bluuebunny/crossref_metadata_2025_split huggingface dataset:
Promoter-Specific Targeting of Human SWI-SNF Complex by Epstein-Barr Virus Nuclear Protein 2
Authors: Wu Daniel Y. , Krumm Anton , Schubach William H. 
Published: July 2002

ABSTRACT
The multiprotein human SWI-SNF (hSWI-SNF) complex is a chromatin-remodeling machine that facilitates transcription by overcoming chromatin-mediated gene repression. We had previously shown that hSNF5/INI1, an intrinsic, consistent component of the hSWI/SNF complex, is associated with Epstein-Barr nuclear antigen 2 (EBNA2) and have proposed that EBNA2 directs this complex to key EBNA2-responsive viral and cellular genes. Using chromatin immunoprecipitation and quantitative PCR, we show that antibodies directed against components of the hSWI-SNF complex preferentially precipitate chromatin-associated DNA that contains a targeted EBNA2-responsive element in the context of both episomal and cellular chromatin. This enrichment does not occur in EBNA2-negative cells or when the EBNA2-responsive element is mutated. The stable association of the hSWI-SNF complex with the EBNA2-responsive promoter can also be disrupted by deletion of the TATA element, suggesting that EBNA2 in itself is insufficient to mediate stable targeting of the hSWI-SNF complex. These results demonstrate that recruitment of the hSWI-SNF complex to selected promoters can occur in vivo through its interaction with site-specific activator proteins and that stable targeting may require the presence of basal transcription factors.
Biology of Attenuated Modified Vaccinia Virus Ankara Recombinant Vector in Mice: Virus Fate and Activation of B- and T-Cell Immune Responses in Comparison with the Western Reserve Strain and Advantages as a Vaccine
Authors: Ramírez Juan C. , Gherardi M. Magdalena , Esteban Mariano 
Published: July 2002

ABSTRACTThe modified vaccinia virus Ankara (MVA) strain is a candidate vector for vaccination against pathogens and tumors, due to safety concerns and the proven ability of recombinants based on this vector to trigger protection against pathogens in animals. In this study we addressed the fate of the MVA vector in BALB/c mice after intraperitoneal inoculation in comparison with that of the replication-competent Western Reserve (WR) strain by measuring levels of expression of the reporter luciferase gene, the capability to infect target tissues from the site of inoculation, and the length of time of virus persistence. We evaluated the extent of humoral and cellular immune responses induced against the virus antigens and a recombinant product (β-galactosidase). We found that MVA infects the same target tissues as the WR strain; surprisingly, within 6 h postinoculation the levels of expression of antigens were higher in tissues from MVA-infected mice than in tissues from mice infected with wild-type virus but at later times postinoculation were 2 to 4 log units higher in tissues from WR-infected mice. In spite of this, antibodies and cellular immune responses to viral vector antigens were considerably lower in MVA-inoculated mice than in WR virus-inoculated mice. In contrast, the cellular immune response to a foreign antigen expressed from MVA was similar to and even higher than that triggered by the recombinant WR virus. MVA elicited a Th1 type of immune response, and the main proinflammatory cytokines induced were interleukin-6 and tumor necrosis factor alpha. Our findings have defined the biological characteristics of MVA infection in tissues and the immune parameters activated in the course of virus infection. These results are of significance with respect to optimal use of MVA as a vaccine.
Infection of Dendritic Cells by the Maedi-Visna Lentivirus
Authors: Ryan Susanna , Tiley Laurence , McConnell Ian , Blacklaws Barbara 
Published: July 2002

ABSTRACTThe early stages of lentivirus infection of dendritic cells have been studied in an in vivo model. Maedi-visna virus (MVV) is a natural pathogen of sheep with a tropism for macrophages, but the infection of dendritic cells has not been proven, largely because of the difficulties of definitively distinguishing the two cell types. Afferent lymphatic dendritic cells from sheep have been phenotypically characterized and separated from macrophages. Dendritic cells purified from experimentally infected sheep have been demonstrated not only to carry infectious MVV but also to be hosts of the virus themselves. The results of the in vivo infection experiments are supported by infections of purified afferent lymph dendritic cells in vitro, in which late reverse transcriptase products are demonstrated by PCR. The significance of the infection of afferent lymph dendritic cells is discussed in relation to the initial spread of lentivirus infection and the requirement for CD4 T cells.
Template Nucleotide Moieties Required for De Novo Initiation of RNA Synthesis by a Recombinant Viral RNA-Dependent RNA Polymerase
Authors: Kim Min-Ju , Zhong Weidong , Hong Zhi , Kao C. Cheng 
Published: July 2002

