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

Autores
Schlesinger, Mariana; Vilchez Larrea, Salomé Catalina; Haikarainen, Teemu; Narwal, Mohit; Venkannagari, Harikanth; Flawia, Mirtha Maria; Lehtiö, Lari; Fernandez Villamil, Silvia Hebe
Año de publicación
2016
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Poly(ADP-ribose) (PAR) metabolism participates in several biological processes such as DNA damage signaling 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 higher eukaryotes, Trypanosoma brucei contains only one PARP and PARG. Up to date, the function of these enzymes has remained elusive in this parasite. The aim of this work is to unravel the role that PAR plays in genotoxic stress response.MethodsThe optimal conditions for the activity of purified recombinant TbPARP were determined by using a fluorometric activity assay followed by screening of PARP inhibitors. Sensitivity to a genotoxic agent, H2O2, was assessed by counting motile parasites over the total number in a Neubauer chamber, in presence of a potent PARP inhibitor 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 Anova followed by Bonferroni test. Finally, PAR influence was studied in cell death pathways by flow cytometry.ResultsAbolition of a functional PARP either by using potent inhibitors present or in PARP-silenced parasites had no effect on parasite growth in culture; however, PARP-inhibited and PARP down-regulated parasites presented an increased resistance against H2O2 treatment when compared to their wild type counterparts. PARP over-expressing and PARG-silenced parasites displayed polymer accumulation in the nucleus and, as expected, showed diminished resistance when exposed to the same genotoxic stimulus. Indeed, they suffered a necrotic death pathway, while an apoptosis-like mechanism was observed in control cultures. Surprisingly, PARP migrated to the nucleus and synthesized PAR only after a genomic stress in wild type parasites while PARG occurred always in this organelle.ConclusionsPARP over-expressing and PARG-silenced cells presented PAR accumulation in the nucleus, even in absence of oxidative stress. Procyclic death pathway after genotoxic damage depends on basal nuclear PAR. This evidence demonstrates that the polymer may have a toxic action by itself since the consequences of an exacerbated PARP activity cannot fully explain the increment in sensitivity observed here. Moreover, the unusual localization of PARP and PARG would reveal a novel regulatory mechanism, making them invaluable model systems.
Fil: Schlesinger, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
Fil: Vilchez Larrea, Salomé Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
Fil: Haikarainen, Teemu. University of Oulu; Finlandia
Fil: Narwal, Mohit. University of Oulu; Finlandia
Fil: Venkannagari, Harikanth. University of Oulu; Finlandia
Fil: Flawia, Mirtha Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
Fil: Lehtiö, Lari. University of Oulu; Finlandia
Fil: Fernandez Villamil, Silvia Hebe. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
Materia
Trypanosoma brucei
PAR
PARP
PARG
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/44182

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repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Disrupted ADP-ribose metabolism with nuclear Poly (ADP-ribose) accumulation leads to different cell death pathways in presence of hydrogen peroxide in procyclic Trypanosoma bruceiSchlesinger, MarianaVilchez Larrea, Salomé CatalinaHaikarainen, TeemuNarwal, MohitVenkannagari, HarikanthFlawia, Mirtha MariaLehtiö, LariFernandez Villamil, Silvia HebeTrypanosoma bruceiPARPARPPARGhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Poly(ADP-ribose) (PAR) metabolism participates in several biological processes such as DNA damage signaling 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 higher eukaryotes, Trypanosoma brucei contains only one PARP and PARG. Up to date, the function of these enzymes has remained elusive in this parasite. The aim of this work is to unravel the role that PAR plays in genotoxic stress response.MethodsThe optimal conditions for the activity of purified recombinant TbPARP were determined by using a fluorometric activity assay followed by screening of PARP inhibitors. Sensitivity to a genotoxic agent, H2O2, was assessed by counting motile parasites over the total number in a Neubauer chamber, in presence of a potent PARP inhibitor 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 Anova followed by Bonferroni test. Finally, PAR influence was studied in cell death pathways by flow cytometry.ResultsAbolition of a functional PARP either by using potent inhibitors present or in PARP-silenced parasites had no effect on parasite growth in culture; however, PARP-inhibited and PARP down-regulated parasites presented an increased resistance against H2O2 treatment when compared to their wild type counterparts. PARP over-expressing and PARG-silenced parasites displayed polymer accumulation in the nucleus and, as expected, showed diminished resistance when exposed to the same genotoxic stimulus. Indeed, they suffered a necrotic death pathway, while an apoptosis-like mechanism was observed in control cultures. Surprisingly, PARP migrated to the nucleus and synthesized PAR only after a genomic stress in wild type parasites while PARG occurred always in this organelle.ConclusionsPARP over-expressing