bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression of an Antigenic Adenovirus Epitope in a Group B Coxsackievirus
Authors: Höfling Katja , Tracy Steven , Chapman Nora , Kim Kyung-Soo , Smith Leser J. 
Published: July 2002

ABSTRACT
Group B coxsackieviruses (CVB) cause human myocarditis, while human adenovirus type 2 (Ad2) is implicated as an agent of this disease. The L1 loop of the Ad2 hexon protein has been demonstrated to be antigenic in rabbits. To evaluate the feasibility of a multivalent vaccine strain against the CVB and Ad2, we cloned the sequence encoding the Ad2 hexon L1 loop, flanked by dissimilar sequences encoding the protease 2A (2Apro) recognition sites, into the genome of an attenuated strain of CVB type 3 (CVB3/0) at the junction of 2Apro and the capsid protein 1D. Progeny virus (CVB3-PL2-Ad2L1) was obtained following transfection of the construct into HeLa cells. Replication of CVB3-PL2-Ad2L1 in diverse cell cultures demonstrated that the yield of the chimeric virus was between 0.5 to 2 log units less than the parental strain. Western blot analyses of the CVB3 capsid protein 1D in CVB3-PL2-Ad2L1-infected HeLa cells demonstrated production of the expected capsid protein. Viral proteins were detected earlier and in approximately fourfold greater amounts in CVB3-PL2-Ad2L1-infected HeLa cells than in CVB3/0-infected cells. Cleavage of the CVB3-PL2-Ad2L1 polyprotein by 2Apro was slowed, accompanied by an accumulation of the fusion 1D-L1 loop protein. Reverse transcription-PCR sequence analysis of CVB3-PL2-Ad2L1 RNA demonstrated that the Ad2 hexon polypeptide coding sequence was maintained in the chimeric viral genome through at least 10 passages in HeLa cells. Mice inoculated with CVB3-PL2-Ad2L1 demonstrated a brief viremia with no replication detectable in the heart but prolonged replication of virus in the pancreas in the absence of pathologic changes in either organ. CVB3-PL2-Ad2L1 induced binding and neutralizing anti-Ad2 antibodies, in addition to antibodies against CVB3 in mice. CVB3-PL2-Ad2L1 was used to challenge mice previously inoculated with CVB3/0 and with preexisting anti-CVB3 neutralizing-antibody titers; anti-Ad2 neutralizing and binding antibodies were induced in these mice at higher levels than in mice without anti-CVB3 immunity. The data demonstrate that a CVB vector can stably express an antigenic polypeptide of Ad2 from within the CVB open reading frame that results in the induction of protective immune responses against both viruses.
Identification of a Key Target Sequence To Block Human Immunodeficiency Virus Type 1 Replication within the
gag-pol
Transframe Domain
Authors: Sei Shizuko , Yang Quan-en , O'Neill Dennis , Yoshimura Kazuhisa , Nagashima Kunio , Mitsuya Hiroaki 
Published: July 2002

ABSTRACT

Although the full sequence of the human immunodeficiency virus type 1 (HIV-1) genome has been known for more than a decade, effective genetic antivirals have yet to be developed. Here we show that, of 22 regions examined, one highly conserved sequence (ACTCTTTGGCAACGA) near the 3′ end of the HIV-1
gag-pol
transframe region, encoding viral protease residues 4 to 8 and a C-terminal Vpr-binding motif of p6
Gag
protein in two different reading frames, can be successfully targeted by an antisense peptide nucleic acid oligomer named PNA
PR2
. A disrupted translation of
gag-pol
mRNA induced at the PNA
PR2
-annealing site resulted in a decreased synthesis of Pr160
Gag-Pol
polyprotein, hence the viral protease, a predominant expression of Pr55
Gag
devoid of a fully functional p6
Gag
protein, and the excessive intracellular cleavage of Gag precursor proteins, hindering the processes of virion assembly. Treatment with PNA
PR2
abolished virion production by up to 99% in chronically HIV-1-infected H9 cells and in peripheral blood mononuclear cells infected with clinical HIV-1 isolates with the multidrug-resistant phenotype. This particular segment of the
gag-pol
transframe gene appears to offer a distinctive advantage over other regions in invading viral structural genes and restraining HIV-1 replication in infected cells and may potentially be exploited as a novel antiviral genetic target.
Octamerization Enables Soluble CD46 Receptor To Neutralize Measles Virus In Vitro and In Vivo
Authors: Christiansen Dale , Devaux Patricia , Réveil Brigitte , Evlashev Alexey , Horvat Branka , Lamy Josette , Rabourdin-Combe Chantal , Cohen Jacques H. M. , Gerlier Denis 
Published: July 2002

