Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity

Autores
D'antuono, Alejandra Lorena; Loureiro, Maria Eugenia; Foscaldi, Sabrina Andrea; Marino, Cristina Ester; Lopez, Nora Mabel
Año de publicación
2014
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
The arenavirus nucleoprotein (NP) is the main protein component of viral nucleocapsids and is strictly required for viral genome replication mediated by the L polymerase. Homo-oligomerization of NP is presumed to play an important role in nucleocapsid assembly, albeit the underlying mechanism and the relevance of NP-NP interaction in nucleocapsid activity are still poorly understood. Here, we evaluate the contribution of the New World Tacaribe virus (TCRV) NP self-interaction to nucleocapsid functional activity. We show that alanine substitution of N-terminal residues predicted to be available for NP-NP interaction strongly affected NP self-association, as determined by coimmunoprecipitation assays, produced a drastic inhibition of transcription and replication of a TCRV minigenome RNA, and impaired NP binding to RNA. Mutagenesis and functional analysis also revealed that, while dispensable for NP self-interaction, key amino acids at the C-terminal domain were essential for RNA synthesis. Furthermore, mutations at these C-terminal residues rendered NP unable to bind RNA both in vivo and in vitro but had no effect on the interaction with the L polymerase. In addition, while all oligomerization-defective variants tested exhibited unaltered capacities to sustain NP-L interaction, NP deletion mutants were fully incompetent to bind L, suggesting that, whereas NP self-association is dispensable, the integrity of both the N-terminal and C-terminal domains is required for binding the L polymerase. Overall, our results suggest that NP self-interaction mediated by the N-terminal domain may play a critical role in TCRV nucleocapsid assembly and activity and that the C-terminal domain of NP is implicated in RNA binding. IMPORTANCE: The mechanism of arenavirus functional nucleocapsid assembly is still poorly understood. No detailed information is available on the nucleocapsid structure, and the regions of full-length NP involved in binding to viral RNA remain to be determined. In this report, novel findings are provided on critical interactions between the viral ribonucleoprotein components. We identify several amino acid residues in both the N-terminal and C-terminal domains of TCRV NP that differentially contribute to NP-NP and NP-RNA interactions and analyze their relevance for binding of NP to the L polymerase and for nucleocapsid activity. Our results provide insight into the contribution of NP self-interaction to RNP assembly and activity and reveal the involvement of the NP C-terminal domain in RNA binding.
Fil: D'antuono, Alejandra Lorena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
Fil: Loureiro, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
Fil: Foscaldi, Sabrina Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
Fil: Marino, Cristina Ester. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina
Fil: Lopez, Nora Mabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
Materia
Tcrv
Nucleoprotein
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:11336/19535

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network_name_str CONICET Digital (CONICET)
spelling Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activityD'antuono, Alejandra LorenaLoureiro, Maria EugeniaFoscaldi, Sabrina AndreaMarino, Cristina EsterLopez, Nora MabelTcrvNucleoproteinhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1The arenavirus nucleoprotein (NP) is the main protein component of viral nucleocapsids and is strictly required for viral genome replication mediated by the L polymerase. Homo-oligomerization of NP is presumed to play an important role in nucleocapsid assembly, albeit the underlying mechanism and the relevance of NP-NP interaction in nucleocapsid activity are still poorly understood. Here, we evaluate the contribution of the New World Tacaribe virus (TCRV) NP self-interaction to nucleocapsid functional activity. We show that alanine substitution of N-terminal residues predicted to be available for NP-NP interaction strongly affected NP self-association, as determined by coimmunoprecipitation assays, produced a drastic inhibition of transcription and replication of a TCRV minigenome RNA, and impaired NP binding to RNA. Mutagenesis and functional analysis also revealed that, while dispensable for NP self-interaction, key amino acids at the C-terminal domain were essential for RNA synthesis. Furthermore, mutations at these C-terminal residues rendered NP unable to bind RNA both in vivo and in vitro but had no effect on the interaction with the L polymerase. In addition, while all oligomerization-defective variants tested exhibited unaltered capacities to sustain NP-L interaction, NP deletion mutants were fully incompetent to bind L, suggesting that, whereas NP self-association is dispensable, the integrity of both the N-terminal and C-terminal domains is required for binding the L polymerase. Overall, our results suggest that NP self-interaction mediated by the N-terminal domain may play a critical role in TCRV nucleocapsid assembly and activity and that the C-terminal domain of NP is implicated in RNA binding. IMPORTANCE: The mechanism of arenavirus functional nucleocapsid assembly is still poorly understood. No detailed information is available on the nucleocapsid structure, and the regions