ABSTRACT
The recombinant RNA-dependent RNA polymerase of the bovine viral diarrhea virus specifically requires a cytidylate at the 3′ end for the de novo initiation of RNA synthesis (C. C. Kao, A. M. Del Vecchio, and W. Zhong, Virology 253:1–7, 1999). Using RNAs containing nucleotide analogs, we found that the N3 and C4-amino group at the initiation cytidine were required for RNA synthesis. However, the ribose C2′-hydroxyl of the initiating cytidylate can accept several modifications and retain the ability to direct synthesis. The only unacceptable modification is a protonated C2′-amino group. Quite strikingly, the recognition of the functional groups for the initiation cytidylate and other template nucleotides are different. For example, a C5-methyl group in cytidine can direct RNA synthesis at all template positions except at the initiation cytidylate and C2′-amino modifications are tolerated better after the +11 position. When a 4-thiouracil (4sU) base analog that allows only imperfect base pairing with the nascent RNA is placed at different positions in the template, the efficiency of synthesis is correlated with the calculated stability of the template-nascent RNA duplex adjacent to the position of the 4sU. These results define the requirements for the specific interactions required for the initiation of RNA synthesis and will be compared to the mechanisms of initiation by other RNA-dependent and DNA-dependent RNA polymerases.
The N-Terminal V3 Loop Glycan Modulates the Interaction of Clade A and B Human Immunodeficiency Virus Type 1 Envelopes with CD4 and Chemokine Receptors
Authors: Malenbaum Susan E. , Yang David , Cavacini Lisa , Posner Marshall , Robinson James , Cheng-Mayer Cecilia 
Published: July 2002

ABSTRACTWe investigated the underlying mechanism by which the highly conserved N-terminal V3 loop glycan of gp120 conferred resistance to neutralization of human immunodeficiency virus type 1 (HIV-1). We find that the presence or absence of this V3 glycan on clade A and B viruses accorded various degrees of susceptibility to neutralization by antibodies to the CD4 binding site, CD4-induced epitopes, and chemokine receptors. Our data suggest that this carbohydrate moiety on gp120 blocks access to the binding site for CD4 and modulates the chemokine receptor binding site of phenotypically diverse clade A and clade B isolates. Its presence also contributes to the masking of CD4-induced epitopes on clade B envelopes. These findings reveal a common mechanism by which diverse HIV-1 isolates escape immune recognition. Furthermore, the observation that conserved functional epitopes of HIV-1 are more exposed on V3 glycan-deficient envelope glycoproteins provides a basis for exploring the use of these envelopes as vaccine components.
The Carboxy-Terminal Fragment of Nucleolin Interacts with the Nucleocapsid Domain of Retroviral Gag Proteins and Inhibits Virion Assembly
Authors: Bacharach Eran , Gonsky Jason , Alin Kimona , Orlova Marianna , Goff Stephen P. 
Published: July 2002

ABSTRACT
A yeast two-hybrid screen for cellular proteins that interact with the murine leukemia virus (MuLV) Gag protein resulted in the identification of nucleolin, a host protein known to function in ribosome assembly. The interacting fusions contained the carboxy-terminal 212 amino acids of nucleolin [Nuc(212)]. The nucleocapsid (NC) portion of Gag was necessary and sufficient to mediate the binding to Nuc(212). The interaction of Gag with Nuc(212) could be demonstrated in vitro and was manifested in vivo by the NC-dependent incorporation of Nuc(212) inside MuLV virions. Overexpression of Nuc(212), but not full-length nucleolin, potently and specifically blocked MuLV virion assembly and/or release. A mutant of MuLV, selected to specifically disrupt the binding to Nuc(212), was found to be severely defective for virion assembly. This mutant harbors a single point mutation in capsid (CA) adjacent to the CA-NC junction, suggesting a role for this region in Moloney MuLV assembly. These experiments demonstrate that selection for proteins that bind assembly domain(s) can yield potent inhibitors of virion assembly. These experiments also raise the possibility that a nucleolin-Gag interaction may be involved in virion assembly.
Identification of a Novel Consensus Sequence at the Cleavage Site of the Lassa Virus Glycoprotein
Authors: Lenz Oliver , ter Meulen Jan , Feldmann Heinz , Klenk Hans-Dieter , Garten Wolfgang 
Published: July 2002

