bluuebunny/crossref_metadata_2025_split huggingface dataset:
Hepatitis C Virus Nonstructural 5A Protein Induces Interleukin-8, Leading to Partial Inhibition of the Interferon-Induced Antiviral Response
Authors: Polyak Stephen J. , Khabar Khalid S. A. , Paschal Denise M. , Ezelle Heather J. , Duverlie Gilles , Barber Glen N. , Levy David E. , Mukaida Naofumi , Gretch David R. 
Published: July 2002

ABSTRACTHepatitis C virus (HCV), a major cause of liver disease worldwide, is frequently resistant to the antiviral alpha interferon (IFN). The HCV nonstructural 5A (NS5A) protein has been implicated in HCV antiviral resistance in many studies. NS5A antagonizes the IFN antiviral response in vitro, and one mechanism is via inhibition of a key IFN-induced enzyme, the double-stranded-RNA-activated protein kinase (PKR). In the present study we determined if NS5A uses other strategies to subvert the IFN system. Expression of full-length NS5A proteins from patients who exhibited a complete response (FL-NS5A-CR) or were nonresponsive (FL-NS5A-NR) to IFN therapy in HeLa cells had no effect on IFN induction of IFN-stimulated gene factor 3 (ISGF-3). Expression of mutant NS5A proteins lacking 110 (NS5A-ΔN110), 222 (NS5A-ΔN222), and 334 amino-terminal amino acids and mutants lacking 117 and 230 carboxy-terminal amino acids also had no effect on ISGF-3 induction by IFN. Expression of FL-NS5A-CR and FL-NS5A-NR did not affect IFN-induced STAT-1 tyrosine phosphorylation or upregulation of PKR and major histocompatibility complex class I antigens. However, NS5A expression in human cells induced interleukin 8 (IL-8) mRNA and protein, and this effect correlated with inhibition of the antiviral effects of IFN in an in vitro bioassay. NS5A induced transcription of a reporter gene driven by the IL-8 promoter, and the first 133 bp of the IL-8 promoter made up the minimal domain required for NS5A transactivation. NS5A-ΔN110 and NS5A-ΔN222 stimulated the IL-8 promoter to higher levels than did the full-length NS5A protein, and this correlated with increased nuclear localization of the proteins. Additional mutagenesis of the IL-8 promoter suggested that NF-κB and AP-1 were important in NS5A-ΔN222 transactivation in the presence of tumor necrosis factor alpha and that NF–IL-6 was inhibitory to this process. This study suggests that NS5A inhibits the antiviral actions of IFN by at least two mechanisms and provides the first evidence for a biological effect of the transcriptional activity of the NS5A protein. During HCV infection, viral proteins may induce chemokines that contribute to HCV antiviral resistance and pathogenesis.
ICP0, ICP4, or VP16 Expressed from Adenovirus Vectors Induces Reactivation of Latent Herpes Simplex Virus Type 1 in Primary Cultures of Latently Infected Trigeminal Ganglion Cells
Authors: Halford William P. , Kemp Clinton D. , Isler Jennifer A. , Davido David J. , Schaffer Priscilla A. 
Published: July 2002

ABSTRACT
In a previous study, we demonstrated that infected-cell polypeptide 0 (ICP0) is necessary for the efficient reactivation of herpes simplex virus type 1 (HSV-1) in primary cultures of latently infected trigeminal ganglion (TG) cells (W. P. Halford and P. A. Schaffer, J. Virol. 75:3240–3249, 2001). The present study was undertaken to determine whether ICP0 is sufficient to trigger HSV-1 reactivation in latently infected TG cells. To test this hypothesis, replication-defective adenovirus vectors that express wild-type and mutant forms of ICP0 under the control of a tetracycline response element (TRE) promoter were constructed. Similar adenovirus vectors encoding wild-type ICP4, wild-type and mutant forms of the HSV-1 origin-binding protein (OBP), and wild-type and mutant forms of VP16 were also constructed. The TRE promoter was induced by coinfection of Vero cells with the test vector and an adenovirus vector that expresses the reverse tetracycline-regulated transactivator in the presence of doxycycline. Northern blot analysis demonstrated that transcription of the OBP gene in the adenovirus expression vector increased as a function of doxycycline concentration over a range of 0.1 to 10 μM. Likewise, Western blot analysis demonstrated that addition of 3 μM doxycycline to adenovirus vector-infected Vero cells resulted in a 100-fold increase in OBP expression. Wild-type forms of ICP0, ICP4, OBP, and VP16 expressed from adenovirus vectors were functional based on their ability to complement plaque formation in Vero cells by replication-defective HSV-1 strains with mutations in these genes. Adenovirus vectors that express wild-type forms of ICP0, ICP4, or VP16 induced reactivation of HSV-1 in 86% ± 5%, 86% ± 5%, and 97% ± 5% of TG cell cultures, respectively (means ± standard deviations). In contrast, vectors that express wild-type OBP or mutant forms of ICP0, OBP, or VP16 induced reactivation in 5% ± 5%, 8% ± 0%, 0% ± 0%, and 13% ± 6% of TG cell cultures, respectively. In control infections, an adenovirus vector expressed green fluorescent protein efficiently in TG neurons but did not induce HSV-1 reactivation. Therefore, expression of ICP0, ICP4, or VP16 is sufficient to induce HSV-1 reactivation in latently infected TG cell cultures. We conclude that this system provides a powerful tool for determining which cellular and viral proteins are sufficient to induce HSV-1 reactivation from neuronal latency.
Requirement for Uracil-DNA Glycosylase during the Transition to Late-Phase Cytomegalovirus DNA Replication
Authors: Courcelle Charmain Tan , Courcelle Justin , Prichard Mark N. , Mocarski Edward S. 
Published: July 2002

