Imaging of the alphavirus capsid protein during virus replication

Yan Zheng, Margaret Kielian

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Alphaviruses are enveloped viruses with highly organized structures. The nucleocapsid (NC) core contains a capsid protein lattice enclosing the plus-sense RNA genome, and it is surrounded by a lipid bilayer containing a lattice of the E1 and E2 envelope glycoproteins. Capsid protein is synthesized in the cytoplasm and particle budding occurs at the plasma membrane (PM), but the traffic and assembly of viral components and the exit of virions from host cells are not well understood. To visualize the dynamics of capsid protein during infection, we developed a Sindbis virus infectious clone tagged with a tetracysteine motif. Tagged capsid protein could be fluorescently labeled with biarsenical dyes in living cells without effects on virus growth, morphology, or protein distribution. Live cell imaging and colocalization experiments defined distinct groups of capsid foci in infected cells. We observed highly motile internal puncta that colocalized with E2 protein, which may represent the transport machinery that capsid protein uses to reach the PM. Capsid was also found in larger nonmotile internal structures that colocalized with cellular G3BP and viral nsP3. Thus, capsid may play an unforeseen role in these previously observed G3BP-positive foci, such as regulation of cellular stress granules. Capsid puncta were also observed at the PM. These puncta colocalized with E2 and recruited newly synthesized capsid protein; thus, they may be sites of virus assembly and egress. Together, our studies provide the first dynamic views of the alphavirus capsid protein in living cells and a system to define detailed mechanisms during alphavirus infection.

Original languageEnglish (US)
Pages (from-to)9579-9589
Number of pages11
JournalJournal of Virology
Volume87
Issue number17
DOIs
StatePublished - 2013

Fingerprint

Alphavirus
Capsid Proteins
Virus Replication
virus replication
coat proteins
image analysis
capsid
Capsid
plasma membrane
Cell Membrane
Alphavirus Infections
cells
Sindbis virus
Sindbis Virus
Viruses
Virus Release
Virus Assembly
Nucleocapsid
nucleocapsid
Viral Structures

ASJC Scopus subject areas

  • Immunology
  • Virology

Cite this

Imaging of the alphavirus capsid protein during virus replication. / Zheng, Yan; Kielian, Margaret.

In: Journal of Virology, Vol. 87, No. 17, 2013, p. 9579-9589.

Research output: Contribution to journalArticle

@article{a1fe90fdf6534841bfd6d620945e9a5e,
title = "Imaging of the alphavirus capsid protein during virus replication",
abstract = "Alphaviruses are enveloped viruses with highly organized structures. The nucleocapsid (NC) core contains a capsid protein lattice enclosing the plus-sense RNA genome, and it is surrounded by a lipid bilayer containing a lattice of the E1 and E2 envelope glycoproteins. Capsid protein is synthesized in the cytoplasm and particle budding occurs at the plasma membrane (PM), but the traffic and assembly of viral components and the exit of virions from host cells are not well understood. To visualize the dynamics of capsid protein during infection, we developed a Sindbis virus infectious clone tagged with a tetracysteine motif. Tagged capsid protein could be fluorescently labeled with biarsenical dyes in living cells without effects on virus growth, morphology, or protein distribution. Live cell imaging and colocalization experiments defined distinct groups of capsid foci in infected cells. We observed highly motile internal puncta that colocalized with E2 protein, which may represent the transport machinery that capsid protein uses to reach the PM. Capsid was also found in larger nonmotile internal structures that colocalized with cellular G3BP and viral nsP3. Thus, capsid may play an unforeseen role in these previously observed G3BP-positive foci, such as regulation of cellular stress granules. Capsid puncta were also observed at the PM. These puncta colocalized with E2 and recruited newly synthesized capsid protein; thus, they may be sites of virus assembly and egress. Together, our studies provide the first dynamic views of the alphavirus capsid protein in living cells and a system to define detailed mechanisms during alphavirus infection.",
author = "Yan Zheng and Margaret Kielian",
year = "2013",
doi = "10.1128/JVI.01299-13",
language = "English (US)",
volume = "87",
pages = "9579--9589",
journal = "Journal of Virology",
issn = "0022-538X",
publisher = "American Society for Microbiology",
number = "17",

}

TY - JOUR

T1 - Imaging of the alphavirus capsid protein during virus replication

AU - Zheng, Yan

AU - Kielian, Margaret

PY - 2013

Y1 - 2013

N2 - Alphaviruses are enveloped viruses with highly organized structures. The nucleocapsid (NC) core contains a capsid protein lattice enclosing the plus-sense RNA genome, and it is surrounded by a lipid bilayer containing a lattice of the E1 and E2 envelope glycoproteins. Capsid protein is synthesized in the cytoplasm and particle budding occurs at the plasma membrane (PM), but the traffic and assembly of viral components and the exit of virions from host cells are not well understood. To visualize the dynamics of capsid protein during infection, we developed a Sindbis virus infectious clone tagged with a tetracysteine motif. Tagged capsid protein could be fluorescently labeled with biarsenical dyes in living cells without effects on virus growth, morphology, or protein distribution. Live cell imaging and colocalization experiments defined distinct groups of capsid foci in infected cells. We observed highly motile internal puncta that colocalized with E2 protein, which may represent the transport machinery that capsid protein uses to reach the PM. Capsid was also found in larger nonmotile internal structures that colocalized with cellular G3BP and viral nsP3. Thus, capsid may play an unforeseen role in these previously observed G3BP-positive foci, such as regulation of cellular stress granules. Capsid puncta were also observed at the PM. These puncta colocalized with E2 and recruited newly synthesized capsid protein; thus, they may be sites of virus assembly and egress. Together, our studies provide the first dynamic views of the alphavirus capsid protein in living cells and a system to define detailed mechanisms during alphavirus infection.

AB - Alphaviruses are enveloped viruses with highly organized structures. The nucleocapsid (NC) core contains a capsid protein lattice enclosing the plus-sense RNA genome, and it is surrounded by a lipid bilayer containing a lattice of the E1 and E2 envelope glycoproteins. Capsid protein is synthesized in the cytoplasm and particle budding occurs at the plasma membrane (PM), but the traffic and assembly of viral components and the exit of virions from host cells are not well understood. To visualize the dynamics of capsid protein during infection, we developed a Sindbis virus infectious clone tagged with a tetracysteine motif. Tagged capsid protein could be fluorescently labeled with biarsenical dyes in living cells without effects on virus growth, morphology, or protein distribution. Live cell imaging and colocalization experiments defined distinct groups of capsid foci in infected cells. We observed highly motile internal puncta that colocalized with E2 protein, which may represent the transport machinery that capsid protein uses to reach the PM. Capsid was also found in larger nonmotile internal structures that colocalized with cellular G3BP and viral nsP3. Thus, capsid may play an unforeseen role in these previously observed G3BP-positive foci, such as regulation of cellular stress granules. Capsid puncta were also observed at the PM. These puncta colocalized with E2 and recruited newly synthesized capsid protein; thus, they may be sites of virus assembly and egress. Together, our studies provide the first dynamic views of the alphavirus capsid protein in living cells and a system to define detailed mechanisms during alphavirus infection.

UR - http://www.scopus.com/inward/record.url?scp=84883285628&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84883285628&partnerID=8YFLogxK

U2 - 10.1128/JVI.01299-13

DO - 10.1128/JVI.01299-13

M3 - Article

C2 - 23785213

AN - SCOPUS:84883285628

VL - 87

SP - 9579

EP - 9589

JO - Journal of Virology

JF - Journal of Virology

SN - 0022-538X

IS - 17

ER -