scholarly journals Role of Sindbis Virus Capsid Protein Region II in Nucleocapsid Core Assembly and Encapsidation of Genomic RNA

2008 ◽  
Vol 82 (9) ◽  
pp. 4461-4470 ◽  
Author(s):  
Ranjit Warrier ◽  
Benjamin R. Linger ◽  
Barbara L. Golden ◽  
Richard J. Kuhn
Keyword(s):  
Amino Acids ◽  
Capsid Protein ◽  
Sindbis Virus ◽  
Rna Virus ◽  
Genomic Rna ◽  
Viral Glycoproteins ◽  
Infected Cells ◽  
Viral Genomic Rna ◽  
Region Ii

ABSTRACT Sindbis virus is an enveloped positive-sense RNA virus in the alphavirus genus. The nucleocapsid core contains the genomic RNA surrounded by 240 copies of a single capsid protein. The capsid protein is multifunctional, and its roles include acting as a protease, controlling the specificity of RNA that is encapsidated into nucleocapsid cores, and interacting with viral glycoproteins to promote the budding of mature virus and the release of the genomic RNA into the newly infected cell. The region comprising amino acids 81 to 113 was previously implicated in two processes, the encapsidation of the viral genomic RNA and the stable accumulation of nucleocapsid cores in the cytoplasm of infected cells. In the present study, specific amino acids within this region responsible for the encapsidation of the genomic RNA have been identified. The region that is responsible for nucleocapsid core accumulation has considerable overlap with the region that controls encapsidation specificity.

scholarly journals Rubella virus-like replicon particles: analysis of encapsidation determinants and non-structural roles of capsid protein in early post-entry replication

2012 ◽  
Vol 93 (3) ◽  
pp. 516-525 ◽  
Author(s):  
Claudia Claus ◽  
Wen-Pin Tzeng ◽  
U. G Liebert ◽  
Teryl K. Frey
Keyword(s):  
Capsid Protein ◽  
Rubella Virus ◽  
Sindbis Virus ◽  
Packaging Signal ◽  
Initiation Of Replication ◽  
Replicase Proteins ◽  
Infected Cells ◽  
In Trans ◽  
Post Entry

Rubella virus (RUBV) contains a plus-strand RNA genome with two ORFs, one encoding the non-structural replicase proteins (NS-ORF) and the second encoding the virion structural proteins (SP-ORF). This study describes development and use of a trans-encapsidation system for the assembly of infectious RUBV-like replicon particles (VRPs) containing RUBV replicons (self replicating genomes with the SP-ORF replaced with a reporter gene). First, this system was used to map signals within the RUBV genome that mediate packaging of viral RNA. Mutations within a proposed packaging signal did not significantly affect relative packaging efficiency. The insertion of various fragments derived from the RUBV genome into Sindbis virus replicons revealed that there are several regions within the RUBV genome capable of enhancing encapsidation of heterologous replicon RNAs. Secondly, the trans-encapsidation system was used to analyse the effect of alterations within the capsid protein (CP) on release of VRPs and subsequent initiation of replication in newly infected cells. Deletion of the N-terminal eight amino acids of the CP reduced VRP titre significantly, which could be partially complemented by native CP provided in trans, indicating that this mutation affected an entry or post-entry event in the replication cycle. To test this hypothesis, the trans-encapsidation system was used to demonstrate the rescue of a lethal deletion within P150, one of the virus replicase proteins, by CP contained within the virus particle. This novel finding substantiated the functional role of CP in early post-entry replication.

scholarly journals RNA-Protein Interaction Analysis of SARS-CoV-2 5′ and 3′ Untranslated Regions Reveals a Role of Lysosome-Associated Membrane Protein-2a during Viral Infection

mSystems ◽  
2021 ◽  
Author(s):  
Rohit Verma ◽  
Sandhini Saha ◽  
Shiv Kumar ◽  
Shailendra Mani ◽  
Tushar Kanti Maiti ◽  
...  
Keyword(s):  
Interaction Analysis ◽  
Rna Virus ◽  
Untranslated Regions ◽  
Host Proteins ◽  
Positive Strand ◽  
Replication Complex ◽  
Positive Strand Rna Virus ◽  
Positive Strand Rna ◽  
Viral Genomic Rna