ABSTRACT
Cytomegalovirus gene UL114, a homolog of mammalian uracil-DNA glycosylase (UNG), is required for efficient viral DNA replication. In quiescent fibroblasts, UNG mutant virus replication is delayed for 48 h and follows the virus-induced expression of cellular UNG. In contrast, mutant virus replication proceeds without delay in actively growing fibroblasts that express host cell UNG. In the absence of viral or host cell UNG expression, mutant virus fails to proceed to late-phase DNA replication, characterized by rapid DNA amplification. The data suggest that uracil incorporated early during wild-type viral DNA replication must be removed by virus or host UNG prior to late-phase amplification and encapsidation into progeny virions. The process of uracil incorporation and excision may introduce strand breaks to facilitate the transition from early-phase replication to late-phase amplification.
Identification of Aleutian Mink Disease Parvovirus Capsid Sequences Mediating Antibody-Dependent Enhancement of Infection, Virus Neutralization, and Immune Complex Formation
Authors: Bloom Marshall E. , Best Sonja M. , Hayes Stanley F. , Wells Richard D. , Wolfinbarger James B. , McKenna Robert , Agbandje-McKenna Mavis 
Published: July 2002

ABSTRACTAleutian mink disease parvovirus (ADV) causes a persistent infection associated with circulating immune complexes, immune complex disease, hypergammaglobulinemia, and high levels of antiviral antibody. Although antibody can neutralize ADV infectivity in Crandell feline kidney cells in vitro, virus is not cleared in vivo, and capsid-based vaccines have proven uniformly ineffective. Antiviral antibody also enables ADV to infect macrophages, the target cells for persistent infection, by Fc-receptor-mediated antibody-dependent enhancement (ADE). The antibodies involved in these unique aspects of ADV pathogenesis may have specific targets on the ADV capsid. Prominent differences exist between the structure of ADV and other, more-typical parvoviruses, which can be accounted for by short peptide sequences in the flexible loop regions of the capsid proteins. In order to determine whether these short sequences are targets for antibodies involved in ADV pathogenesis, we studied heterologous antibodies against several peptides present in the major capsid protein, VP2. Of these antibodies, a polyclonal rabbit antibody to peptide VP2:428-446 was the most interesting. The anti-VP2:428-446 antibody aggregated virus particles into immune complexes, mediated ADE, and neutralized virus infectivity in vitro. Thus, antibody against this short peptide can be implicated in key facets of ADV pathogenesis. Structural modeling suggested that surface-exposed residues of VP2:428-446 are readily accessible for antibody binding. The observation that antibodies against a single target peptide in the ADV capsid can mediate both neutralization and ADE may explain the failure of capsid-based vaccines.
Infectious Simian/Human Immunodeficiency Virus with Human Immunodeficiency Virus Type 1 Subtype C from an African Isolate: Rhesus Macaque Model
Authors: Ndung'u Thumbi , Lu Yichen , Renjifo Boris , Touzjian Neal , Kushner Nicholas , Pena-Cruz Victor , Novitsky Vladimir A. , Lee Tun-Hou , Essex Max 
Published: July 2002