ABSTRACTA chimeric fusion protein encompassing the CD46 ectodomain linked to the C-terminal part of the C4b binding protein (C4bp) α chain (sCD46-C4bpα) was produced in eukaryotic cells. This protein, secreted as a disulfide-linked homo-octamer, was recognized by a panel of anti-CD46 antibodies with varying avidities. Unlike monomeric sCD46, the octameric sCD46-C4bpα protein was devoid of complement regulatory activity. However, sCD46-C4bpα was able to bind to the measles virus hemagglutinin protein expressed on murine cells with a higher avidity than soluble monomeric sCD46. Moreover, the octameric sCD46-C4bpα protein was significantly more efficient than monomeric sCD46 in inhibiting virus binding to CD46, in blocking virus induced cell-cell fusion, and in neutralizing measles virus in vitro. In addition, the octameric sCD46-C4bpα protein, but not the monomeric sCD46, fully protected CD46 transgenic mice against a lethal intracranial measles virus challenge.
Retrovirus Vectors Bearing Jaagsiekte Sheep Retrovirus Env Transduce Human Cells by Using a New Receptor Localized to Chromosome 3p21.3
Authors: Rai Sharath K. , DeMartini James C. , Miller A. Dusty 
Published: July 2002

ABSTRACT

Jaagsiekte sheep retrovirus (JSRV) is a type D retrovirus associated with a contagious lung tumor of sheep, ovine pulmonary carcinoma. Other than sheep, JSRV is known to infect goats, but there is no evidence of human infection. Until now it has not been possible to study the host range for JSRV because of the inability to grow this virus in culture. Here we show that the JSRV envelope protein (Env) can be used to pseudotype Moloney murine leukemia virus (MoMLV)-based retrovirus vectors and that such vectors can transduce human cells in culture. We constructed hybrid retrovirus packaging cells that express the JSRV Env and the MoMLV Gag-Pol proteins and can produce JSRV-pseudotype vectors at titers of up to 10
6
alkaline phosphatase-positive focus-forming units/ml. Using this high-titer virus, we have studied the host range for JSRV, which includes sheep, human, monkey, bovine, dog, and rabbit cells but not mouse, rat, or hamster cells. Considering the inability of the JSRV-pseudotype vector to transduce hamster cells, we used the hamster cell line-based Stanford G3 panel of whole human genome radiation hybrids to phenotypically map the JSRV receptor (
JVR
) gene within the p21.3 region of human chromosome 3. JVR is likely a new retrovirus receptor, as none of the previously identified retrovirus receptors localizes to the same position. Several chemokine receptors that have been shown to serve as coreceptors for lentivirus infection are clustered in the same region of chromosome 3; however, careful examination shows that the JSRV receptor does not colocalize with any of these genes.
Coreceptor Competition for Association with CD4 May Change the Susceptibility of Human Cells to Infection with T-Tropic and Macrophagetropic Isolates of Human Immunodeficiency Virus Type 1
Authors: Lee Shirley , Lapham Cheryl K. , Chen Hong , King Lisa , Manischewitz Jody , Romantseva Tatiana , Mostowski Howard , Stantchev Tzanko S. , Broder Christopher C. , Golding Hana 
Published: July 2002