Replication of a positive-strand RNA virus involves an RNA-protein complex consisting of viral genomic RNA, host RNA(s), virus-encoded proteins, and host proteins. Dissecting out individual components of the replication complex will help decode the mechanism of viral replication. 5′ and 3′ UTRs in positive-strand RNA viruses play essential regulatory roles in virus replication.

scholarly journals Multiple capsid protein binding sites mediate selective packaging of the alphavirus genomic RNA

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Rebecca S. Brown ◽  
Dimitrios G. Anastasakis ◽  
Markus Hafner ◽  
Margaret Kielian
Keyword(s):  
Capsid Protein ◽  
Binding Sites ◽  
Semliki Forest Virus ◽  
Virus Assembly ◽  
Genomic Rna ◽  
Packaging Signal ◽  
Genome Packaging ◽  
Genome Recognition ◽  
Infected Cells

Abstract The alphavirus capsid protein (Cp) selectively packages genomic RNA (gRNA) into the viral nucleocapsid to produce infectious virus. Using photoactivatable ribonucleoside crosslinking and an innovative biotinylated Cp retrieval method, here we comprehensively define binding sites for Semliki Forest virus (SFV) Cp on the gRNA. While data in infected cells demonstrate Cp binding to the proposed genome packaging signal (PS), mutagenesis experiments show that PS is not required for production of infectious SFV or Chikungunya virus. Instead, we identify multiple Cp binding sites that are enriched on gRNA-specific regions and promote infectious SFV production and gRNA packaging. Comparisons of binding sites in cytoplasmic vs. viral nucleocapsids demonstrate that budding causes discrete changes in Cp-gRNA interactions. Notably, Cp’s top binding site is maintained throughout virus assembly, and specifically binds and assembles with Cp into core-like particles in vitro. Together our data suggest a model for selective alphavirus genome recognition and assembly.

scholarly journals Old World hantaviruses do not produce detectable amounts of dsRNA in infected cells and the 5′ termini of their genomic RNAs are monophosphorylated

2011 ◽  
Vol 92 (5) ◽  
pp. 1199-1204 ◽  
Author(s):  
Hao Wang ◽  
Antti Vaheri ◽  
Friedemann Weber ◽  
Alexander Plyusnin
Keyword(s):  
Rna Virus ◽  
Innate Immune ◽  
Hantaan Virus ◽  
Signalling Pathways ◽  
Genomic Rna ◽  
Genomic Rnas ◽  
Old World ◽  
Interferon Induction ◽  
Infected Cells ◽  
Negative Sense

dsRNA and 5′-triphosphate RNA are considered critical activators of the innate immune response because of their interaction with pattern recognition receptors. It has been reported that no dsRNA is detected in negative-sense RNA virus-infected cells and that Hantaan virus (HTNV) genomic RNA bears a 5′ monophosphate group. In this paper we examine the 5′ termini of genomic RNAs of and dsRNA production by two major groups of Old World hantaviruses. No detectable amounts of dsRNA were found in infected cells. Also, the genomic RNAs of these hantaviruses bear a 5′ monophosphate group and therefore are unable to trigger interferon induction. Taken together with the earlier data on HTNV, these results suggest that in addition to the dsRNA and genomic RNA, which may be only minimally involved in the induction of innate immunity, other cellular signalling pathways may also be involved and that these await further investigation.

scholarly journals Role of RNA in Facilitating Gag/Gag-Pol Interaction

2002 ◽  
Vol 76 (8) ◽  
pp. 4131-4137 ◽  
Author(s):  
Ahmad Khorchid ◽  
Rabih Halwani ◽  
Mark A. Wainberg ◽  
Lawrence Kleiman
Keyword(s):  
Human Immunodeficiency Virus ◽  
Genomic Rna ◽  
Immunodeficiency Virus ◽  
Membrane Bound ◽  
Bulk Membrane ◽  
Triton X ◽  
Viral Genomic Rna ◽  
Hiv 1