of full-length NP involved in binding to viral RNA remain to be determined. In this report, novel findings are provided on critical interactions between the viral ribonucleoprotein components. We identify several amino acid residues in both the N-terminal and C-terminal domains of TCRV NP that differentially contribute to NP-NP and NP-RNA interactions and analyze their relevance for binding of NP to the L polymerase and for nucleocapsid activity. Our results provide insight into the contribution of NP self-interaction to RNP assembly and activity and reveal the involvement of the NP C-terminal domain in RNA binding.Fil: D'antuono, Alejandra Lorena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; ArgentinaFil: Loureiro, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; ArgentinaFil: Foscaldi, Sabrina Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; ArgentinaFil: Marino, Cristina Ester. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Lopez, Nora Mabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; ArgentinaAmerican Society For Microbiology2014-06info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/vnd.openxmlformats-officedocument.wordprocessingml.documentapplication/pdfhttp://hdl.handle.net/11336/19535D'antuono, Alejandra Lorena; Loureiro, Maria Eugenia; Foscaldi, Sabrina Andrea; Marino, Cristina Ester; Lopez, Nora Mabel; Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity; American Society For Microbiology; Journal Of Virology; 88; 11; 6-2014; 6492-65050022-538X1098-5514CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/http://jvi.asm.org/content/88/11/6492.longinfo:eu-repo/semantics/altIdentifier/doi/10.1128/JVI.00321-14info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:17:00Zoai:ri.conicet.gov.ar:11336/19535instacron: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 10:17:00.863CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
title Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
spellingShingle Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
D'antuono, Alejandra Lorena
Tcrv
Nucleoprotein
title_short Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
title_full Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
title_fullStr Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
title_full_unstemmed Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
title_sort Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity
dc.creator.none.fl_str_mv D'antuono, Alejandra Lorena
Loureiro, Maria Eugenia
Foscaldi, Sabrina Andrea
Marino, Cristina Ester
Lopez, Nora Mabel
author D'antuono, Alejandra Lorena
author_facet D'antuono, Alejandra Lorena
Loureiro, Maria Eugenia
Foscaldi, Sabrina Andrea
Marino, Cristina Ester
Lopez, Nora Mabel
author_role author
author2 Loureiro, Maria Eugenia
Foscaldi, Sabrina Andrea
Marino, Cristina Ester
Lopez, Nora Mabel
author2_role author
author
author
author
dc.subject.none.fl_str_mv Tcrv
Nucleoprotein
topic Tcrv
Nucleoprotein
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv The arenavirus nucleoprotein (NP) is the main protein component of viral nucleocapsids and is strictly required for viral genome replication mediated by the L polymerase. Homo-oligomerization of NP is presumed to play an important role in nucleocapsid assembly, albeit the underlying mechanism and the relevance of NP-NP interaction in nucleocapsid activity are still poorly understood. Here, we evaluate the contribution of the New World Tacaribe virus (TCRV) NP self-interaction to nucleocapsid functional activity. We show that alanine substitution of N-terminal residues predicted to be available for NP-NP interaction strongly affected NP self-association, as determined by coimmunoprecipitation assays, produced a drastic inhibition of transcription and replication of a TCRV minigenome RNA, and impaired NP binding to RNA. Mutagenesis and functional analysis also revealed that, while dispensable for NP self-interaction, key amino acids at the C-terminal domain were essential for RNA synthesis. Furthermore, mutations at these C-terminal residues rendered NP unable to bind RNA both in vivo and in vitro but had no effect on the interaction with the L polymerase. In addition, while all oligomerization-defective variants tested exhibited unaltered capacities to sustain NP-L interaction, NP deletion mutants were fully incompetent to bind L, suggesting that, whereas NP self-association is dispensable, the integrity of both the N-terminal and C-terminal domains is required for binding the L polymerase. Overall, our results suggest that NP self-interaction mediated by the N-terminal domain may play a critical role in TCRV nucleocapsid assembly and activity and that the C-terminal domain of NP is implicated in RNA binding. IMPORTANCE: The mechanism of arenavirus functional nucleocapsid assembly is still poorly understood. No detailed information is available on the nucleocapsid structure, and the regions of full-length NP involved in binding to viral RNA remain to be determined. In this report, novel findings are provided on critical interactions between the viral ribonucleoprotein components. We identify several amino acid residues in both the N-terminal and C-terminal domains of TCRV NP that differentially contribute to NP-NP and NP-RNA interactions and analyze their relevance for binding of NP to the L polymerase and for nucleocapsid activity. Our results provide insight into the contribution of NP self-interaction to RNP assembly and activity and reveal the involvement of the NP C-terminal domain in RNA binding.