ABSTRACTThe chemokine receptors CCR5 and CXCR4 were found to function in vivo as the principal coreceptors for M-tropic and T-tropic human immunodeficiency virus (HIV) strains, respectively. Since many primary cells express multiple chemokine receptors, it was important to determine if the efficiency of virus-cell fusion is influenced not only by the presence of the appropriate coreceptor (CXCR4 or CCR5) but also by the levels of other coreceptors expressed by the same target cells. We found that in cells with low to medium surface CD4 density, coexpression of CCR5 and CXCR4 resulted in a significant reduction in the fusion with CXCR4 domain (X4) envelope-expressing cells and in their susceptibility to infection with X4 viruses. The inhibition could be reversed either by increasing the density of surface CD4 or by antibodies against the N terminus and second extracellular domains of CCR5. In addition, treatment of macrophages with a combination of anti-CCR5 antibodies or β-chemokines increased their fusion with X4 envelope-expressing cells. Conversely, overexpression of CXCR4 compared with CCR5 inhibited CCR5-dependent HIV-dependent fusion in 3T3.CD4.401 cells. Thus, coreceptor competition for association with CD4 may occur in vivo and is likely to have important implications for the course of HIV type 1 infection, as well as for the outcome of coreceptor-targeted therapies.
The Hepatitis B Virus Core Promoter Is Strongly Activated by the Liver Nuclear Receptor Fetoprotein Transcription Factor or by Ectopically Expressed Steroidogenic Factor 1
Authors: Gilbert Stéphane , Galarneau Luc , Lamontagne Alain , Roy Sylvie , Bélanger Luc 
Published: July 2002

ABSTRACT

Orphan nuclear receptor fetoprotein transcription factor (FTF) was previously identified as a specific regulator of the α
1
-fetoprotein gene during early liver development and in response to hormonal signals (L. Galarneau, J.-F. Paré, D. Allard, D. Hamel, L. Lévesque, J. D. Tugwood, S. Green, and L. Bélanger, Mol. Cell. Biol. 16:3853–3865, 1996). Here we report a functional analysis of FTF interactions with the hepatitis B virus (HBV) nucleocapsid promoter. DNA-protein-binding assays show that the HBV core promoter contains two high-affinity FTF-binding sites and a third, lower-affinity site shared with other receptors. Transfections in HepG2, Hep3B, and PLC/PRF/5 hepatoma cells using chloramphenicol acetyltransferase reporter genes with the nucleocapsid promoter linked or not linked to enhancer I indicate that FTF is a potent activator of the HBV core promoter, more efficient than HNF4α, HNF3α, HNF3β, or C/EBPα. Steroidogenic factor 1, a close FTF homolog which binds to the same DNA motif and is expressed ectopically in HepG2 cells, seems to be an even stronger inducer than FTF. Point mutations of the FTF-binding sites indicate direct FTF activatory effects on the core promoter and the use of both high-affinity sites for productive interaction between the core promoter and enhancer I. Coexpression assays further indicate that FTF and HNF4α are the most efficient partners for coactivation of the pregenomic core promoter, which may largely account for the hepatic tropism and the early amplification of HBV infection. Carboxy terminus-truncated FTF behaves as a dominant negative mutant to compete all three FTF sites and strongly deactivate core promoter interactions with enhancer I; this suggests possible new ways to interfere with HBV infection.
Simian Immunodeficiency Virus Utilizes Human and Sooty Mangabey but Not Rhesus Macaque STRL33 for Efficient Entry
Authors: Pöhlmann Stefan , Lee Benhur , Meister Silke , Krumbiegel Mandy , Leslie George , Doms Robert W. , Kirchhoff Frank 
Published: July 2002

ABSTRACT
It has been established that many simian immunodeficiency virus (SIV) isolates utilize the orphan receptors GPR15 and STRL33 about as efficiently as the chemokine receptor CCR5 for entry into target cells. Most studies were performed, however, with coreceptors of human origin. We found that SIV from captive rhesus macaques (SIVmac) can utilize both human and simian CCR5 and GPR15 with comparable efficiencies. Strikingly, however, only human STRL33 (huSTRL33), not rhesus macaque STRL33 (rhSTRL33), functioned efficiently as an entry cofactor for a variety of isolates of SIVmac and SIV from sooty mangabeys. A single amino acid substitution of S30R in huSTRL33 impaired coreceptor activity, and the reverse change in rhSTRL33 greatly increased coreceptor activity. In comparison, species-specific sequence variations in N-terminal tyrosines in STRL33 had only moderate effects on SIV entry. These results show that a serine residue located just outside of the cellular membrane in the N terminus of STRL33 is critical for SIV coreceptor function. Interestingly, STRL33 derived from sooty mangabeys, a natural host of SIV, also contained a serine at the corresponding position and was used efficiently as an entry cofactor. These results suggest that STRL33 is not a relevant coreceptor in the SIV/macaque model but may play a role in SIV replication and transmission in naturally infected sooty mangabeys.
Pseudorabies Virus Glycoprotein K Requires the UL20 Gene Product for Processing
Authors: Dietz Petra , Klupp Barbara G. , Fuchs Walter , Köllner Bernd , Weiland Emilie , Mettenleiter Thomas C. 
Published: July 2002