ABSTRACT

The Lassa virus glycoprotein consists of an amino-terminal and a carboxy-terminal cleavage fragment designated GP-1 and GP-2, respectively, that are derived by proteolysis from the precursor GP-C. The membrane-anchored GP-2 obtained from purified virions of the Josiah strain revealed the N-terminal tripeptide GTF
262
when analyzed by Edman degradation. Upstream of this site, GP-C contains the tetrapeptide sequence RRLL
259
, which is conserved in all Lassa virus isolates published to date. Systematic mutational analysis of vector-expressed GP-C revealed that the motif R-X (L/I/V)-L
259
(where X stands for L, I, or V) is essential for cleavage of the peptide bond between leucine
259
and glycine
260
. This cleavage motif is homologous to the consensus sequence recognized by a novel class of cellular endoproteases which have so far not been implicated in the processing of viral glycoproteins.
Toward a More Accurate Quantitation of the Activity of Recombinant Retroviruses: Alternatives to Titer and Multiplicity of Infection
Authors: Andreadis Stylianos , Lavery Thomas , Davis Howard E. , Le Doux Joseph M. , Yarmush Martin L. , Morgan Jeffrey R. 
Published: July 2002

ABSTRACT

In this paper, we present a mathematical model with experimental support of how several key parameters govern the adsorption of active retrovirus particles onto the surface of adherent cells. These parameters, including time of adsorption, volume of virus, and the number, size, and type of target cells, as well as the intrinsic properties of the virus, diffusion coefficient, and half-life (
t
1/2
), have been incorporated into a mathematical expression that describes the rate at which active virus particles adsorb to the cell surface. From this expression, we have obtained estimates of
C
vo
, the starting concentration of active retrovirus particles. In contrast to titer,
C
vo
is independent of the specific conditions of the assay. The relatively slow diffusion (
D
= 2 × 10
−8
cm
2
/s) and rapid decay (
t
1/2
= 6 to 7 h) of retrovirus particles explain why
C
vo
values are significantly higher than titer values. Values of
C
vo
also indicate that the number of defective particles in a retrovirus stock is much lower than previously thought, which has implications especially for the use of retroviruses for in vivo gene therapy. With this expression, we have also computed AVC (active viruses/cell), the number of active retrovirus particles that would adsorb per cell during a given adsorption time. In contrast to multiplicity of infection, which is based on titer and is subject to the same inaccuracies, AVC is based on the physicochemical parameters of the transduction assay and so is a more reliable alternative.
Assembly of Spikes into Coronavirus Particles Is Mediated by the Carboxy-Terminal Domain of the Spike Protein
Authors: Godeke Gert-Jan , de Haan Cornelis A. M. , Rossen John W. A. , Vennema Harry , Rottier Peter J. M. 
Published: July 2002

ABSTRACT
The type I glycoprotein S of coronavirus, trimers of which constitute the typical viral spikes, is assembled into virions through noncovalent interactions with the M protein. Here we demonstrate that incorporation is mediated by the short carboxy-terminal segment comprising the transmembrane and endodomain. To this aim, we used the virus-like particle (VLP) system that we developed earlier for the mouse hepatitis virus strain A59 (MHV-A59) and which we describe now also for the unrelated coronavirus feline infectious peritonitis virus (FIPV; strain 79-1146). Two chimeric MHV-FIPV S proteins were constructed, consisting of the ectodomain of the one virus and the transmembrane and endodomain of the other. These proteins were tested for their incorporation into VLPs of either species. They were found to assemble only into viral particles of the species from which their carboxy-terminal domain originated. Thus, the 64-terminal-residue sequence suffices to draw the 1308 (MHV)- or 1433 (FIPV)-amino-acid-long mature S protein into VLPs. Both chimeric S proteins appeared to cause cell fusion when expressed individually, suggesting that they were biologically fully active. This was indeed confirmed by incorporating one of the proteins into virions which thereby acquired a new host cell tropism, as will be reported elsewhere.
De Novo Initiation of RNA Synthesis by Hepatitis C Virus Nonstructural Protein 5B Polymerase
Authors: Zhong Weidong , Uss Annette S. , Ferrari Eric , Lau Johnson Y. N. , Hong Zhi 
Published: July 2002

