Novel Gene Product of Thogoto Virus Segment 6 Codes for an Interferon Antagonist

ABSTRACT Thogoto virus (THOV) is a tick-transmitted orthomyxovirus with a genome of six negative-stranded RNA segments. The sixth segment encodes two different transcripts: a spliced transcript that is translated into the matrix protein (M) and an unspliced transcript. Here, we report that the unspliced transcript encodes an elongated form of M named ML. A THOV isolate deficient in ML expression was an efficient interferon inducer, whereas ML-expressing wild-type strains were poor interferon inducers. These results were confirmed with recombinant THOVs rescued from cDNAs. Expression of ML efficiently suppressed activation of the beta interferon promoter by double-stranded RNA. These results indicate that ML is an accessory protein that functions as a potent interferon antagonist by blocking transcriptional activation of alpha/beta interferons.

Download Full-text

Bunyamwera Bunyavirus Nonstructural Protein NSs Counteracts the Induction of Alpha/Beta Interferon

Journal of Virology ◽

10.1128/jvi.76.16.7949-7955.2002 ◽

2002 ◽

Vol 76 (16) ◽

pp. 7949-7955 ◽

Cited By ~ 157

Author(s):

Friedemann Weber ◽

Anne Bridgen ◽

John K. Fazakerley ◽

Hein Streitenfeld ◽

Nina Kessler ◽

...

Keyword(s):

Transcriptional Activation ◽

Defense Mechanisms ◽

Reverse Genetics ◽

Cultured Cells ◽

Nonstructural Protein ◽

Wild Type ◽

Double Stranded Rna ◽

Wild Type Counterpart ◽

Alpha Beta ◽

Bunyamwera Virus

ABSTRACT Production of alpha/beta interferons (IFN-α/β) in response to viral infection is one of the main defense mechanisms of the innate immune system. Many viruses therefore encode factors that subvert the IFN system to enhance their virulence. Bunyamwera virus (BUN) is the prototype of the Bunyaviridae family. By using reverse genetics, we previously produced a recombinant virus lacking the nonstructural protein NSs (BUNdelNSs) and showed that NSs is a nonessential gene product that contributes to viral pathogenesis. Here we demonstrate that BUNdelNSs is a strong inducer of IFN-α/β, whereas in cells infected with the wild-type counterpart expressing NSs (wild-type BUN), neither IFN nor IFN mRNA could be detected. IFN induction by BUNdelNSs correlated with activation of NF-κB and was dependent on virally produced double-stranded RNA and on the IFN transcription factor IRF-3. Furthermore, both in cultured cells and in mice lacking a functional IFN-α/β system, BUNdelNSs replicated to wild-type BUN levels, whereas in IFN-competent systems, wild-type BUN grew more efficiently. These results suggest that BUN NSs is an IFN induction antagonist that blocks the transcriptional activation of IFN-α/β in order to increase the virulence of Bunyamwera virus.

Download Full-text

PPEY Motif within the Rabies Virus (RV) Matrix Protein Is Essential for Efficient Virion Release and RV Pathogenicity

Journal of Virology ◽

10.1128/jvi.00889-08 ◽

2008 ◽

Vol 82 (19) ◽

pp. 9730-9738 ◽

Cited By ~ 56

Author(s):

Christoph Wirblich ◽

Gene S. Tan ◽

Amy Papaneri ◽

Peter J. Godlewski ◽

Jan Marc Orenstein ◽

...

Keyword(s):

Viral Replication ◽

Rabies Virus ◽

Rna Synthesis ◽

Matrix Protein ◽

Virus Production ◽

Wild Type Virus ◽

Wild Type ◽

Conserved Sequence ◽

Virion Release ◽

The Matrix

ABSTRACT Late (L) domains containing the highly conserved sequence PPXY were first described for retroviruses, and later research confirmed their conservation and importance for efficient budding of several negative-stranded RNA viruses. Rabies virus (RV), a member of the Rhabdoviridae family, contains the sequence PPEY (amino acids 35 to 38) within the N terminus of the matrix (M) protein, but the functions of this potential L-domain in the viral life cycle, viral pathogenicity, and immunogenicity have not been established. Here we constructed a series of recombinant RVs containing mutations within the PPEY motif and analyzed their effects on viral replication and RV pathogenicity. Our results indicate that the first proline at position 35 is the most important for viral replication, whereas P36 and Y38 have a lesser but still noticeable impact. The reduction in viral replication was most likely due to inhibition of virion release, because initially no major impact on RV RNA synthesis was observed. In addition, results from electron microscopy demonstrated that the M4A mutant virus (PPEY→SAEA) displayed a more cell-associated phenotype than that of wild-type RV. Furthermore, all mutations within the PPEY motif resulted in reduced spread of the recombinant RVs as indicated by a reduction in focus size. Importantly, recombinant PPEY L-domain mutants were highly attenuated in mice yet still elicited potent antibody responses against RV G protein that were as high as those observed after infection with wild-type virus. Our data indicate that the RV PPEY motif has L-domain activity essential for efficient virus production and pathogenicity but is not essential for immunogenicity and thus can be targeted to increase the safety of rabies vaccine vectors.

