PAR accumulation in the nucleus, even inabsence of oxidative stress. Procyclic death pathway after genotoxic damage depends on basal nuclear PAR. Thisevidence demonstrates that the polymer may have a toxic action by itself since the consequences of an exacerbatedPARP activity cannot fully explain the increment in sensitivity observed here. Moreover, the unusual localization ofPARP and PARG would reveal a novel regulatory mechanism, making them invaluable model systems.