Faculty Opinions recommendation of Viral translation is coupled to transcription in Sindbis virus-infected cells.

2007 ◽  
Author(s):  
Lee Gehrke
Keyword(s):  
Sindbis Virus ◽  
Viral Translation ◽  
Infected Cells

scholarly journals Viral Translation Is Coupled to Transcription in Sindbis Virus-Infected Cells

2007 ◽  
Vol 81 (13) ◽  
pp. 7061-7068 ◽  
Author(s):  
Miguel A. Sanz ◽  
Alfredo Castelló ◽  
Luis Carrasco
Keyword(s):  
Sindbis Virus ◽  
Late Phase ◽  
Mrna Translation ◽  
Encephalomyocarditis Virus ◽  
Host Protein ◽  
Viral Mrnas ◽  
Viral Translation ◽  
Bhk Cells ◽  
Infected Cells

ABSTRACT During the late phase of Sindbis virus infection, the viral subgenomic mRNA is translated efficiently in BHK cells, whereas host protein synthesis is inhibited. However, transfection of in vitro-generated Sindbis virus subgenomic mRNA leads to efficient translation in uninfected BHK cells, whereas it is a poor substrate in infected cells. Therefore, the structure of the subgenomic mRNA itself is not sufficient to confer its translatability in infected cells. In this regard, translation of the subgenomic mRNA requires synthesis from the viral transcription machinery. The lack of translation of transfected viral mRNAs in infected cells is not due to their degradation nor is it a consequence of competition between viral transcripts and transfected mRNAs, because a replicon that cannot produce subgenomic mRNA also interferes with exogenous mRNA translation. Interestingly, subgenomic mRNA is translated more efficiently when it is transfected into uninfected cells than when it is transcribed from a transfected replicon. Finally, a similar behavior was observed for other RNA viruses, such as vesicular stomatitis virus and encephalomyocarditis virus. These findings support the notion that translation is coupled to transcription in cells infected with different animal viruses.

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.

Effect of Interferon on RNA Synthesis in Sindbis Virus Infected Cells

1966 ◽  
Vol 121 (2) ◽  
pp. 630-632 ◽  
Author(s):  
H. B. Levy ◽  
L. F. Snellbaker ◽  
S. Baron
Keyword(s):  
Sindbis Virus ◽  
Rna Synthesis ◽  
Infected Cells

scholarly journals Effects of inducing or inhibiting apoptosis on Sindbis virus replication in mosquito cells

2008 ◽  
Vol 89 (11) ◽  
pp. 2651-2661 ◽  
Author(s):  
Hua Wang ◽  
Carol D. Blair ◽  
Ken E. Olson ◽  
Rollie J. Clem
Keyword(s):  
Virus Replication ◽  
Persistent Infection ◽  
Sindbis Virus ◽  
Actinomycin D ◽  
Virus Production ◽  
Cell Types ◽  
Lytic Infection ◽  
Mosquito Cells ◽  
Infected Cells ◽  
Apoptotic Genes

Sindbis virus (SINV) is a mosquito-borne virus in the genus Alphavirus, family Togaviridae. Like most alphaviruses, SINVs exhibit lytic infection (apoptosis) in many mammalian cell types, but are generally thought to cause persistent infection with only moderate cytopathic effects in mosquito cells. However, there have been several reports of apoptotic-like cell death in mosquitoes infected with alphaviruses or flaviviruses. Given that apoptosis has been shown to be an antiviral response in other systems, we have constructed recombinant SINVs that express either pro-apoptotic or anti-apoptotic genes in order to test the effects of inducing or inhibiting apoptosis on SINV replication in mosquito cells. Recombinant SINVs expressing the pro-apoptotic genes reaper (rpr) from Drosophila or michelob_x (mx) from Aedes aegypti caused extensive apoptosis in cells from the mosquito cell line C6/36, thus changing the normal persistent infection observed with SINV to a lytic infection. Although the infected cells underwent apoptosis, high levels of virus replication were still observed during the initial infection. However, virus production subsequently decreased compared with persistently infected cells, which continued to produce high levels of virus over the next several days. Infection of C6/36 cells with SINV expressing the baculovirus caspase inhibitor P35 inhibited actinomycin D-induced caspase activity and protected infected cells from actinomycin D-induced apoptosis, but had no observable effect on virus replication. This study is the first to test directly whether inducing or inhibiting apoptosis affects arbovirus replication in mosquito cells.

