bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Herpes Simplex Virus Type 2 R1 Protein Kinase (ICP10 PK) Functions as a Dominant Regulator of Apoptosis in Hippocampal Neurons Involving Activation of the ERK Survival Pathway and Upregulation of the Antiapoptotic Protein Bag-1
Authors: Perkins D. , Pereira E. F. R. , Aurelian L. 
Published: December 2002

ABSTRACTHerpes simplex virus types 1 and 2 (HSV-1 and HSV-2) can trigger or block apoptosis in a cell type-dependent manner. We have recently shown that the protein kinase activity of the large subunit of the HSV-2 ribonucleotide reductase (R1) protein (ICP10 PK) blocks apoptosis in cultured hippocampal neurons by activating the extracellular signal-regulated kinase (ERK) survival pathway (Perkins et al., J. Virol. 76:1435-1449, 2002). The present studies were designed to better elucidate the mechanism of ICP10 PK-induced neuroprotection and determine whether HSV-1 has similar activity. The data indicate that apoptosis inhibition by ICP10 PK involves a c-Raf-1-dependent mechanism and induction of the antiapoptotic protein Bag-1 by the activated ERK survival pathway. Also associated with neuroprotection by ICP10 PK are increased activation/stability of the transcription factor CREB and stabilization of the antiapoptotic protein Bcl-2. HSV-1 and the ICP10 PK-deleted HSV-2 mutant ICP10ΔPK activate JNK, c-Jun, and ATF-2, induce the proapoptotic protein BAD, and trigger apoptosis in hippocampal neurons. c-Jun activation and apoptosis are inhibited in hippocampal cultures infected with HSV-1 in the presence of the JNK inhibitor SP600125, suggesting that JNK/c-Jun activation is required for HSV-1-induced apoptosis. Ectopically delivered ICP10 PK (but not its PK-negative mutant p139) inhibits apoptosis triggered by HSV-1 or ICP10ΔPK. Collectively, the data indicate that ICP10 PK-induced activation of the ERK survival pathway results in Bag-1 upregulation and overrides the proapoptotic JNK/c-Jun signal induced by other viral proteins.
Viability of a Drug-Resistant Human Immunodeficiency Virus Type 1 Protease Variant: Structural Insights for Better Antiviral Therapy
Authors: Prabu-Jeyabalan Moses , Nalivaika Ellen A. , King Nancy M. , Schiffer Celia A. 
Published: December 2002

ABSTRACT
Under the selective pressure of protease inhibitor therapy, patients infected with human immunodeficiency virus (HIV) often develop drug-resistant HIV strains. One of the first drug-resistant mutations to arise in the protease, particularly in patients receiving indinavir or ritonavir treatment, is V82A, which compromises the binding of these and other inhibitors but allows the virus to remain viable. To probe this drug resistance, we solved the crystal structures of three natural substrates and two commercial drugs in complex with an inactive drug-resistant mutant (D25N/V82A) HIV-1 protease. Through structural analysis and comparison of the protein-ligand interactions, we found that Val82 interacts more closely with the drugs than with the natural substrate peptides. The V82A mutation compromises these interactions with the drugs while not greatly affecting the substrate interactions, which is consistent with previously published kinetic data. Coupled with our earlier observations, these findings suggest that future inhibitor design may reduce the probability of the appearance of drug-resistant mutations by targeting residues that are essential for substrate recognition.
TRAF1 Is a Critical Regulator of JNK Signaling by the TRAF-Binding Domain of the Epstein-Barr Virus-Encoded Latent Infection Membrane Protein 1 but Not CD40
Authors: Eliopoulos Aristides G. , Waites Elyse R. , Blake Sarah M. S. , Davies Clare , Murray Paul , Young Lawrence S. 
Published: December 2002

