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Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles

BACKGROUND: Paramyxoviruses are assembled at the plasma membrane budding sites after synthesis of all the structural components in the cytoplasm. Although viral ribonuclocapsid (vRNP) is an essential component of infectious virions, the process of vRNP translocation to assembly sites is poorly under...

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Detalles Bibliográficos
Autores principales: Chambers, Raychel, Takimoto, Toru
Formato: Texto
Lenguaje:English
Publicado: Public Library of Science 2010
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2881874/
https://www.ncbi.nlm.nih.gov/pubmed/20543880
http://dx.doi.org/10.1371/journal.pone.0010994
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author Chambers, Raychel
Takimoto, Toru
author_facet Chambers, Raychel
Takimoto, Toru
author_sort Chambers, Raychel
collection PubMed
description BACKGROUND: Paramyxoviruses are assembled at the plasma membrane budding sites after synthesis of all the structural components in the cytoplasm. Although viral ribonuclocapsid (vRNP) is an essential component of infectious virions, the process of vRNP translocation to assembly sites is poorly understood. METHODOLOGY/PRINCIPAL FINDINGS: To analyze real-time trafficking of vRNPs in live infected cells, we created a recombinant Sendai virus (SeV), rSeVLeGFP, which expresses L protein fused to enhanced green fluorescent protein (eGFP). The rSeVLeGFP showed similar growth kinetics compared to wt SeV, and newly synthesized LeGFP could be detected as early as 8 h postinfection. The majority of LeGFP co-localized with other components of vRNPs, NP and P proteins, suggesting the fluorescent signals of LeGFP represent the locations of vRNPs. Analysis of LeGFP movement using time-lapse digital video microscopy revealed directional and saltatory movement of LeGFP along microtubules. Treatment of the cells with nocodazole restricted vRNP movement and reduced progeny virion production without affecting viral protein synthesis, suggesting the role of microtubules in vRNP trafficking and virus assembly. Further study with an electron microscope showed close association of vRNPs with intracellular vesicles present in infected cells. In addition, the vRNPs co-localized with Rab11a protein, which is known to regulate the recycling endocytosis pathway and Golgi-to-plasma membrane trafficking. Simultaneous movement between LeGFP and Rab11a was also observed in infected cells, which constitutively express mRFP-tagged Rab11a. Involvement of recycling endosomes in vRNP translocation was also suggested by the fact that vRNPs move concomitantly with recycling transferrin labeled with Alexa 594. CONCLUSIONS/SIGNIFICANCE: Collectively, our results strongly suggest a previously unrecognized involvement of the intracellular vesicular trafficking pathway in vRNP translocation and provide new insights into the transport of viral structural components to the assembly sites of enveloped viruses.
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spelling pubmed-28818742010-06-11 Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles Chambers, Raychel Takimoto, Toru PLoS One Research Article BACKGROUND: Paramyxoviruses are assembled at the plasma membrane budding sites after synthesis of all the structural components in the cytoplasm. Although viral ribonuclocapsid (vRNP) is an essential component of infectious virions, the process of vRNP translocation to assembly sites is poorly understood. METHODOLOGY/PRINCIPAL FINDINGS: To analyze real-time trafficking of vRNPs in live infected cells, we created a recombinant Sendai virus (SeV), rSeVLeGFP, which expresses L protein fused to enhanced green fluorescent protein (eGFP). The rSeVLeGFP showed similar growth kinetics compared to wt SeV, and newly synthesized LeGFP could be detected as early as 8 h postinfection. The majority of LeGFP co-localized with other components of vRNPs, NP and P proteins, suggesting the fluorescent signals of LeGFP represent the locations of vRNPs. Analysis of LeGFP movement using time-lapse digital video microscopy revealed directional and saltatory movement of LeGFP along microtubules. Treatment of the cells with nocodazole restricted vRNP movement and reduced progeny virion production without affecting viral protein synthesis, suggesting the role of microtubules in vRNP trafficking and virus assembly. Further study with an electron microscope showed close association of vRNPs with intracellular vesicles present in infected cells. In addition, the vRNPs co-localized with Rab11a protein, which is known to regulate the recycling endocytosis pathway and Golgi-to-plasma membrane trafficking. Simultaneous movement between LeGFP and Rab11a was also observed in infected cells, which constitutively express mRFP-tagged Rab11a. Involvement of recycling endosomes in vRNP translocation was also suggested by the fact that vRNPs move concomitantly with recycling transferrin labeled with Alexa 594. CONCLUSIONS/SIGNIFICANCE: Collectively, our results strongly suggest a previously unrecognized involvement of the intracellular vesicular trafficking pathway in vRNP translocation and provide new insights into the transport of viral structural components to the assembly sites of enveloped viruses. Public Library of Science 2010-06-07 /pmc/articles/PMC2881874/ /pubmed/20543880 http://dx.doi.org/10.1371/journal.pone.0010994 Text en Chambers, Takimoto. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Chambers, Raychel
Takimoto, Toru
Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles
title Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles
title_full Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles
title_fullStr Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles
title_full_unstemmed Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles
title_short Trafficking of Sendai Virus Nucleocapsids Is Mediated by Intracellular Vesicles
title_sort trafficking of sendai virus nucleocapsids is mediated by intracellular vesicles
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2881874/
https://www.ncbi.nlm.nih.gov/pubmed/20543880
http://dx.doi.org/10.1371/journal.pone.0010994
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