scholarly journals Disruption of Stress Granule Formation by the Multifunctional Cricket Paralysis Virus 1A Protein

2016 ◽  
Vol 91 (5) ◽  
Author(s):  
Anthony Khong ◽  
Craig H. Kerr ◽  
Clarence H. L. Yeung ◽  
Kathleen Keatings ◽  
Arabinda Nayak ◽  
...  
Keyword(s):  
Virus Infection ◽  
Stress Granule ◽  
Granule Formation ◽  
Viral Translation ◽  
Content Type ◽  
Cellular Processes ◽  
Cellular Environment ◽  
Viral Yield ◽  
Infected Cells ◽  
Paralysis Virus

ABSTRACT Stress granules (SGs) are cytosolic ribonucleoprotein aggregates that are induced during cellular stress. Several viruses modulate SG formation, suggesting that SGs have an impact on virus infection. However, the mechanisms and impact of modulating SG assembly in infected cells are not completely understood. In this study, we identify the dicistrovirus cricket paralysis virus 1A (CrPV-1A) protein that functions to inhibit SG assembly during infection. Moreover, besides inhibiting RNA interference, CrPV-1A also inhibits host transcription, which indirectly modulates SG assembly. Thus, CrPV-1A is a multifunctional protein. We identify a key R146A residue that is responsible for these effects, and mutant CrPV(R146A) virus infection is attenuated in Drosophila melanogaster S2 cells and adult fruit flies and results in increased SG formation. Treatment of CrPV(R146A)-infected cells with actinomycin D, which represses transcription, restores SG assembly suppression and viral yield. In summary, CrPV-1A modulates several cellular processes to generate a cellular environment that promotes viral translation and replication. IMPORTANCE RNA viruses encode a limited set of viral proteins to modulate an array of cellular processes in order to facilitate viral replication and inhibit antiviral defenses. In this study, we identified a viral protein, called CrPV-1A, within the dicistrovirus cricket paralysis virus that can inhibit host transcription, modulate viral translation, and block a cellular process called stress granule assembly. We also identified a specific amino acid within CrPV-1A that is important for these cellular processes and that mutant viruses containing mutations of CrPV-1A attenuate virus infection. We also demonstrate that the CrPV-1A protein can also modulate cellular processes in human cells, suggesting that the mode of action of CrPV-1A is conserved. We propose that CrPV-1A is a multifunctional, versatile protein that creates a cellular environment in virus-infected cells that permits productive virus infection.

Confocal Analysis of the Distribution and Persistence of Sindbis Virus (TaV-GFP) Infection in Midguts of Aedes aegypti Mosquitoes

2020 ◽  
Vol 26 (2) ◽  
pp. 267-274
Author(s):  
Jason J. Saredy ◽  
Florence Y. Chim ◽  
Zoë L. Lyski ◽  
Yani P. Ahearn ◽  
Doria F. Bowers
Keyword(s):  
Aedes Aegypti ◽  
Sindbis Virus ◽  
Fluorescent Protein ◽  
Blood Feeding ◽  
Target Tissue ◽  
Secondary Target ◽  
Posterior Midgut ◽  
Infected Cells ◽  
Green Fluorescent ◽  
Global Threat

AbstractBiological transmission of arthropod-borne viruses (arboviruses) to vertebrate hosts by hematophagous insects poses a global threat because such arboviruses can result in a range of serious public health infectious diseases. Sindbis virus (SINV), the prototype Alphavirus, was used to track infections in the posterior midgut (PMG) of Aedes aegypti adult mosquitoes. Females were fed viremic blood containing a virus reporter, SINV [Thosea asigna virus-green fluorescent protein (TaV-GFP)], that leaves a fluorescent signal in infected cells. We assessed whole-mount PMGs to identify primary foci, secondary target tissues, distribution, and virus persistence. Following a viremic blood meal, PMGs were dissected and analyzed at various days of post blood-feeding. We report that virus foci indicated by GFP in midgut epithelial cells resulted in a 9.8% PMG infection and a 10.8% dissemination from these infected guts. The number of virus foci ranged from 1 to 3 per individual PMG and was more prevalent in the PMG-middle > PMG-frontal > PMG-caudal regions. SINV TaV-GFP was first observed in the PMG (primary target tissue) at 3 days post blood-feeding, was sequestered in circumscribed foci, replicated in PMG peristaltic muscles (secondary target tissue) following dissemination, and GFP was observed to persist in PMGs for 30 days postinfection.