ABSTRACTThe oncogenic Epstein-Barr virus (EBV)-encoded latent infection membrane protein 1 (LMP1) mimics a constitutive active tumor necrosis factor (TNF) family receptor in its ability to recruit TNF receptor-associated factors (TRAFs) and TNF receptor-associated death domain protein (TRADD) in a ligand-independent manner. As a result, LMP1 constitutively engages signaling pathways, such as the JNK and p38 mitogen-activated protein kinases (MAPK), the transcription factor NF-κB, and the JAK/STAT cascade, and these activities may explain many of its pleiotropic effects on cell phenotype, growth, and transformation. In this study we demonstrate the ability of the TRAF-binding domain of LMP1 to signal on the JNK/AP-1 axis in a cell type- dependent manner that critically involves TRAF1 and TRAF2. Thus, expression of this LMP1 domain in TRAF1-positive lymphoma cells promotes significant JNK activation, which is blocked by dominant-negative TRAF2 but not TRAF5. However, TRAF1 is absent in many established epithelial cell lines and primary nasopharyngeal carcinoma (NPC) biopsy specimens. In these cells, JNK activation by the TRAF-binding domain of LMP1 depends on the reconstitution of TRAF1 expression. The critical role of TRAF1 in the regulation of TRAF2-dependent JNK signaling is particular to the TRAF-binding domain of LMP1, since a homologous region in the cytoplasmic tail of CD40 or the TRADD-interacting domain of LMP1 signal on the JNK axis independently of TRAF1 status. These data further dissect the signaling components used by LMP1 and identify a novel role for TRAF1 as a modulator of oncogenic signals.
Mouse Cytomegalovirus Immediate-Early Protein 1 Binds with Host Cell Repressors To Relieve Suppressive Effects on Viral Transcription and Replication during Lytic Infection
Authors: Tang Qiyi , Maul Gerd G. 
Published: December 2002

ABSTRACT
Herpesviruses start their transcriptional cascade at nuclear domain 10 (ND10). The deposition of virus genomes at these nuclear sites occurs due to the binding of the interferon-inducible repressor protein promyelocytic leukemia protein (PML) and/or Daxx to a viral DNA-protein complex. However, the presence of repressive proteins at the nuclear site of virus transcription has remained unexplained. We investigated the mouse cytomegalovirus (MCMV) immediate-early 1 protein (IE1), which is necessary for productive infection at low multiplicities of infection and therefore likely to be involved in overcoming cellular repression. Temporal analysis of IE1 distribution revealed its initial segregation into ND10 by binding to PML and/or Daxx and IE1-dependent recruitment of the transcriptional repressor histone deacetylase-2 (HDAC-2) to this site. However, these protein aggregates are dissociated in cells producing sufficient IE1 through titration of PML, Daxx, and HDAC-2. Importantly, binding of IE1 to HDAC-2 decreased deacetylation activity. Moreover, inhibition of HDAC by trichostatin-A resulted in an increase in viral protein synthesis, an increase in cells starting the formation of prereplication compartments, and an increase in the total infectious viruses produced. Thus, IE1, like trichostatin-A, reverses the repressive effect of HDAC evident in the presence of acetylated histones in the immediate-early promoter region. Since HDAC also binds to the promoter region of IE1, as determined by the chromatin immunoprecipitation assay, these combined results suggest that IE1 inhibits or reverses HDAC-mediated repression of the infecting viral genomes, possibly by a process akin to activation of heterochromatin. We propose that even permissive cells can repress transcription of infecting viral genomes through repressors, including HDAC, Daxx, and PML, and the segregation of IE1 to ND10 that would inactivate those repressors. The virus can counter this repression by overexpressing IE1 when present in sufficient copy number, thus reducing the availability and effectiveness of these repressors.
Polymorphism and Structural Maturation of Bunyamwera Virus in Golgi and Post-Golgi Compartments
Authors: Salanueva Iñigo J. , Novoa Reyes R. , Cabezas Pilar , López-Iglesias Carmen , Carrascosa José L. , Elliott Richard M. , Risco Cristina 
Published: December 2002

ABSTRACT

The Golgi apparatus is the assembly site for a number of complex enveloped viruses. Using high-preservation methods for electron microscopy, we have detected two previously unknown maturation steps in the morphogenesis of Bunyamwera virus in BHK-21 cells. The first maturation takes place inside the Golgi stack, where annular immature particles transform into dense, compact structures. Megalomicin, a drug that disrupts the
trans
side of the Golgi complex, reversibly blocks transformation, showing that a functional
trans
-Golgi is needed for maturation. The second structural change seems to take place during the egress of viral particles from cells, when a coat of round-shaped spikes becomes evident. A fourth viral assembly was detected in infected cells: rigid tubular structures assemble in the Golgi region early in infection and frequently connect with mitochondria. In Vero cells, the virus induces an early and spectacular fragmentation of intracellular membranes while productive infection progresses. Assembly occurs in fragmented Golgi stacks and generates tubular structures, as well as the three spherical viral forms. These results, together with our previous studies with nonrelated viruses, show that the Golgi complex contains key factors for the structural transformation of a number of enveloped viruses that assemble intracellularly.
Construction of an Excisable Bacterial Artificial Chromosome Containing a Full-Length Infectious Clone of Herpes Simplex Virus Type 1: Viruses Reconstituted from the Clone Exhibit Wild-Type Properties In Vitro and In Vivo
Authors: Tanaka Michiko , Kagawa Hiroyuki , Yamanashi Yuji , Sata Tetsutaro , Kawaguchi Yasushi 
Published: December 2002