Download Full-text

The unique degradation pathway of the PTS2 receptor, Pex7, is dependent on the PTS receptor/coreceptor, Pex5 and Pex20

Molecular Biology of the Cell ◽

10.1091/mbc.e13-12-0716 ◽

2014 ◽

Vol 25 (17) ◽

pp. 2634-2643 ◽

Cited By ~ 11

Author(s):

Danielle Hagstrom ◽

Changle Ma ◽

Soumi Guha-Polley ◽

Suresh Subramani

Keyword(s):

Matrix Protein ◽

Protein Import ◽

Degradation Pathway ◽

Matrix Proteins ◽

Receptor Recycling ◽

Wild Type ◽

The Matrix ◽

Peroxisomal Targeting ◽

Targeting Signals ◽

The Stability

Peroxisomal matrix protein import uses two peroxisomal targeting signals (PTSs). Most matrix proteins use the PTS1 pathway and its cargo receptor, Pex5. The PTS2 pathway is dependent on another receptor, Pex7, and its coreceptor, Pex20. We found that during the matrix protein import cycle, the stability and dynamics of Pex7 differ from those of Pex5 and Pex20. In Pichia pastoris, unlike Pex5 and Pex20, Pex7 is constitutively degraded in wild-type cells but is stabilized in pex mutants affecting matrix protein import. Degradation of Pex7 is more prevalent in cells grown in methanol, in which the PTS2 pathway is nonessential, in comparison with oleate, suggesting regulation of Pex7 turnover. Pex7 must shuttle into and out of peroxisomes before it is polyubiquitinated and degraded by the proteasome. The shuttling of Pex7, and consequently its degradation, is dependent on the receptor recycling pathways of Pex5 and Pex20 and relies on an interaction between Pex7 and Pex20. We also found that blocking the export of Pex20 from peroxisomes inhibits PTS1-mediated import, suggesting sharing of limited components in the export of PTS receptors/coreceptors. The shuttling and stability of Pex7 are divergent from those of Pex5 and Pex20, exemplifying a novel interdependence of the PTS1 and PTS2 pathways.

Download Full-text

TWO CHROMOSOMAL GENES REQUIRED FOR KILLING EXPRESSION IN KILLER STRAINS OF SACCHAROMYCES CEREVISIAE

Genetics ◽

10.1093/genetics/82.3.429 ◽

1976 ◽

Vol 82 (3) ◽

pp. 429-442

Author(s):

Reed B Wickner ◽

Michael J Leibowitz

Keyword(s):

Saccharomyces Cerevisiae ◽

Molecular Weight ◽

Mutant Phenotype ◽

Wild Type ◽

Double Stranded Rna ◽

Killer Strain ◽

Chromosomal Genes ◽

Complementation Groups ◽

Killer Plasmid ◽

Type Strains

ABSTRACT The killer character of yeast is determined by a 1.4 × 106 molecular weight double-stranded RNA plasmid and at least 12 chromosomal genes. Wild-type strains of yeast that carry this plasmid (killers) secrete a toxin which is lethal only to strains not carrying this plasmid (sensitives). —— We have isolated 28 independent recessive chromosomal mutants of a killer strain that have lost the ability to secrete an active toxin but remain resistant to the effects of the toxin and continue to carry the complete cytoplasmic killer genome. These mutants define two complementation groups, kex1 and kex2. Kex1 is located on chromosome VII between ade5 and lys5. Kex2 is located on chromosome XIV, but it does not show meiotic linkage to any gene previously located on this chromosome. —— When the killer plasmid of kex1 or kex2 strains is eliminated by curing with heat or cycloheximide, the strains become sensitive to killing. The mutant phenotype reappears among the meiotic segregants in a cross with a normal killer. Thus, the kex phenotype does not require an alteration of the killer plasmid. —— Kex1 and kex2 strains each contain near-normal levels of the 1.4 × 106 molecular weight double-stranded RNA, whose presence is correlated with the presence of the killer genome.

Download Full-text

Transcriptional activation of virulence genes in wild-type strains of Listeria monocytogenes in response to a change in the extracellular medium composition

Research in Microbiology ◽

10.1016/0923-2508(96)84712-7 ◽

1996 ◽

Vol 147 (5) ◽

pp. 371-384 ◽

Cited By ~ 112

Author(s):

M.-T. Ripio ◽

G. Domínguez-Bernal ◽

M. Suárez ◽

K. Brehm ◽

P. Berche ◽

...

