Long-range RNA-RNA interactions circularize the dengue virus genome

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Secondary and tertiary RNA structures present in viral RNA genomes play essential regulatory roles during translation, RNA replication, and assembly of new viral particles. In the case of flaviviruses, RNA-RNA interactions between the 5′ and 3′ ends of the genome have been proposed to be required fo...

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Autores principales:	Alvarez, D.E., Lodeiro, M.F., Ludueña, S.J., Pietrasanta, L.I., Gamarnik, A.V.
Formato:	Artículo publishedVersion
Lenguaje:	Inglés
Publicado:	2005
Materias:	virus RNA article atomic force microscopy base pairing binding assay cyclization Dengue virus Flavivirus genetic transfection immunofluorescence in vitro study nonhuman priority journal protein nucleic acid interaction RNA binding RNA extraction RNA replication RNA sequence untranslated region virus genome virus mutation virus transcription Animals Base Sequence Binding Sites Cell Line Cricetinae Dengue Virus Microscopy, Atomic Force Models, Molecular Molecular Sequence Data Mutagenesis, Site-Directed Nucleic Acid Conformation RNA RNA, Viral Virus Replication
Acceso en línea:	http://hdl.handle.net/20.500.12110/paper_0022538X_v79_n11_p6631_Alvarez
Aporte de:	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires

id	paperaa:paper_0022538X_v79_n11_p6631_Alvarez
record_format	dspace
spelling	paperaa:paper_0022538X_v79_n11_p6631_Alvarez2023-06-12T16:44:52Z Long-range RNA-RNA interactions circularize the dengue virus genome J. Virol. 2005;79(11):6631-6643 Alvarez, D.E. Lodeiro, M.F. Ludueña, S.J. Pietrasanta, L.I. Gamarnik, A.V. virus RNA article atomic force microscopy base pairing binding assay cyclization Dengue virus Flavivirus genetic transfection immunofluorescence in vitro study nonhuman priority journal protein nucleic acid interaction RNA binding RNA extraction RNA replication RNA sequence untranslated region virus genome virus mutation virus transcription Animals Base Sequence Binding Sites Cell Line Cricetinae Dengue Virus Microscopy, Atomic Force Models, Molecular Molecular Sequence Data Mutagenesis, Site-Directed Nucleic Acid Conformation RNA RNA, Viral Virus Replication Dengue virus Secondary and tertiary RNA structures present in viral RNA genomes play essential regulatory roles during translation, RNA replication, and assembly of new viral particles. In the case of flaviviruses, RNA-RNA interactions between the 5′ and 3′ ends of the genome have been proposed to be required for RNA replication. We found that two RNA elements present at the ends of the dengue virus genome interact in vitro with high affinity. Visualization of individual molecules by atomic force microscopy reveled that physical interaction between these RNA elements results in cyclization of the viral RNA. Using RNA binding assays, we found that the putative cyclization sequences, known as 5′ and 3′ CS, present in all mosquito-borne flaviviruses, were necessary but not sufficient for RNA-RNA interaction. Additional sequences present at the 5′ and 3′ untranslated regions of the viral RNA were also required for RNA-RNA complex formation. We named these sequences 5′ and 3′ UAR (upstream AUG region). In order to investigate the functional role of 5′-3′ UAR complementarity, these sequences were mutated either separately, to destroy base pairing, or simultaneously, to restore complementarity in the context of full-length dengue virus RNA. Nonviable viruses were recovered after transfection of dengue virus RNA carrying mutations either at the 5′ or 3′ UAR, while the RNA containing the compensatory mutations was able to replicate. Since sequence complementarity between the ends of the genome is required for dengue virus viability, we propose that cyclization of the RNA is a required conformation for viral replication. Copyright © 2005, American Society for Microbiology. All Rights Reserved. 2005 info:eu-repo/semantics/article info:ar-repo/semantics/artículo info:eu-repo/semantics/publishedVersion application/pdf eng info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_0022538X_v79_n11_p6631_Alvarez
institution	Universidad de Buenos Aires
institution_str	I-28
repository_str	R-134
collection	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA)
language	Inglés
orig_language_str_mv	eng
topic	virus RNA article atomic force microscopy base pairing binding assay cyclization Dengue virus Flavivirus genetic transfection immunofluorescence in vitro study nonhuman priority journal protein nucleic acid interaction RNA binding RNA extraction RNA replication RNA sequence untranslated region virus genome virus mutation virus transcription Animals Base Sequence Binding Sites Cell Line Cricetinae Dengue Virus Microscopy, Atomic Force Models, Molecular Molecular Sequence Data Mutagenesis, Site-Directed Nucleic Acid Conformation RNA RNA, Viral Virus Replication Dengue virus
spellingShingle	virus RNA article atomic force microscopy base pairing binding assay cyclization Dengue virus Flavivirus genetic transfection immunofluorescence in vitro study nonhuman priority journal protein nucleic acid interaction RNA binding RNA extraction RNA replication RNA sequence untranslated region virus genome virus mutation virus transcription Animals Base Sequence Binding Sites Cell Line Cricetinae Dengue Virus Microscopy, Atomic Force Models, Molecular Molecular Sequence Data Mutagenesis, Site-Directed Nucleic Acid Conformation RNA RNA, Viral Virus Replication Dengue virus Alvarez, D.E. Lodeiro, M.F. Ludueña, S.J. Pietrasanta, L.I. Gamarnik, A.V. Long-range RNA-RNA interactions circularize the dengue virus genome
topic_facet	virus RNA article atomic force microscopy base pairing binding assay cyclization Dengue virus Flavivirus genetic transfection immunofluorescence in vitro study nonhuman priority journal protein nucleic acid interaction RNA binding RNA extraction RNA replication RNA sequence untranslated region virus genome virus mutation virus transcription Animals Base Sequence Binding Sites Cell Line Cricetinae Dengue Virus Microscopy, Atomic Force Models, Molecular Molecular Sequence Data Mutagenesis, Site-Directed Nucleic Acid Conformation RNA RNA, Viral Virus Replication Dengue virus
description	Secondary and tertiary RNA structures present in viral RNA genomes play essential regulatory roles during translation, RNA replication, and assembly of new viral particles. In the case of flaviviruses, RNA-RNA interactions between the 5′ and 3′ ends of the genome have been proposed to be required for RNA replication. We found that two RNA elements present at the ends of the dengue virus genome interact in vitro with high affinity. Visualization of individual molecules by atomic force microscopy reveled that physical interaction between these RNA elements results in cyclization of the viral RNA. Using RNA binding assays, we found that the putative cyclization sequences, known as 5′ and 3′ CS, present in all mosquito-borne flaviviruses, were necessary but not sufficient for RNA-RNA interaction. Additional sequences present at the 5′ and 3′ untranslated regions of the viral RNA were also required for RNA-RNA complex formation. We named these sequences 5′ and 3′ UAR (upstream AUG region). In order to investigate the functional role of 5′-3′ UAR complementarity, these sequences were mutated either separately, to destroy base pairing, or simultaneously, to restore complementarity in the context of full-length dengue virus RNA. Nonviable viruses were recovered after transfection of dengue virus RNA carrying mutations either at the 5′ or 3′ UAR, while the RNA containing the compensatory mutations was able to replicate. Since sequence complementarity between the ends of the genome is required for dengue virus viability, we propose that cyclization of the RNA is a required conformation for viral replication. Copyright © 2005, American Society for Microbiology. All Rights Reserved.
format	Artículo Artículo publishedVersion
author	Alvarez, D.E. Lodeiro, M.F. Ludueña, S.J. Pietrasanta, L.I. Gamarnik, A.V.
author_facet	Alvarez, D.E. Lodeiro, M.F. Ludueña, S.J. Pietrasanta, L.I. Gamarnik, A.V.
author_sort	Alvarez, D.E.
title	Long-range RNA-RNA interactions circularize the dengue virus genome
title_short	Long-range RNA-RNA interactions circularize the dengue virus genome
title_full	Long-range RNA-RNA interactions circularize the dengue virus genome
title_fullStr	Long-range RNA-RNA interactions circularize the dengue virus genome
title_full_unstemmed	Long-range RNA-RNA interactions circularize the dengue virus genome
title_sort	long-range rna-rna interactions circularize the dengue virus genome
publishDate	2005
url	http://hdl.handle.net/20.500.12110/paper_0022538X_v79_n11_p6631_Alvarez
work_keys_str_mv	AT alvarezde longrangernarnainteractionscircularizethedenguevirusgenome AT lodeiromf longrangernarnainteractionscircularizethedenguevirusgenome AT luduenasj longrangernarnainteractionscircularizethedenguevirusgenome AT pietrasantali longrangernarnainteractionscircularizethedenguevirusgenome AT gamarnikav longrangernarnainteractionscircularizethedenguevirusgenome
_version_	1769810278409568256

Long-range RNA-RNA interactions circularize the dengue virus genome

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