Fil: D'antuono, Alejandra Lorena. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
Fil: Loureiro, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
Fil: Foscaldi, Sabrina Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
Fil: Marino, Cristina Ester. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina
Fil: Lopez, Nora Mabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencias y Tecnología "dr. Cesar Milstein"; Argentina
description The arenavirus nucleoprotein (NP) is the main protein component of viral nucleocapsids and is strictly required for viral genome replication mediated by the L polymerase. Homo-oligomerization of NP is presumed to play an important role in nucleocapsid assembly, albeit the underlying mechanism and the relevance of NP-NP interaction in nucleocapsid activity are still poorly understood. Here, we evaluate the contribution of the New World Tacaribe virus (TCRV) NP self-interaction to nucleocapsid functional activity. We show that alanine substitution of N-terminal residues predicted to be available for NP-NP interaction strongly affected NP self-association, as determined by coimmunoprecipitation assays, produced a drastic inhibition of transcription and replication of a TCRV minigenome RNA, and impaired NP binding to RNA. Mutagenesis and functional analysis also revealed that, while dispensable for NP self-interaction, key amino acids at the C-terminal domain were essential for RNA synthesis. Furthermore, mutations at these C-terminal residues rendered NP unable to bind RNA both in vivo and in vitro but had no effect on the interaction with the L polymerase. In addition, while all oligomerization-defective variants tested exhibited unaltered capacities to sustain NP-L interaction, NP deletion mutants were fully incompetent to bind L, suggesting that, whereas NP self-association is dispensable, the integrity of both the N-terminal and C-terminal domains is required for binding the L polymerase. Overall, our results suggest that NP self-interaction mediated by the N-terminal domain may play a critical role in TCRV nucleocapsid assembly and activity and that the C-terminal domain of NP is implicated in RNA binding. IMPORTANCE: The mechanism of arenavirus functional nucleocapsid assembly is still poorly understood. No detailed information is available on the nucleocapsid structure, and the regions of full-length NP involved in binding to viral RNA remain to be determined. In this report, novel findings are provided on critical interactions between the viral ribonucleoprotein components. We identify several amino acid residues in both the N-terminal and C-terminal domains of TCRV NP that differentially contribute to NP-NP and NP-RNA interactions and analyze their relevance for binding of NP to the L polymerase and for nucleocapsid activity. Our results provide insight into the contribution of NP self-interaction to RNP assembly and activity and reveal the involvement of the NP C-terminal domain in RNA binding.
publishDate 2014
dc.date.none.fl_str_mv 2014-06
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/11336/19535
D'antuono, Alejandra Lorena; Loureiro, Maria Eugenia; Foscaldi, Sabrina Andrea; Marino, Cristina Ester; Lopez, Nora Mabel; Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity; American Society For Microbiology; Journal Of Virology; 88; 11; 6-2014; 6492-6505
0022-538X
1098-5514
CONICET Digital
CONICET
url http://hdl.handle.net/11336/19535
identifier_str_mv D'antuono, Alejandra Lorena; Loureiro, Maria Eugenia; Foscaldi, Sabrina Andrea; Marino, Cristina Ester; Lopez, Nora Mabel; Differential contribution of Tacaribe arenavirus Nucleoprotein N-terminal and C-terminal residues to nucleocapsid functional activity; American Society For Microbiology; Journal Of Virology; 88; 11; 6-2014; 6492-6505
0022-538X
1098-5514
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://jvi.asm.org/content/88/11/6492.long
info:eu-repo/semantics/altIdentifier/doi/10.1128/JVI.00321-14
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/vnd.openxmlformats-officedocument.wordprocessingml.document
application/pdf
dc.publisher.none.fl_str_mv American Society For Microbiology
publisher.none.fl_str_mv American Society For Microbiology
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
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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