ABSTRACT

Human immunodeficiency virus type 1 (HIV-1) subtype C is responsible for more than 56% of all infections in the HIV and AIDS pandemic. It is the predominant subtype in the rapidly expanding epidemic in southern Africa. To develop a relevant model that would facilitate studies of transmission, pathogenesis, and vaccine development for this subtype, we generated SHIV
MJ4
, a simian/human immunodeficiency virus (SHIV) chimera based on HIV-1 subtype C. SHIV
MJ4
contains the majority of
env
, the entire second exon of
tat
, and a partial sequence of the second exon of
rev
, all derived from a CCR5-tropic, primary isolate envelope clone from southern Africa. SHIV
MJ4
replicated efficiently in human, rhesus, and pig-tailed macaque peripheral blood mononuclear cells (PBMCs) in vitro but not in CEMx174 cells. To assess in vivo infectivity, SHIV
MJ4
was intravenously inoculated into four rhesus macaques (
Macaca mulatta
). All four animals became infected as determined through virus isolation, PCR analysis, and viral loads of 10
7
to 10
8
copies of viral RNA per ml of plasma during the primary infection phase. We have established a CCR5-tropic SHIV
MJ4
/rhesus macaque model that may be useful in the studies of HIV-1 subtype C immunology and biology and may also facilitate the evaluation of vaccines to control the spread of HIV-1 subtype C in southern Africa and elsewhere.
Rat CRM1 Is Responsible for the Poor Activity of Human T-Cell Leukemia Virus Type 1 Rex Protein in Rat Cells
Authors: Hakata Yoshiyuki , Yamada Masami , Shida Hisatoshi 
Published: July 2002

ABSTRACT
Rat models of human T-cell leukemia virus type 1 (HTLV-1)-related diseases such as adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis have been reported. However, these models do not completely reproduce human diseases partly because HTLV-1 replicates poorly in rats. We investigated here the possible reason for this. We found that the activity of Rex in rat cells is quite low compared to that in human cells. As Rex function depends largely on the CRM1 protein, whose human type (human CRM1 [hCRM1]) directly binds to Rex and exports it from the nucleus to the cytoplasm, we assessed whether rat CRM1 (rCRM1) could act as well as hCRM1 as a cofactor for Rex activity. We first cloned a cDNA encoding rCRM1 and found that both rCRM1 and hCRM1 could bind to and export Rex protein to the cytoplasm with similar efficiencies. However, unlike hCRM1, rCRM1 could hardly support Rex function because of its poor ability in inducing the Rex-Rex interaction required for RNA export into the cytoplasm. These observations suggest that the poor ability of rCRM1 to act as a cofactor for Rex function may be responsible for the poor replication of HTLV-1 in rats.
Functional Specialization and Evolution of Leader Proteinases in the Family
Closteroviridae
Authors: Peng Chih-Wen , Peremyslov Valera V. , Mushegian Arcady R. , Dawson William O. , Dolja Valerian V. 
Published: July 2002

ABSTRACT

Members of the
Closteroviridae
and
Potyviridae
families of the plant positive-strand RNA viruses encode one or two papain-like leader proteinases. In addition to a C-terminal proteolytic domain, each of these proteinases possesses a nonproteolytic N-terminal domain. We compared functions of the several leader proteinases using a gene swapping approach. The leader proteinase (L-Pro) of
Beet yellows virus
(BYV; a closterovirus) was replaced with L1 or L2 proteinases of
Citrus tristeza virus
(CTV; another closterovirus), P-Pro proteinase of
Lettuce infectious yellows virus
(LIYV; a crinivirus), and HC-Pro proteinase of
Tobacco etch virus
(a potyvirus). Each foreign proteinase efficiently processed the chimeric BYV polyprotein in vitro. However, only L1 and P-Pro, not L2 and HC-Pro, were able to rescue the amplification of the chimeric BYV variants. The combined expression of L1 and L2 resulted in an increased RNA accumulation compared to that of the parental BYV. Remarkably, this L1-L2 chimera exhibited reduced invasiveness and inability to move from cell to cell. Similar analyses of the BYV hybrids, in which only the papain-like domain of L-Pro was replaced with those derived from L1, L2, P-Pro, and HC-Pro, also revealed functional specialization of these domains. In subcellular-localization experiments, distinct patterns were observed for the leader proteinases of BYV, CTV, and LIYV. Taken together, these results demonstrated that, in addition to a common proteolytic activity, the leader proteinases of closteroviruses possess specialized functions in virus RNA amplification, virus invasion, and cell-to-cell movement. The phylogenetic analysis suggested that functionally distinct L1 and L2 of CTV originated by a gene duplication event.
Hantavirus Nucleocapsid Protein Is Expressed as a Membrane-Associated Protein in the Perinuclear Region
Authors: Ravkov Eugene V. , Compans Richard W. 
Published: July 2002

