Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action

Detalhes bibliográficos
Autor(a) principal: Freire, João M.
Data de Publicação: 2013
Outros Autores: Veiga, Ana Salomé, Figueiredo, Inês Rego de, Torre, Beatriz G. de la, Santos, Nuno C., Andreu, David, Poian, Andrea T. Da, Castanho, Miguel A. R. B.
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10451/10714
Resumo: © 2013 FEBS
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spelling Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of actionCapsid proteinCell penetrating peptideDengue virusGene therapyTime-resolvedFlow cytometry© 2013 FEBSCell penetrating peptides (CPPs) can be used as drug delivery systems for different therapeutic molecules. In this work two novel CPPs, pepR and pepM, designed from two domains of the dengue virus (DENV) capsid protein, were studied for their ability to deliver nucleic acids into cells as non-covalently bound cargo. Translocation studies were performed by confocal microscopy in HepG2, BHK and HEK cell lineages, astrocytes and peripheral blood mononuclear cells. Combined studies in HepG2 cells, astrocytes and BHK cells, at 4 and 37 °C or using specific endocytosis inhibitors, revealed that pepR and pepM use distinct internalization routes: pepM translocates lipid membranes directly, while pepR uses an endocytic pathway. To confirm these results, a methodology was developed to monitor the translocation kinetics of both peptides by real-time flow cytometry. Kinetic constants were determined, and the amount of nucleic acids delivered was estimated. Additional studies were performed in order to understand the molecular bases of the peptide-mediated translocation. Peptide– nucleic acid and peptide–lipid membrane interactions were studied quantitatively based on the intrinsic fluorescence of the peptides. pepR and pepM bound ssDNA to the same extent. Partition studies revealed that both peptides bind preferentially to anionic lipid membranes, adopting an α-helical conformation. However, fluorescence quenching studies suggest that pepM is deeply inserted into the lipid bilayer, in contrast with pepR. Moreover, only pepM is able to promote the fusion and aggregation of vesicles composed of zwitterionic lipids. Altogether, the results show that DENV capsid protein derived peptides serve as good templates for novel CPP-based nucleic acid delivery strategies, defining different routes for cell entry.This work was supported by Fundação para a Ciência e Tecnologia – Ministério da Educação e Ciência (FCT-MEC, Portugal) (PTDC/QUI-BIQ/112929/2009), by the European Union [projects FP7-PEOPLE IRSES MEMPEPACROSS and FP7-HEALTH-F3-2008-223414 (LEISHDRUG)], by the Spanish Ministry of Economy and Competitiveness (SAF2011-24899), by the Generalitat de Catalunya (2009 SGR 492), by the Brazilian Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), by Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ) and by the National Institute of Science and Technology in Dengue (INCTDengue). JMF also acknowledges FCT-MEC for PhD fellowship SFRH/BD/70423/2010. MC acknowledges a grant from Programa Ciência Sem Fronteiras PVE171/2012 (CAPES and CNPq, Brazil). The authors thank Sónia Sá Santos (IMM, Lisbon, Portugal) for the kind gift of the primary astrocyte cultures.WileyRepositório da Universidade de LisboaFreire, João M.Veiga, Ana SaloméFigueiredo, Inês Rego deTorre, Beatriz G. de laSantos, Nuno C.Andreu, DavidPoian, Andrea T. DaCastanho, Miguel A. R. B.2014-03-10T14:06:57Z20132013-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10451/10714engFEBS Journal 281 (2014) 191–2151742-464Xhttp://dx.doi.org/10.1111/febs.12587metadata only accessinfo:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-11-08T15:56:22Zoai:repositorio.ul.pt:10451/10714Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-19T21:34:38.222315Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse
dc.title.none.fl_str_mv Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
title Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
spellingShingle Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
Freire, João M.
Capsid protein
Cell penetrating peptide
Dengue virus
Gene therapy
Time-resolved
Flow cytometry
title_short Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
title_full Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
title_fullStr Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
title_full_unstemmed Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
title_sort Nucleic acid delivery by cell penetrating peptides derived from dengue virus capsid protein : design and mechanism of action
author Freire, João M.
author_facet Freire, João M.
Veiga, Ana Salomé
Figueiredo, Inês Rego de
Torre, Beatriz G. de la
Santos, Nuno C.
Andreu, David
Poian, Andrea T. Da
Castanho, Miguel A. R. B.
author_role author
author2 Veiga, Ana Salomé
Figueiredo, Inês Rego de
Torre, Beatriz G. de la
Santos, Nuno C.
Andreu, David
Poian, Andrea T. Da
Castanho, Miguel A. R. B.
author2_role author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Repositório da Universidade de Lisboa
dc.contributor.author.fl_str_mv Freire, João M.
Veiga, Ana Salomé
Figueiredo, Inês Rego de
Torre, Beatriz G. de la
Santos, Nuno C.
Andreu, David
Poian, Andrea T. Da
Castanho, Miguel A. R. B.
dc.subject.por.fl_str_mv Capsid protein
Cell penetrating peptide
Dengue virus
Gene therapy
Time-resolved
Flow cytometry
topic Capsid protein
Cell penetrating peptide
Dengue virus
Gene therapy
Time-resolved
Flow cytometry
description © 2013 FEBS
publishDate 2013
dc.date.none.fl_str_mv 2013
2013-01-01T00:00:00Z
2014-03-10T14:06:57Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/article
format article
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://hdl.handle.net/10451/10714
url http://hdl.handle.net/10451/10714
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv FEBS Journal 281 (2014) 191–215
1742-464X
http://dx.doi.org/10.1111/febs.12587
dc.rights.driver.fl_str_mv metadata only access
info:eu-repo/semantics/openAccess
rights_invalid_str_mv metadata only access
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv Wiley
publisher.none.fl_str_mv Wiley
dc.source.none.fl_str_mv reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
instacron:RCAAP
instname_str Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
instacron_str RCAAP
institution RCAAP
reponame_str Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
collection Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
repository.name.fl_str_mv Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
repository.mail.fl_str_mv
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