ABSTRACT We have examined the influence of RNA upon the interaction of Gag-Pol with Gag during human immunodeficiency virus type 1 (HIV-1) assembly. COS7 cells were transfected with protease-negative HIV-1 proviral DNA, and Gag/Gag-Pol complexes were detected by coimmunoprecipitation with anti-integrase. In COS7 cells, Gag/Gag-Pol is found almost entirely in pelletable, membrane-bound complexes. Exposure of cells to 1% Triton X-100 releases Gag/Gag-Pol from bulk membrane, but the complexes remain pelletable. The role of RNA in facilitating the interaction between Gag and Gag-Pol was examined in these bulk membrane-free, pelletable complexes. The specific presence of viral genomic RNA is not required to maintain the Gag/Gag-Pol interaction, but some type of RNA is, since exposure to RNase destabilized the Gag/Gag-Pol complex. When present only in Gag, the nucleocapsid mutation R7R10K11S, which inhibits Gag binding to RNA, inhibits the formation of both Gag and Gag/Gag-Pol complexes. When present only in Gag-Pol, this mutation has no effect upon complex formation. This result indicates that Gag-Pol may not interact directly with RNA but rather requires RNA-facilitated Gag multimerization for its interaction with Gag.

scholarly journals Location and Role of Free Cysteinyl Residues in the Sindbis Virus E1 and E2 Glycoproteins

2007 ◽  
Vol 81 (12) ◽  
pp. 6231-6240 ◽  
Author(s):  
Christopher B. Whitehurst ◽  
Erik J. Soderblom ◽  
Michelle L. West ◽  
Raquel Hernandez ◽  
Michael B. Goshe ◽  
...  
Keyword(s):  
Virus Particle ◽  
Disulfide Bonds ◽  
Sindbis Virus ◽  
Rna Virus ◽  
Virus Maturation ◽  
Virus Particles ◽  
Native Virus ◽  
Mature Virus Particle ◽  
Liquid Chromatography Tandem Mass

ABSTRACT Sindbis virus is a single-stranded positive-sense RNA virus. It is composed of 240 copies of three structural proteins: E1, E2, and capsid. These proteins form a mature virus particle composed of two nested T=4 icosahedral shells. A complex network of disulfide bonds in the E1 and E2 glycoproteins is developed through a series of structural intermediates as virus maturation occurs (M. Mulvey and D. T. Brown, J. Virol. 68:805-812, 1994; M. Carleton et al., J. Virol. 71:1558-1566, 1997). To better understand the nature of this disulfide network, E1 and E2 cysteinyl residues were labeled with iodoacetamide in the native virus particle and analyzed by liquid chromatography-tandem mass spectrometry. This analysis identified cysteinyl residues of E1 and E2, which were found to be label accessible in the native virus particle, as well as those that were either label inaccessible or blocked by their involvement in disulfide bonds. Native virus particles alkylated with iodoacetamide demonstrated a 4-log decrease in viral infectivity. This suggests that the modification of free cysteinyl residues results in the loss of infectivity by destabilizing the virus particle or that a rearrangement of disulfide bonds, which is required for infectivity, is blocked by the modification. Although modification of these residues prevented infectivity, it did not alter the ability of virus to fuse cells after exposure to acidic pH; thus, modification of free cysteinyl residues biochemically separated the process of infection from the process of membrane fusion.

scholarly journals HIV-1 Virus Interactions With Host Proteins: Interaction of the N-terminal Domain of the HIV-1 Capsid Protein With Human Calmodulin

2019 ◽  
Vol 14 (5) ◽  
pp. 1934578X1984919
Author(s):  
Ywh-Min Tzou ◽  
Ronald Shin ◽  
N. Rama Krishna
Keyword(s):  
Capsid Protein ◽  
Host Factors ◽  
Host Proteins ◽  
Terminal Domain ◽  
Immunodeficiency Virus ◽  
Infected Cells ◽  
Virus Interactions ◽  
Precise Role ◽  
Hiv 1