ABSTRACT

Black Creek Canal virus (BCCV) is a New World hantavirus which is associated with hantavirus pulmonary syndrome. We have examined the site of expression of the BCCV nucleocapsid protein (NBCCV) in the absence of BCCV glycoproteins and found that the majority of the protein is localized to the Golgi region. Immunofluorescence analysis of BHK21 cells expressing the NBCCV and La Crosse virus nucleocapsid protein (NLACV) showed different intracellular localization patterns of these proteins within the same cell: NLACV is cytoplasmic, whereas NBCCV is perinuclear. NBCCV was found to be colocalized with α-mannosidase II, a marker for the Golgi complex. Also, NBCCV was found to be associated with microsomal membranes following cell fractionation. Sedimentation analysis in density gradients revealed that the membrane association of NBCCV is sensitive to treatments with high-salt and high-pH solutions, which indicates that NBCCV is a peripheral membrane protein. Analysis of NBCCV truncation mutants revealed that the 141-amino-acid C-terminal portion of this protein was capable of targeting green fluorescent protein to the perinuclear region. The difference in the intracellular localization between the NBCCV and NLACV proteins suggests that the mechanisms involved in the morphogenesis of New World hantaviruses are distinct from that documented for other members of the
Bunyaviridae
family.
Increased Expression of MIP-1α and MIP-1β mRNAs in the Brain Correlates Spatially and Temporally with the Spongiform Neurodegeneration Induced by a Murine Oncornavirus
Authors: Askovic Srdjan , Favara Cynthia , McAtee Frank J. , Portis John L. 
Published: July 2002

ABSTRACTThe chimeric murine oncornavirus FrCasEcauses a rapidly progressive paralytic disease associated with spongiform neurodegeneration throughout the neuroaxis. Neurovirulence is determined by the sequence of the viral envelope gene and by the capacity of the virus to infect microglia. The neurocytopathic effect of this virus appears to be indirect, since the cells which degenerate are not infected. In the present study we have examined the possible role of inflammatory responses in this disease and have used as a control the virus F43. F43 is an highly neuroinvasive but avirulent virus which differs from FrCasEonly in 3′polandenvsequences. Like FrCasE, F43 infects large numbers of microglial cells, but it does not induce spongiform neurodegeneration. RNAase protection assays were used to detect differential expression of genes encoding a variety of cytokines, chemokines, and inflammatory cell-specific markers. Tumor necrosis factor alpha (TNF-α) and TNF-β mRNAs were upregulated in advanced stages of disease but not early, even in regions with prominent spongiosis. Surprisingly there was no evidence for upregulation of the cytokines interleukin-1α (IL-1α), IL-1β, and IL-6 or of the microglial marker F4/80 at any stage of this disease. In contrast, increased levels of the β-chemokines MIP-1α and -β were seen early in the disease and were concentrated in regions of the brain rich in spongiosis, and the magnitude of responses was similar to that observed in the brains of mice injected with the glutamatergic neurotoxin ibotenic acid. MIP-1α and MIP-1β mRNAs were also upregulated in F43-inoculated mice, but the responses were three- to fivefold lower and occurred later in the course of infection than was observed in FrCasE-inoculated mice. These results suggest that the robust increase in expression of MIP-1α and MIP-1β in the brain represents a correlate of neurovirulence in this disease, whereas the TNF responses are likely secondary events.
Random, Asynchronous, and Asymmetric Transcriptional Activity of Enhancer-Flanking Major Immediate-Early Genes
ie1/3
and
ie2
during Murine Cytomegalovirus Latency in the Lungs
Authors: Grzimek Natascha K. A. , Dreis Doris , Schmalz Susanne , Reddehase Matthias J. 
Published: July 2002