ABSTRACT

In recent years, several laboratories have reported on the cloning of herpes simplex virus type 1 (HSV-1) genomes as bacterial artificial chromosomes (BACs) in
Escherichia coli
and on procedures to manipulate these genomes by using the bacterial recombination machinery. However, the HSV-BACs reported so far are either replication incompetent or infectious, with a deletion of one or more viral genes due to the BAC vector insertion. For use as a multipurpose clone in research on HSV-1, we attempted to generate infectious HSV-BACs containing the full genome of HSV-1 without any loss of viral genes. Our results were as follows. (i)
E. coli
(YEbac102) harboring the full-length HSV-1 genome (pYEbac102) in which a BAC flanked by
loxP
sites was inserted into the intergenic region between U
L
3 and U
L
4 was constructed. (ii) pYEbac102 was an infectious molecular clone, given that its transfection into rabbit skin cells resulted in production of infectious virus (YK304). (iii) The BAC vector sequence was almost perfectly excisable from the genome of the reconstituted virus YK304 by coinfection of Vero cells with YK304 and a recombinant adenovirus, AxCANCre, expressing Cre recombinase. (iv) As far as was examined, the reconstituted viruses from pYEbac102 could not be phenotypically differentiated from wild-type viruses in vitro and in vivo. Thus, the viruses grew as well in Vero cells as did the wild-type virus and exhibited wild-type virulence in mice on intracerebral inoculation. (v) The infectious molecular clone pYEbac102 is in fact useful for mutagenesis of the HSV-1 genome by bacterial genetics, and a recombinant virus carrying amino acid substitutions in both copies of the α0 gene was generated. pYEbac102 will have multiple applications to the rapid generation of genetically engineered HSV-1 recombinants in basic research into HSV-1 and in the development of HSV vectors in human therapy.
Cellular Gene Expression upon Human Immunodeficiency Virus Type 1 Infection of CD4+-T-Cell Lines
Authors: van 't Wout Angélique B. , Lehrman Ginger K. , Mikheeva Svetlana A. , O'Keeffe Gemma C. , Katze Michael G. , Bumgarner Roger E. , Geiss Gary K. , Mullins James I. 
Published: December 2002

ABSTRACTThe expression levels of ∼4,600 cellular RNA transcripts were assessed in CD4+-T-cell lines at different times after infection with human immunodeficiency virus type 1 strain BRU (HIV-1BRU) using DNA microarrays. We found that several classes of genes were inhibited by HIV-1BRUinfection, consistent with the G2arrest of HIV-1-infected cells induced by Vpr. These included genes involved in cell division and transcription, a family of DEAD-box proteins (RNA helicases), and all genes involved in translation and splicing. However, the overall level of cell activation and signaling was increased in infected cells, consistent with strong virus production. These included a subgroup of transcription factors, including EGR1 and JUN, suggesting they play a specific role in the HIV-1 life cycle. Some regulatory changes were cell line specific; however, the majority, including enzymes involved in cholesterol biosynthesis, of changes were regulated in most infected cell lines. Compendium analysis comparing gene expression profiles of our HIV-1 infection experiments to those of cells exposed to heat shock, interferon, or influenza A virus indicated that HIV-1 infection largely induced specific changes rather than simply activating stress response or cytokine response pathways. Thus, microarray analysis confirmed several known HIV-1 host cell interactions and permitted identification of specific cellular pathways not previously implicated in HIV-1 infection. Continuing analyses are expected to suggest strategies for impacting HIV-1 replication in vivo by targeting these pathways.
The 3C-Like Proteinase of an Invertebrate Nidovirus Links Coronavirus and Potyvirus Homologs
Authors: Ziebuhr John , Bayer Sonja , Cowley Jeff A. , Gorbalenya Alexander E. 
Published: December 2002