and PARG-silenced cells presented PAR accumulation in the nucleus, even in absence of oxidative stress. Procyclic death pathway after genotoxic damage depends on basal nuclear PAR. This evidence demonstrates that the polymer may have a toxic action by itself since the consequences of an exacerbated PARP activity cannot fully explain the increment in sensitivity observed here. Moreover, the unusual localization of PARP and PARG would reveal a novel regulatory mechanism, making them invaluable model systems.Fil: Schlesinger, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; ArgentinaFil: Vilchez Larrea, Salomé Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; ArgentinaFil: Haikarainen, Teemu. University of Oulu; FinlandiaFil: Narwal, Mohit. University of Oulu; FinlandiaFil: Venkannagari, Harikanth. University of Oulu; FinlandiaFil: Flawia, Mirtha Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; ArgentinaFil: Lehtiö, Lari. University of Oulu; FinlandiaFil: Fernandez Villamil, Silvia Hebe. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; ArgentinaBioMed Central2016-03info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/44182Schlesinger, Mariana; Vilchez Larrea, Salomé Catalina; Haikarainen, Teemu; Narwal, Mohit; Venkannagari, Harikanth; et al.; 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; BioMed Central; Parasites and Vectors; 9; 173; 3-2016; 1-151756-3305CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/http://parasitesandvectors.biomedcentral.com/articles/10.1186/s13071-016-1461-1info:eu-repo/semantics/altIdentifier/doi/10.1186/s13071-016-1461-1info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:27:03Zoai:ri.conicet.gov.ar:11336/44182instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 10:27:04.082CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv 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
title 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
spellingShingle 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
Schlesinger, Mariana
Trypanosoma brucei
PAR
PARP
PARG
title_short 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
title_full 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
title_fullStr 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
title_full_unstemmed 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
title_sort 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
dc.creator.none.fl_str_mv Schlesinger, Mariana
Vilchez Larrea, Salomé Catalina
Haikarainen, Teemu
Narwal, Mohit
Venkannagari, Harikanth
Flawia, Mirtha Maria
Lehtiö, Lari
Fernandez Villamil, Silvia Hebe
author Schlesinger, Mariana
author_facet Schlesinger, Mariana
Vilchez Larrea, Salomé Catalina
Haikarainen, Teemu
Narwal, Mohit
Venkannagari, Harikanth
Flawia, Mirtha Maria
Lehtiö, Lari
Fernandez Villamil, Silvia Hebe
author_role author
author2 Vilchez Larrea, Salomé Catalina
Haikarainen, Teemu
Narwal, Mohit
Venkannagari, Harikanth
Flawia, Mirtha Maria
Lehtiö, Lari
Fernandez Villamil, Silvia Hebe
author2_role author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Trypanosoma brucei
PAR
PARP
PARG
topic Trypanosoma brucei
PAR
PARP
PARG
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Poly(ADP-ribose) (PAR) metabolism participates in several biological processes such as DNA damage signaling 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 higher eukaryotes, Trypanosoma brucei contains only one PARP and PARG. Up to date, the function of these enzymes has remained elusive in this parasite. The aim of this work is to unravel the role that PAR plays in genotoxic stress response.MethodsThe optimal conditions for the activity of purified recombinant TbPARP were determined by using a fluorometric activity assay followed by screening of PARP inhibitors. Sensitivity to a genotoxic agent, H2O2, was assessed by counting motile parasites over the total number in a Neubauer chamber, in presence of a potent PARP inhibitor 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 Anova followed by Bonferroni test. Finally, PAR influence was studied in cell death pathways by flow cytometry.ResultsAbolition of a functional PARP either by using potent inhibitors present or in PARP-silenced parasites had no effect on parasite growth in culture; however, PARP-inhibited and PARP down-regulated parasites presented an increased resistance against H2O2 treatment when compared to their wild type counterparts. PARP over-expressing and PARG-silenced parasites displayed polymer accumulation in the nucleus and, as expected, showed diminished resistance when exposed to the same genotoxic stimulus. Indeed, they suffered a necrotic death pathway, while an apoptosis-like mechanism was observed in control cultures. Surprisingly, PARP migrated to the nucleus and synthesized PAR only after a genomic stress in wild type parasites while PARG occurred always in this organelle.ConclusionsPARP over-expressing and PARG-silenced cells presented PAR accumulation in the nucleus, even in absence of oxidative stress. Procyclic death pathway after genotoxic damage depends on basal nuclear PAR. This evidence demonstrates that the polymer may have a toxic action by itself since the consequences of an exacerbated PARP activity cannot fully explain the increment in sensitivity observed here. Moreover, the unusual localization of PARP and PARG would reveal a novel regulatory mechanism, making them invaluable model systems.