scholarly journals Effects of Palmitoylation of Replicase Protein nsP1 on Alphavirus Infection

2000 ◽  
Vol 74 (15) ◽  
pp. 6725-6733 ◽  
Author(s):  
Tero Ahola ◽  
Pekka Kujala ◽  
Minna Tuittila ◽  
Titta Blom ◽  
Pirjo Laakkonen ◽  
...  
Keyword(s):  
Sindbis Virus ◽  
Semliki Forest Virus ◽  
Rna Replication ◽  
Cell Types ◽  
Alkaline Solutions ◽  
Nonstructural Proteins ◽  
Replication Complex ◽  
Replication Sites ◽  
Infected Cells ◽  
Alphavirus Infection

ABSTRACT The membrane-associated alphavirus RNA replication complex contains four virus-encoded subunits, the nonstructural proteins nsP1 to nsP4. Semliki Forest virus (SFV) nsP1 is hydrophobically modified by palmitoylation of cysteines 418 to 420. Here we show that Sindbis virus nsP1 is also palmitoylated on the same site (cysteine 420). When mutations preventing nsP1 palmitoylation were introduced into the genomes of these two alphaviruses, the mutant viruses remained viable and replicated to high titers, although their growth was slightly delayed. The subcellular distribution of palmitoylation-defective nsP1 was altered in the mutant: it no longer localized to filopodial extensions, and a fraction of it was soluble. The ultrastructure of the alphavirus replication sites appeared normal, and the localization of the other nonstructural proteins was unaltered in the mutants. In both wild-type- and mutant-virus-infected cells, SFV nsP3 and nsP4 could be extracted from membranes only by alkaline solutions whereas the nsP2-membrane association was looser. Thus, the membrane binding properties of the alphavirus RNA replication complex were not determined by the palmitoylation of nsP1. The nsP1 palmitoylation-defective alphaviruses produced normal plaques in several cell types, but failed to give rise to plaques in HeLa cells, although they induced normal apoptosis of these cells. The SFV mutant was apathogenic in mice: it caused blood viremia, but no infectious virus was detected in the brain.

scholarly journals Development of Sindbis Viruses Encoding nsP2/GFP Chimeric Proteins and Their Application for Studying nsP2 Functioning

2007 ◽  
Vol 81 (10) ◽  
pp. 5046-5057 ◽  
Author(s):  
Svetlana Atasheva ◽  
Rodion Gorchakov ◽  
Robert English ◽  
Ilya Frolov ◽  
Elena Frolova
Keyword(s):  
Virus Replication ◽  
Sindbis Virus ◽  
Fluorescent Protein ◽  
Protein Complexes ◽  
Chimeric Protein ◽  
Specific Protein ◽  
Heterologous Proteins ◽  
Peptide Tags ◽  
Infected Cells ◽  
Green Fluorescent

ABSTRACT Sindbis virus (SINV) is one of almost 30 currently known alphaviruses. In infected cells, it produces only a few proteins that function in virus replication and interfere with the development of the antiviral response. One of the viral nonstructural proteins, nsP2, not only exhibits protease and RNA helicase activities that are directly involved in viral RNA replication but also plays critical roles in the development of transcriptional and translational shutoffs in the SINV-infected cells. These multiple activities of nsP2 complicate investigations of this protein's functions and further understanding of its structure. Using a transposon-based approach, we generated a cDNA library of SINV genomes with a green fluorescent protein (GFP) gene randomly inserted into nsP2 and identified a number of sites that can be used for GFP cloning without a strong effect on virus replication. Recombinant SIN viruses encoding nsP2/GFP chimeric protein were capable of growth in tissue culture and interfering with cellular functions. SINV, expressing GFP in the nsP2, was used to isolate nsP2-specific protein complexes formed in the cytoplasm of the infected cells. These complexes contained viral nsPs, all of the cellular proteins that we previously coisolated with SINV nsP3, and some additional protein factors that were not found before in detectable concentrations. The random insertion library-based approach, followed by the selection of the viable variants expressing heterologous proteins, can be applied for mapping the domain structure of the viral nonstructural and structural proteins, cloning of peptide tags for isolation of the protein-specific complexes, and studying their formation by using live-cell imaging. This approach may also be applicable to presentation of additional antigens and retargeting of viruses to new receptors.

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