ABSTRACT

RNA-dependent RNA polymerase (RdRp) encoded by positive-strand RNA viruses is critical to the replication of viral RNA genome. Like other positive-strand RNA viruses, replication of hepatitis C virus (HCV) RNA is mediated through a negative-strand intermediate, which is generated through copying the positive-strand genomic RNA. Although it has been demonstrated that HCV NS5B alone can direct RNA replication through a copy-back primer at the 3′ end, de novo initiation of RNA synthesis is likely to be the mode of RNA replication in infected cells. In this study, we demonstrate that a recombinant HCV NS5B protein has the ability to initiate de novo RNA synthesis in vitro. The NS5B used HCV 3′ X-tail RNA (98 nucleotides) as the template to synthesize an RNA product of monomer size, which can be labeled by [γ-
32
P]nucleoside triphosphate. The de novo initiation activity was further confirmed by using small synthetic RNAs ending with dideoxynucleotides at the 3′ termini. In addition, HCV NS5B preferred GTP as the initiation nucleotide. The optimal conditions for the de novo initiation activity have been determined. Identification and characterization of the de novo priming or initiation activity by HCV NS5B provides an opportunity to screen for inhibitors that specifically target the initiation step.
Epstein-Barr Virus Entry Utilizing HLA-DP or HLA-DQ as a Coreceptor
Authors: Haan Keith M. , Kwok William W. , Longnecker Richard , Speck Peter 
Published: July 2002

ABSTRACTEpstein-Barr virus (EBV) glycoprotein gp350/gp220 association with cellular CD21 facilitates virion attachment to B lymphocytes. Membrane fusion requires the additional interaction between virion gp42 and cellular HLA-DR. This binding is thought to catalyze membrane fusion through a further association with the gp85-gp25 (gH-gL) complex. Cell lines expressing CD21 but lacking expression of HLA class II molecules are resistant to infection by a recombinant EBV expressing enhanced green fluorescent protein. Surface expression of HLA-DR, HLA-DP, or HLA-DQ confers susceptibility to EBV infection on resistant cells that express CD21. Therefore, HLA-DP or HLA-DQ can substitute for HLA-DR and serve as a coreceptor in EBV entry.
The Kinetics of VP5 mRNA Expression Is Not Critical for Viral Replication in Cultured Cells
Authors: Lieu Pauline T. , Wagner Edward K. 
Published: July 2002

ABSTRACTWe generated recombinant viruses in which the kinetics of expression of the leaky-late VP5 mRNA was altered. We then analyzed the effect of such alterations on viral replication in cultured cells. The VP5 promoter and leader sequences from positions −36 to +20, containing the TATA box and an initiator element, were deleted and replaced with a strong early (dUTPase), an equal-strength leaky-late (VP16), or a strict-late (UL38) promoter. We found that recombinant viruses containing the dUTPase promoter inserted in the VP5 locus expressed VP5-encoding mRNA with early kinetics, while virus with the UL38 promoter inserted expressed such mRNA with strict-late kinetics. Further, in spite of differences in its functional architecture, the VP16 promoter fully substituted for the VP5 promoter. Western blot analysis demonstrated that the amounts of VP5 capsid protein produced by the recombinant viruses differed somewhat; however, on complementing C32 and noncomplementing Vero cells, such viruses replicated to titers equivalent to those of the rescued wild-type virus controls. Multistep virus growth in mouse embryo fibroblasts, rabbit skin cells, and Vero cells also demonstrated equivalent replication efficiencies for both recombinant and wild-type viruses. Further, recombinant viruses did not show any impairment in their ability to replicate on serum-starved or quiescent human lung fibroblasts. We conclude that the kinetics of the essential VP5 mRNA expression is not critical for viral replication in cultured cells.
Identification and Characterization of a Shared TNFR-Related Receptor for Subgroup B, D, and E Avian Leukosis Viruses Reveal Cysteine Residues Required Specifically for Subgroup E Viral Entry
Authors: Adkins Heather B. , Brojatsch Jürgen , Young John A. T. 
Published: July 2002

ABSTRACT

Genetic and receptor interference data have indicated the presence of one or more cellular receptors for subgroup B, D, and E avian leukosis viruses (ALV) encoded by the
s1
allele of the chicken
tvb
locus. Despite the prediction that these viruses use the same receptor, they exhibit a nonreciprocal receptor interference pattern: ALV-B and ALV-D can interfere with infection by all three viral subgroups, but ALV-E only interferes with infection by subgroup E viruses. We identified a
tvb
s1
cDNA clone which encodes a tumor necrosis factor receptor-related receptor for ALV-B, -D, and -E. The nonreciprocal receptor interference pattern was reconstituted in transfected human 293 cells by coexpressing the cloned receptor with the envelope (Env) proteins of either ALV-B or ALV-E. This pattern of interference was also observed when soluble ALV surface (SU)-immunoglobulin fusion proteins were bound to this cellular receptor before viral challenge. These data demonstrate that viral Env-receptor interactions can account for the nonreciprocal interference between ALV subgroups B, D, and E. Furthermore, they indicate that a single chicken gene located at
tvb
s1
encodes receptors for these three viral subgroups. The TVB
S1
protein differs exclusively at residue 62 from the published subgroup B- and D-specific receptor, encoded by the
s3
allele of
tvb
. Residue 62 is a cysteine in TVB
S1
but is a serine in TVB
S3
, giving TVB
S1
an even number of cysteines in the extracellular domain. We present evidence for a disulfide bond requirement in TVB
S1
for ALV-E infection but not for ALV-B infection. Thus, ALV-B and ALV-E interact in fundamentally different ways with this shared receptor, a finding that may account for the observed biological differences between these two ALV subgroups.
Receptor Specificities of Human Respiroviruses
Authors: Suzuki Takashi , Portner Allen , Scroggs Ruth Ann , Uchikawa Makoto , Koyama Noriko , Matsuo Kazuko , Suzuki Yasuo , Takimoto Toru 
Published: July 2002