ABSTRACT

Glycoprotein K (gK) of pseudorabies virus (PrV) has recently been identified as a virion component which is dispensable for viral entry but required for direct cell-to-cell spread. Electron microscopic data suggested a possible function of gK in virus egress by preventing immediate fusion of released virus particles with the plasma membrane (B. G. Klupp, J. Baumeister, P. Dietz, H. Granzow, and T. C. Mettenleiter, J. Virol. 72:1949–1958, 1998). For more detailed analysis, a PrV mutant with a deletion of the UL53 (gK) open reading frame (ORF) from codons 48 to 275 was constructed, and the protein was analyzed with two monoclonal antibodies directed against PrV gK. The salient findings of this report are as follows. (i) From the PrV UL53 ORF, a functional gK is translated only from the first in-frame methionine. From the second in-frame methionine, a nonfunctional product is expressed which is not incorporated into virions. (ii) When constitutively expressed in a stable cell line without other viral proteins, gK is only incompletely processed. After superinfection with gK-deletion mutants, proper processing is restored and mature gK is incorporated into virions. (iii) The UL20 gene product is specifically required for processing of gK. gK is not correctly processed in a UL20 deletion mutant of PrV, and superinfection of gK-expressing cells with PrV-UL20
−
does not restore processing. However, all other known structural viral glycoproteins appear to be processed normally in PrV-UL20
−
-infected cells. (iv) Coexpression of gK and UL20 restored gK processing at least partially. Thus, our data show that the UL20 gene product is required for proper processing of PrV gK.
A Metastable Form of the Large Envelope Protein of Duck Hepatitis B Virus: Low-pH Release Results in a Transition to a Hydrophobic, Potentially Fusogenic Conformation
Authors: Grgacic E. V. L. , Schaller H. 
Published: July 2002

ABSTRACTWe have examined the structure and fusion potential of the duck hepatitis B virus (DHBV) envelope proteins by treating subviral particles with deforming agents known to release envelope proteins of viruses from a metastable to a fusion-active state. Exposure of DHBV particles to low pH triggered a major structural change in the large envelope protein (L), resulting in exposure of trypsin sites within its S domain but without affecting the same region in the small surface protein (S) subunits. This conformational change was associated with increased hydrophobicity of the particle surface, most likely arising from surface exposure of the hydrophobic first transmembrane domain (TM1). In the hydrophobic conformation, DHBV particles were able to bind to liposomes and intact cells, while in their absence these particles aggregated, resulting in viral inactivation. These results suggests that some L molecules are in a spring-loaded metastable state which, when released, exposes a previously hidden hydrophobic domain, a transition potentially representing the fusion-active state of the envelope.
Relationship between Human Immunodeficiency Virus Type 1 Gag Multimerization and Membrane Binding
Authors: Ono Akira , Demirov Dimiter , Freed Eric O. 
Published: July 2002

ABSTRACT

The human immunodeficiency virus type 1 (HIV-1) Gag precursor, Pr55
Gag
, is necessary and sufficient for the assembly and release of viruslike particles. Binding of Gag to membrane and Gag multimerization are both essential steps in virus assembly, yet the domains responsible for these events have not been fully defined. In addition, the relationship between membrane binding and Gag-Gag interaction remains to be elucidated. To investigate these issues, we analyzed, in vivo, the membrane-binding and assembly properties of a series of C-terminally truncated Gag mutants. Pr55
Gag
was truncated at the C terminus of matrix (MAstop), between the N- and C-terminal domains of capsid (CA146stop), at the C terminus of capsid (p41stop), at the C terminus of p2 (p43stop), and after the N-terminal 35 amino acids of nucleocapsid (NC35stop). The ability of these truncated Gag molecules to assemble and release viruslike particles and their capacity to copackage into particles when coexpressed with full-length Gag were determined. We demonstrate that the amount of truncated Gag incorporated into particles is incrementally increased by extension from CA146 to NC35, suggesting that multiple sites in this region are involved in Gag multimerization. Using membrane flotation centrifugation, we observe that MA shows significantly reduced membrane binding relative to full-length Gag but that CA146 displays steady-state membrane-binding properties comparable to those of Pr55
Gag
. The finding that the CA146 mutant, which contains only matrix and the N-terminal domain of capsid, exhibits levels of steady-state membrane binding equivalent to those of full-length Gag indicates that strong Gag-Gag interaction domains are not required for the efficient binding of HIV-1 Gag to membrane.
Circular Double-Stranded Forms of TT Virus DNA in the Liver
Authors: Okamoto Hiroaki , Ukita Masato , Nishizawa Tsutomu , Kishimoto Junichi , Hoshi Yuji , Mizuo Hitoshi , Tanaka Takeshi , Miyakawa Yuzo , Mayumi Makoto 
Published: July 2002