ABSTRACT

Gill-associated virus (GAV), a positive-stranded RNA virus of prawns, is the prototype of newly recognized taxa (genus
Okavirus
, family
Roniviridae
) within the order
Nidovirales.
In this study, a putative GAV cysteine proteinase (3C-like proteinase [3CL
pro
]), which is predicted to be the key enzyme involved in processing of the GAV replicase polyprotein precursors, pp1a and pp1ab, was characterized. Comparative sequence analysis indicated that, like its coronavirus homologs, 3CL
pro
has a three-domain organization and is flanked by hydrophobic domains. The putative 3CL
pro
domain including flanking regions (pp1a residues 2793 to 3143) was fused to the
Escherichia coli
maltose-binding protein (MBP) and, when expressed in
E. coli
, was found to possess N-terminal autoprocessing activity that was not dependent on the presence of the 3CL
pro
C-terminal domain. N-terminal sequence analysis of the processed protein revealed that cleavage occurred at the location
2827
LVTHE↓VRTGN
2836
. The
trans
-processing activity of the purified recombinant 3CL
pro
(pp1a residues 2832 to 3126) was used to identify another cleavage site,
6441
KVNHE↓LYHVA
6450
, in the C-terminal pp1ab region. Taken together, the data tentatively identify VxHE↓(L,V) as the substrate consensus sequence for the GAV 3CL
pro
. The study revealed that the GAV and potyvirus 3CL
pro
s possess similar substrate specificities which correlate with structural similarities in their respective substrate-binding sites, identified in sequence comparisons. Analysis of the proteolytic activities of MBP-3CL
pro
fusion proteins carrying replacements of putative active-site residues provided evidence that, in contrast to most other 3C/3CL
pro
s but in common with coronavirus 3CL
pro
s, the GAV 3CL
pro
employs a Cys
2968
-His
2879
catalytic dyad. The properties of the GAV 3CL
pro
define a novel RNA virus proteinase variant that bridges the gap between the distantly related chymotrypsin-like cysteine proteinases of coronaviruses and potyviruses.
Human Immunodeficiency Virus Type 1 Nucleocapsid Zn
2+
Fingers Are Required for Efficient Reverse Transcription, Initial Integration Processes, and Protection of Newly Synthesized Viral DNA
Authors: Buckman James S. , Bosche William J. , Gorelick Robert J. 
Published: December 2002

ABSTRACT

Human immunodeficiency virus type 1 (HIV-1) containing mutations in the nucleocapsid (NC) Zn
2+
finger domains have greatly reduced infectivity, even though genome packaging is largely unaffected in certain cases. To examine replication defects, viral DNA (vDNA) was isolated from cells infected with viruses containing His-to-Cys changes in their Zn
2+
fingers (NC
H23C
and NC
H44C
), an integrase mutant (IN
D116N
), a double mutant (NC
H23C
/IN
D116N
), or wild-type HIV-1. In vitro assays have established potential roles for NC in reverse transcription and integration. In vivo results for these processes were obtained by quantitative PCR, cloning of PCR products, and comparison of the quantity and composition of vDNA generated at discrete points during reverse transcription. Quantitative analysis of the reverse transcription intermediates for these species strongly suggests decreased stability of the DNA produced. Both Zn
2+
finger mutants appear to be defective in DNA synthesis, with the minus- and plus-strand transfer processes being affected while interior portions of the vDNA remain more intact. Sequences obtained from PCR amplification and cloning of 2-LTR circle junction fragments revealed that the NC mutants had a phenotype similar to the IN mutant; removal of the terminal CA dinucleotides necessary for integration of the vDNA is disabled by the NC mutations. Thus, the loss of infectivity in these NC mutants in vivo appears to result from defective reverse transcription and integration processes stemming from decreased protection of the full-length vDNA. Finally, these results indicate that the chaperone activity of NC extends from the management of viral RNA through to the full-length vDNA.
Two Human Immunodeficiency Virus Vaccinal Lipopeptides Follow Different Cross-Presentation Pathways in Human Dendritic Cells
Authors: Andrieu Muriel , Desoutter Jean-François , Loing Estelle , Gaston Jésintha , Hanau Daniel , Guillet Jean-Gérard , Hosmalin Anne 
Published: December 2002

ABSTRACTAn efficient vaccine against human immunodeficiency virus (HIV) must induce good cellular immune responses. To do this, it must be processed and presented by dendritic cells, which are required for primary T-lymphocyte stimulation. We have previously shown that a model lipopeptide containing a short epitopic peptide from HIV-1 was endocytosed and presented in association with major histocompatibility complex class I molecules by human dendritic cells to specific CD8+T lymphocytes, but the cross-presentation pathway needed to be precisely determined. We have studied a longer lipopeptide (Pol461-484) and another lipopeptide (Nef66-97) currently being used in vaccine trials. Like the shorter lipopeptide, the rhodamine-labeled Pol461-484lipopeptide was internalized by endocytosis, as assessed by confocal microscopy. The lipopeptides were processed by dendritic cells and presented to CD8+T cells specific for the HLA-A*0201-restricted Pol476-484and the HLA-A*0301-restricted Nef73-82epitope, respectively. Presentation of both lipopeptides was inhibited by brefeldin A. Presentation of the Pol lipopeptide was inhibited by epoxomycin, a proteasome-specific inhibitor, but not by monensin. This shows that it gained access to the cytosol to be digested by the proteasome. In contrast, presentation of the Nef lipopeptide was not inhibited by epoxomycin but was inhibited by monensin, a classical inhibitor of acid-dependent endosomal enzyme activity, indicating an endocytic processing pathway yielding to major histocompatibility complex class I-restricted presentation. Therefore, the two lipopeptides followed different cross-presentation pathways, both resulting in efficient presentation to CD8+T lymphocytes.
Hepatitis C Virus F Protein Is a Short-Lived Protein Associated with the Endoplasmic Reticulum
Authors: Xu Zhenming , Choi Jinah , Lu Wen , Ou Jing-hsiung 
Published: December 2002