ABSTRACT

The lungs are a major organ site of cytomegalovirus (CMV) pathogenesis, latency, and recurrence. Previous work on murine CMV latency has documented a high load and an even distribution of viral genomes in the lungs after the resolution of productive infection. Initiation of the productive cycle requires expression of the
ie1/3
transcription unit, which is driven by the immediate-early (IE) promoter P
1/3
and generates IE1 and IE3 transcripts by differential splicing. Latency is molecularly defined by the absence of IE3 transcripts specifying the essential transactivator protein IE3. In contrast, IE1 transcripts were found to be generated focally and randomly, reflecting sporadic P
1/3
activity. Selective generation of IE1 transcripts implies molecular control of latency operating after
ie1/3
transcription initiation. P
1/3
is regulated by an upstream enhancer. It is widely assumed that the viral transcriptional program is started by activation of the enhancer through the binding of transcription factors. Accordingly, stochastic transcription during latency might reflect episodes of enhancer activation by the “noise” activity of intrinsic transcription factors. In addition to
ie1/3
, the enhancer controls gene
ie2
, which has its own promoter, P
2
, and is transcribed in opposite direction. We show here that
ie2
is also randomly transcribed during latency. Notably, however,
ie1
and
ie2
were found to be expressed independently. We infer from this finding that expression of the major IE genes is regulated asymmetrically and asynchronously via the combined control unit P
1/3
-E-P
2
. Our data are consistent with a stochastic nature of enhancer action as it is proposed by the “binary” or probability model.
Mitochondrial Aconitase Binds to the 3′ Untranslated Region of the Mouse Hepatitis Virus Genome
Authors: Nanda Santosh K. , Leibowitz Julian L. 
Published: July 2002

ABSTRACTMouse hepatitis virus (MHV), a member of theCoronaviridae, contains a polyadenylated positive-sense single-stranded genomic RNA which is 31 kb long. MHV replication and transcription take place via the synthesis of negative-strand RNA intermediates from a positive-strand genomic template. Acis-acting element previously identified in the 3′ untranslated region binds totrans-acting host factors from mouse fibroblasts and forms at least three RNA-protein complexes. The largest RNA-protein complex formed by thecis-acting element and the lysate from uninfected mouse fibroblasts has a molecular weight of about 200 kDa. The complex observed in gel shift assays has been resolved by second-dimension sodium dodecyl sulfate-polyacrylamide gel electrophoresis into four proteins of approximately 90, 70, 58, and 40 kDa after RNase treatment. Specific RNA affinity chromatography also has revealed the presence of a 90-kDa protein associated with RNA containing thecis-acting element bound to magnetic beads. The 90-kDa protein has been purified from uninfected mouse fibroblast crude lysates. Protein microsequencing identified the 90-kDa protein as mitochondrial aconitase. Antibody raised against purified mitochondrial aconitase recognizes the RNA-protein complex and the 90-kDa protein, which can be released from the complex by RNase digestion. Furthermore, UV cross-linking studies indicate that highly purified mitochondrial aconitase binds specifically to the MHV 3′ protein-binding element. Increasing the intracellular level of mitochondrial aconitase by iron supplementation resulted in increased RNA-binding activity in cell extracts and increased virus production as well as viral protein synthesis at early hours of infection. These results are particularly interesting in terms of identification of an RNA target for mitochondrial aconitase, which has a cytoplasmic homolog, cytoplasmic aconitase, also known as iron regulatory protein 1, a well-recognized RNA-binding protein. The binding properties of mitochondrial aconitase and the functional relevance of RNA binding appear to parallel those of cytoplasmic aconitase.
Herpes Simplex Virus Type 1 Promoter Activity during Latency Establishment, Maintenance, and Reactivation in Primary Dorsal Root Neurons In Vitro
Authors: Arthur Jane L. , Scarpini Cinzia G. , Connor Vivienne , Lachmann Robin H. , Tolkovsky Aviva M. , Efstathiou Stacey 
Published: July 2002

ABSTRACT
A neonatal rat dorsal root ganglion-derived neuronal culture system has been utilized to study herpes simplex virus (HSV) latency establishment, maintenance, and reactivation. We present our initial characterization of viral gene expression in neurons following infection with replication-defective HSV recombinants carrying β-galactosidase and/or green fluorescent protein reporter genes under the control of lytic cycle- or latency-associated promoters. In this system lytic virus reporter promoter activity was detected in up to 58% of neurons 24 h after infection. Lytic cycle reporter promoters were shut down over time, and long-term survival of neurons harboring latent virus genomes was demonstrated. Latency-associated promoter-driven reporter gene expression was detected in neurons from early times postinfection and was stably maintained in up to 83% of neurons for at least 3 weeks. In latently infected cultures, silent lytic cycle promoters could be activated in up to 53% of neurons by nerve growth factor withdrawal or through inhibition of histone deacetylases by trichostatin A. We conclude that the use of recombinant viruses containing reporter genes, under the regulation of lytic and latency promoter control in neuronal cultures in which latency can be established and reactivation can be induced, is a potentially powerful system in which to study the molecular events that occur during HSV infection of neurons.