Keyword(s):

Listeria Monocytogenes ◽

Transcriptional Activation ◽

Virulence Genes ◽

Medium Composition ◽

Wild Type ◽

Extracellular Medium ◽

Type Strains

Download Full-text

Evasion of Host Defenses by Measles Virus: Wild-Type Measles Virus Infection Interferes with Induction of Alpha/Beta Interferon Production

Journal of Virology ◽

10.1128/jvi.74.16.7478-7484.2000 ◽

2000 ◽

Vol 74 (16) ◽

pp. 7478-7484 ◽

Cited By ~ 123

Author(s):

Denise Naniche ◽

Annie Yeh ◽

Danelle Eto ◽

Marianne Manchester ◽

Robert M. Friedman ◽

...

Keyword(s):

Peripheral Blood ◽

Measles Virus ◽

Mononuclear Cells ◽

Human Peripheral Blood ◽

Antiviral Immunity ◽

Peripheral Blood Lymphocytes ◽

Wild Type ◽

Peripheral Blood Mononuclear ◽

Double Stranded Rna ◽

Alpha Beta

ABSTRACT Measles is a highly contagious disease currently responsible for over one million childhood deaths, particularly in the developing world. Since alpha/beta interferons (IFNs) are pivotal players both in nonspecific antiviral immunity and in specific cellular responses, their induction or suppression by measles virus (MV) could influence the outcome of a viral infection. In this study we compare the IFN induction and sensitivity of laboratory-passaged attenuated MV strains Edmonston and Moraten with those of recent wild-type viruses isolated and passaged solely on human peripheral blood mononuclear cells (PBMC) or on the B958 marmoset B-cell line. We report that two PBMC-grown wild-type measles isolates and two B958-grown strains of MV induce 10- to 80-fold-lower production of IFN by phytohemagglutinin-stimulated peripheral blood lymphocytes (PBL) compared to Edmonston and Moraten strains of measles. Preinfection of PBL with these non-IFN-inducing MV isolates prevents Edmonston-induced but not double-stranded-RNA-induced IFN production. This suggests that the wild-type viruses can actively inhibit Edmonston-induced IFN synthesis and that this is not occurring by double-stranded RNA. Furthermore, the wild-type MV is more sensitive than Edmonston MV to the effect of IFN. MV is thus able to suppress the synthesis of the earliest mediator of antiviral immunity, IFN-α/β. This could have important implications in the virulence and spread of MV.

Download Full-text

Activation of the ADE Genes Requires the Chromatin Remodeling Complexes SAGA and SWI/SNF

Eukaryotic Cell ◽

10.1128/ec.00068-07 ◽

2007 ◽

Vol 6 (8) ◽

pp. 1474-1485 ◽

Cited By ~ 6

Author(s):

Rebecca N. Koehler ◽

Nicole Rachfall ◽

Ronda J. Rolfes

Keyword(s):

Chromatin Remodeling ◽

Growth Conditions ◽

Specific Response ◽

Wild Type ◽

Reporter Expression ◽

Activator Proteins ◽

Genome Wide ◽

A Genome ◽

Chromatin Remodeling Complexes ◽

Type Strains

ABSTRACT The activation of the ADE regulon genes requires the pair of transcription factors Bas1 and Pho2. In a genome-wide screen for additional regulators of the pathway, strains with mutations in multiple subunits of the chromatin remodeling complexes SAGA and SWI/SNF were uncovered. These mutants exhibited decreased expression of an ADE5,7-lacZ reporter and native ADE compared to the wild-type strains, but the expression of the BAS1 and PHO2 genes was not substantially decreased. An unregulated Bas1-Pho2 fusion protein depended upon SAGA and SWI/SNF activity to promote transcription of a reporter. A significant but low-level association of Gcn5-myc and Snf2-myc with the ADE5,7 promoter was independent of adenine growth conditions and independent of the presence of the activator proteins Bas1 and Pho2. However, the increase in occupancy of Bas1 and Pho2 at ADE5,7 depended on both SAGA and SWI/SNF. The loss of catalytic activity of both SAGA and SWI/SNF complexes in the gcn5Δ snf2Δ double mutant was severely detrimental to ADE-lacZ reporter expression and native ADE gene expression, indicating complementary roles for these complexes. We conclude that Bas1 and Pho2 do not recruit the SAGA and SWI/SNF complexes to the ADE5,7 promoter but that the remodeling complexes are necessary to increase the binding of Bas1 and Pho2 in response to the adenine regulatory signal. Our data support the model that the SAGA and SWI/SNF complexes engage in global surveillance that is necessary for the specific response by Bas1 and Pho2.