ABSTRACT

Through their hemagglutinin-neuraminidase glycoprotein, parainfluenza viruses bind to sialic acid-containing glycoconjugates to initiate infection. Although the virus-receptor interaction is a key factor of infection, the exact nature of the receptors that human parainfluenza viruses recognize has not been determined. We evaluated the abilities of human parainfluenza virus types 1 (hPIV-1) and 3 (hPIV-3) to bind to different types of gangliosides. Both hPIV-1 and hPIV-3 preferentially bound to neolacto-series gangliosides containing a terminal
N
-acetylneuraminic acid (NeuAc) linked to
N
-acetyllactosamine (Galβ1-4GlcNAc) by the α2-3 linkage (NeuAcα2-3Galβ1-4GlcNAc). Unlike hPIV-1, hPIV-3 bound to gangliosides with a terminal NeuAc linked to Galβ1-4GlcNAc through an α2-6 linkage (NeuAcα2-6Galβ1-4GlcNAc) or to gangliosides with a different sialic acid,
N
-glycolylneuraminic acid (NeuGc), linked to Galβ1-4GlcNAc (NeuGcα2-3Galβ1-4GlcNAc). These results indicate that the molecular species of glycoconjugate that hPIV-1 recognizes are more limited than those recognized by hPIV-3. Further analysis using purified gangliosides revealed that the oligosaccharide core structure is also an important element for binding. Gangliosides that contain branched
N
-acetyllactosaminoglycans in their core structure showed higher avidity than those without them. Agglutination of human, cow, and guinea pig erythrocytes but not equine erythrocytes by hPIV-1 and hPIV-3 correlated well with the presence or the absence of sialic acid-linked branched
N
-acetyllactosaminoglycans on the cell surface. Finally, NeuAcα2-3I, which bound to both viruses, inhibited virus infection of Lewis lung carcinoma-monkey kidney cells in a dose-dependent manner. We conclude that hPIV-1 and hPIV-3 preferentially recognize oligosaccharides containing branched
N
-acetyllactosaminoglycans with terminal NeuAcα2-3Gal as receptors and that hPIV-3 also recognizes NeuAcα2-6Gal- or NeuGcα2-3Gal-containing receptors. These findings provide important information that can be used to develop inhibitors that prevent human parainfluenza virus infection.
Induction of pRb Degradation by the Human Papillomavirus Type 16 E7 Protein Is Essential To Efficiently Overcome p16
INK4a
-Imposed G
1
Cell Cycle Arrest
Authors: Giarrè Marianna , Caldeira Sandra , Malanchi Ilaria , Ciccolini Francesca , Leão Maria João , Tommasino Massimo 
Published: July 2002

ABSTRACT

It has previously been shown that the E7 protein from the cutaneous human papillomavirus type 1 (HPV1), which is associated with benign skin lesions, binds the product of the tumor suppressor gene retinoblastoma (pRb) with an efficiency similar to that of the E7 protein from the oncogenic HPV type 16. Despite this ability, HPV1 E7 does not display any activity in transforming primary cells. In addition, the two viral proteins differ in their mechanisms of targeting pRb. HPV16 E7 promotes pRb destabilization, while cells expressing HPV1 E7 do not show any decrease in pRb levels. In this study, we show that HPV1 E7, in contrast to HPV16 E7, has only a weak activity to neutralize the effect of cyclin-dependent kinase inhibitor p16
INK4a
. By generation of HPV1/16 E7 chimeric proteins, we have identified a central motif in the two E7 proteins, which determines their different abilities to overcome the p16
INK4a
-mediated cell cycle arrest. This motif is located downstream of the pRb-binding domain and comprises only three amino acids in HPV16 E7. Swapping this central motif in the two viral proteins causes an exchange of their activities involved in circumventing the inhibitory function of p16
INK4a
. Most importantly, our data show that the efficiency of the E7 proteins in neutralizing the inhibitory effect of p16
INK4a
correlates with their ability to promote pRb degradation.