The human immunodeficiency virus (HIV-1 virus) exploits several host factors for assembly, infection, and replication within the infected cells. In this work, we describe the evidence for an interaction of the N-terminal domain of the HIV-1 capsid protein with human calmodulin. The precise role of this interaction within the life cycle of the HIV-1 virus is yet to be defined. Potential roles for this interaction in the viral capsid uncoating are discussed.

scholarly journals Noncapped Alphavirus Genomic RNAs and Their Role during Infection

2015 ◽  
Vol 89 (11) ◽  
pp. 6080-6092 ◽  
Author(s):  
K. J. Sokoloski ◽  
K. C. Haist ◽  
T. E. Morrison ◽  
S. Mukhopadhyay ◽  
R. W. Hardy
Keyword(s):  
Sindbis Virus ◽  
Rna Synthesis ◽  
Rna Virus ◽  
Genomic Rna ◽  
Concerted Effort ◽  
Genomic Rnas ◽  
Virus Particles ◽  
Molecular Determinant ◽  
Viral Particles ◽  
Positive Sense

ABSTRACTAlphaviruses are enveloped positive-sense RNA viruses that exhibit a wide host range consisting of vertebrate and invertebrate species. Previously we have reported that the infectivity of Sindbis virus (SINV), the model alphavirus, was largely a function of the cell line producing the viral particles. Mammalian-cell-derived SINV particles, on average, exhibit a higher particle-to-PFU ratio than mosquito cell-derived SINV particles. Nevertheless, the outcome of nonproductive infection, the molecular traits that determine particle infectivity and the biological importance of noninfectious particles were, prior to this study, unknown. Here, we report that the incoming genomic RNAs of noninfectious SINV particles undergo rapid degradation following infection. Moreover, these studies have led to the identification of the absence of the 5′ cap structure as a primary molecular determinant of particle infectivity. We show that the genomic RNAs of alphaviruses are not universally 5′ capped, with a significant number of noncapped genomic RNA produced early in infection. The production of noncapped viral genomic RNAs is important to the establishment and maintenance of alphaviral infection.IMPORTANCEThis report is of importance to the field of virology for three reasons. First, these studies demonstrate that noncapped Sindbis virus particles are produced as a result of viral RNA synthesis. Second, this report is, to our knowledge, the first instance of the direct measurement of the half-life of an incoming genomic RNA from a positive-sense RNA virus. Third, these studies indicate that alphaviral infection is likely a concerted effort of infectious and noninfectious viral particles.

scholarly journals Formation and Function of Liquid-Like Viral Factories in Negative-Sense Single-Stranded RNA Virus Infections

Viruses ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 126
Author(s):  
Justin M. Su ◽  
Maxwell Z. Wilson ◽  
Charles E. Samuel ◽  
Dzwokai Ma
Keyword(s):  
Rna Viruses ◽  
Rna Virus ◽  
Host Responses ◽  
Future Research ◽  
Virus Infections ◽  
Infected Cells ◽  
And Function ◽  
Negative Sense ◽  
Liquid Liquid Phase Separation

Liquid–liquid phase separation (LLPS) represents a major physiochemical principle to organize intracellular membrane-less structures. Studies with non-segmented negative-sense (NNS) RNA viruses have uncovered a key role of LLPS in the formation of viral inclusion bodies (IBs), sites of viral protein concentration in the cytoplasm of infected cells. These studies further reveal the structural and functional complexity of viral IB factories and provide a foundation for their future research. Herein, we review the literature leading to the discovery of LLPS-driven formation of IBs in NNS RNA virus-infected cells and the identification of viral scaffold components involved, and then outline important questions and challenges for IB assembly and disassembly. We discuss the functional implications of LLPS in the life cycle of NNS RNA viruses and host responses to infection. Finally, we speculate on the potential mechanisms underlying IB maturation, a phenomenon relevant to many human diseases.

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