Download Full-text

Alteration of a Novel Dispensable Mitochondrial Ribosomal Small-Subunit Protein, Rsm28p, Allows Translation of Defective COX2 mRNAs

Eukaryotic Cell ◽

10.1128/ec.4.2.337-345.2005 ◽

2005 ◽

Vol 4 (2) ◽

pp. 337-345 ◽

Cited By ~ 13

Author(s):

Elizabeth H. Williams ◽

Nada Bsat ◽

Nathalie Bonnefoy ◽

Christine A. Butler ◽

Thomas D. Fox

Keyword(s):

Mitochondrial Gene ◽

Carbon Sources ◽

Small Subunit ◽

Wild Type ◽

Inner Mitochondrial Membrane ◽

Positive Role ◽

Mitochondrial Ribosomes ◽

The Matrix ◽

Triton X ◽

Type Strains

ABSTRACT Mutations affecting the RNA sequence of the first 10 codons of the Saccharomyces cerevisiae mitochondrial gene COX2 strongly reduce translation of the mRNA, which encodes the precursor of cytochrome c oxidase subunit II. A dominant chromosomal mutation that suppresses these defects is an internal in-frame deletion of 67 codons from the gene YDR494w. Wild-type YDR494w encodes a 361-residue polypeptide with no similarity to proteins of known function. The epitope-tagged product of this gene, now named RSM28, is both peripherally associated with the inner surface of the inner mitochondrial membrane and soluble in the matrix. Epitope-tagged Rsm28p from Triton X-100-solubilized mitochondria sedimented with the small subunit of mitochondrial ribosomes in a sucrose gradient containing 500 mM NH4Cl. Complete deletion of RSM28 caused only a modest decrease in growth on nonfermentable carbon sources in otherwise wild-type strains and enhanced the respiratory defect of the suppressible cox2 mutations. The rsm28 null mutation also reduced translation of an ARG8 m reporter sequence inserted at the COX1, COX2, and COX3 mitochondrial loci. We tested the ability of RSM28-1 to suppress a variety of cox2 and cox3 mutations and found that initiation codon mutations in both genes were suppressed. We conclude that Rsm28p is a dispensable small-subunit mitochondrial ribosomal protein previously undetected in systematic investigations of these ribosomes, with a positive role in translation of several mitochondrial mRNAs.

Download Full-text

Ebola Virus VP35 Protein Binds Double-Stranded RNA and Inhibits Alpha/Beta Interferon Production Induced by RIG-I Signaling

Journal of Virology ◽

10.1128/jvi.02199-05 ◽

2006 ◽

Vol 80 (11) ◽

pp. 5168-5178 ◽

Cited By ~ 293

Author(s):

Washington B. Cárdenas ◽

Yueh-Ming Loo ◽

Michael Gale ◽

Amy L. Hartman ◽

Christopher R. Kimberlin ◽

...

Keyword(s):

Ebola Virus ◽

Binding Activity ◽

Wild Type ◽

Antagonist Activity ◽

Double Stranded Rna ◽

Beta Interferon ◽

Double Stranded Dna ◽

Dsrna Binding ◽

Alpha Beta

ABSTRACT The Ebola virus (EBOV) VP35 protein blocks the virus-induced phosphorylation and activation of interferon regulatory factor 3 (IRF-3), a transcription factor critical for the induction of alpha/beta interferon (IFN-α/β) expression. However, the mechanism(s) by which this blockage occurs remains incompletely defined. We now provide evidence that VP35 possesses double-stranded RNA (dsRNA)-binding activity. Specifically, VP35 bound to poly(rI) · poly(rC)-coated Sepharose beads but not control beads. In contrast, two VP35 point mutants, R312A and K309A, were found to be greatly impaired in their dsRNA-binding activity. Competition assays showed that VP35 interacted specifically with poly(rI) · poly(rC), poly(rA) · poly(rU), or in vitro-transcribed dsRNAs derived from EBOV sequences, and not with single-stranded RNAs (ssRNAs) or double-stranded DNA. We then screened wild-type and mutant VP35s for their ability to target different components of the signaling pathways that activate IRF-3. These experiments indicate that VP35 blocks activation of IRF-3 induced by overexpression of RIG-I, a cellular helicase recently implicated in the activation of IRF-3 by either virus or dsRNA. Interestingly, the VP35 mutants impaired for dsRNA binding have a decreased but measurable IFN antagonist activity in these assays. Additionally, wild-type and dsRNA-binding-mutant VP35s were found to have equivalent abilities to inhibit activation of the IFN-β promoter induced by overexpression of IPS-1, a recently identified signaling molecule downstream of RIG-I, or by overexpression of the IRF-3 kinases IKKε and TBK-1. These data support the hypothesis that dsRNA binding may contribute to VP35 IFN antagonist function. However, additional mechanisms of inhibition, at a point proximal to the IRF-3 kinases, most likely also exist.

Download Full-text