ABSTRACT
Hepatitis C virus (HCV) F protein is a newly discovered HCV gene product that is expressed by translational ribosomal frameshift. Little is known about the biological properties of this protein. By performing pulse-chase labeling experiments, we demonstrate here that the F protein is a labile protein with a half-life of <10 min in Huh7 hepatoma cells and in vitro. The half-life of the F protein could be substantially increased by proteasome inhibitors, suggesting that the rapid degradation of the F protein is mediated by the proteasome pathway. Further immunofluorescence staining and subcellular fractionation experiments indicate that the F protein is primarily associated with the endoplasmic reticulum. This subcellular localization is similar to those of HCV core and NS5A proteins, raising the possibility that the F protein may participate in HCV morphogenesis or replication.
Common Sites of Retroviral Integration in Mouse Hematopoietic Tumors Identified by High-Throughput, Single Nucleotide Polymorphism-Based Mapping and Bacterial Artificial Chromosome Hybridization
Authors: Shen Haifa , Suzuki Takeshi , Munroe David J. , Stewart Claudia , Rasmussen Lynn , Gilbert Debra J. , Jenkins Nancy A. , Copeland Neal G. 
Published: December 2002

ABSTRACT

Retroviral insertional mutagenesis in mouse hematopoietic tumors provides a powerful cancer gene discovery tool. Here, we describe a high-throughput, single nucleotide polymorphism (SNP)-based method, for mapping retroviral integration sites cloned from mouse tumors, and a bacterial artificial chromosome (BAC) hybridization method, for localizing these retroviral integration sites to common sites of retroviral integration (CISs). Several new CISs were identified, including one CIS that mapped near
Notch1
, a gene that has been causally associated with human T-cell tumors. This mapping method is applicable to many different species, including ones where few genetic markers and little genomic sequence information are available. It can also be used to map endogenous proviruses.
Recognition of Native Hepatitis C Virus E1E2 Heterodimers by a Human Monoclonal Antibody
Authors: Cocquerel Laurence , Quinn Elizabeth R. , Flint Mike , Hadlock Kenneth G. , Foung Steven K. H. , Levy Shoshana 
Published: December 2002

ABSTRACT
The majority of hepatitis C virus (HCV)-infected individuals progress from acute to chronic disease, despite the presence of a strong humoral immune response to the envelope glycoproteins E1 and E2. When expressed in mammalian cells, E1 and E2 form both noncovalently linked E1E2 heterodimers, believed to be properly folded, and disulfide-linked, high-molecular-weight aggregates that are misfolded. Previously, we identified 10 human monoclonal antibodies (HMAbs) that bind E2 glycoproteins from different genotypes. Here we demonstrate that one of these HMAbs, CBH-2, is unique in its ability to distinguish between properly folded and misfolded envelope proteins. This HMAb recognizes HCV-E2 only when complexed with E1. The E1E2 complexes recognized by CBH-2 are noncovalently linked heterodimers and not misfolded disulfide-linked, high-molecular-weight aggregates. The E1E2 heterodimers seen by CBH-2 no longer associate with the endoplasmic reticulum chaperone calnexin and are likely to represent the prebudding form of the HCV virion.
Baseline Susceptibility of Primary Human Immunodeficiency Virus Type 1 to Entry Inhibitors
Authors: Labrosse Béatrice , Labernardière Jean-Louis , Dam Elisabeth , Trouplin Virginie , Skrabal Katharina , Clavel François , Mammano Fabrizio 
Published: December 2002

ABSTRACT
Human immunodeficiency virus type 1 plasma viruses from 29 entry inhibitor-naive patients were characterized for their susceptibilities to T-20, AMD3100, and RANTES. A strikingly wide range of susceptibilities to T-20 was observed that was influenced by coreceptor usage but not by the susceptibilities of the viruses to inhibitors that target the chemokine receptors or by polymorphisms in the gp41 N helix.