Fil: Schlesinger, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
Fil: Vilchez Larrea, Salomé Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
Fil: Haikarainen, Teemu. University of Oulu; Finlandia
Fil: Narwal, Mohit. University of Oulu; Finlandia
Fil: Venkannagari, Harikanth. University of Oulu; Finlandia
Fil: Flawia, Mirtha Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
Fil: Lehtiö, Lari. University of Oulu; Finlandia
Fil: Fernandez Villamil, Silvia Hebe. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular ; Argentina
description Poly(ADP-ribose) (PAR) metabolism participates in several biological processes such as DNA damage signaling 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 higher eukaryotes, Trypanosoma brucei contains only one PARP and PARG. Up to date, the function of these enzymes has remained elusive in this parasite. The aim of this work is to unravel the role that PAR plays in genotoxic stress response.MethodsThe optimal conditions for the activity of purified recombinant TbPARP were determined by using a fluorometric activity assay followed by screening of PARP inhibitors. Sensitivity to a genotoxic agent, H2O2, was assessed by counting motile parasites over the total number in a Neubauer chamber, in presence of a potent PARP inhibitor 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 Anova followed by Bonferroni test. Finally, PAR influence was studied in cell death pathways by flow cytometry.ResultsAbolition of a functional PARP either by using potent inhibitors present or in PARP-silenced parasites had no effect on parasite growth in culture; however, PARP-inhibited and PARP down-regulated parasites presented an increased resistance against H2O2 treatment when compared to their wild type counterparts. PARP over-expressing and PARG-silenced parasites displayed polymer accumulation in the nucleus and, as expected, showed diminished resistance when exposed to the same genotoxic stimulus. Indeed, they suffered a necrotic death pathway, while an apoptosis-like mechanism was observed in control cultures. Surprisingly, PARP migrated to the nucleus and synthesized PAR only after a genomic stress in wild type parasites while PARG occurred always in this organelle.ConclusionsPARP over-expressing and PARG-silenced cells presented PAR accumulation in the nucleus, even in absence of oxidative stress. Procyclic death pathway after genotoxic damage depends on basal nuclear PAR. This evidence demonstrates that the polymer may have a toxic action by itself since the consequences of an exacerbated PARP activity cannot fully explain the increment in sensitivity observed here. Moreover, the unusual localization of PARP and PARG would reveal a novel regulatory mechanism, making them invaluable model systems.
publishDate 2016
dc.date.none.fl_str_mv 2016-03
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/44182
Schlesinger, Mariana; Vilchez Larrea, Salomé Catalina; Haikarainen, Teemu; Narwal, Mohit; Venkannagari, Harikanth; et al.; 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; BioMed Central; Parasites and Vectors; 9; 173; 3-2016; 1-15
1756-3305
CONICET Digital
CONICET
url http://hdl.handle.net/11336/44182
identifier_str_mv Schlesinger, Mariana; Vilchez Larrea, Salomé Catalina; Haikarainen, Teemu; Narwal, Mohit; Venkannagari, Harikanth; et al.; 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; BioMed Central; Parasites and Vectors; 9; 173; 3-2016; 1-15
1756-3305
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/http://parasitesandvectors.biomedcentral.com/articles/10.1186/s13071-016-1461-1
info:eu-repo/semantics/altIdentifier/doi/10.1186/s13071-016-1461-1
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https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
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dc.publisher.none.fl_str_mv BioMed Central
publisher.none.fl_str_mv BioMed Central
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
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reponame_str CONICET Digital (CONICET)
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instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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