ABSTRACT

TT virus
(TTV) is an unenveloped, circular, and single-stranded DNA virus commonly infecting human beings worldwide. TTV DNAs in paired serum and liver tissues from three viremic individuals were separated by gel electrophoresis and characterized biophysically. TTV DNAs in sera migrated in sizes ranging from 2.0 to 2.5 kb. TTV DNAs in liver tissues, however, migrated at 2.0 to 2.5 kb as well as at 3.5 to 6.1 kb. Both faster- and slower-migrating forms of TTV DNAs in the liver were found to be circular and of the full genomic length of 3.8 kb. TTV DNAs migrating at 2.0 to 2.5 kb, from either serum or liver tissues, were sensitive to S1 nuclease but resistant to restriction endonucleases, and therefore, they were single-stranded. By contrast, TTV DNAs in liver tissues that migrated at 3.5 to 6.1 kb were resistant to S1 nuclease. They migrated at 3.7 to 4.0 kb after digestion with
Eco
RI, which suggests that they represent circular, double-stranded replicative intermediates of TTV. When TTV DNAs were subjected to strand-specific primer extension and then amplified by PCR with internal primers, those in serum were found to be minus-stranded DNAs while those in liver tissues were found to be a mixture of plus- and minus-stranded DNAs. These results suggest that TTV replicates in the liver via a circular double-stranded DNA.
Nonstructural Protein 5A of Hepatitis C Virus Inhibits the Function of Karyopherin β3
Authors: Chung Kyung Min , Lee Juhang , Kim Jung-Eun , Song Ok-Kyu , Cho Sungchan , Lim Jeongsim , Seedorf Matthias , Hahm Bumsuk , Jang Sung Key 
Published: July 2002

ABSTRACTIt has been suggested that nonstructural protein 5A (NS5A) of hepatitis C virus (HCV) plays a role in the incapacitation of interferon by inactivation of RNA-dependent protein kinase PKR. In order to further investigate the role of NS5A, we tried to identify cellular proteins interacting with NS5A by using the yeast two-hybrid system. The karyopherin β3 gene was isolated from a human liver cell library as a protein interacting with NS5A. The protein-protein interaction between NS5A and karyopherin β3 was confirmed by in vitro binding assay and an in vivo coimmunoprecipitation method. The effect of NS5A on the karyopherin β3 activity was investigated using a yeast cell line containing mutations in bothPSE1andKAP123, genes that are homologous to the human karyopherin β3 gene. Human karyopherin β3 complemented the loss of thePSE1andKAP123functions, supporting growth of the double mutant cells. However, expression of NS5A hampered the growth of the double mutant cells supplemented with human karyopherin β3. On the other hand, expression of NS5A by itself had no effect on the growth of the double mutant expressing wild-type yeastPSE1. This indicates that NS5A may inhibit karyopherin β3 function via protein-protein interaction. The role of NS5A in HCV replication is discussed.
Expression of Hepatitis B Virus X Protein Does Not Alter the Accumulation of Spontaneous Mutations in Transgenic Mice
Authors: Madden Charles R. , Finegold Milton J. , Slagle Betty L. 
Published: July 2002

ABSTRACT

Chronic infection with hepatitis B virus (HBV) is one of the major etiological factors in the development of human hepatocellular carcinoma. Transgenic mice that express the HBV X protein (HBx) have previously been shown to be more sensitive to the effects of hepatocarcinogens, although the mechanism for this cofactor role remains unknown. The ability of HBx to inhibit DNA repair in transiently transfected cell lines suggests one possible pathway. In the present study, primary hepatocytes isolated from transgenic mice that possess the HBV
X
gene under the control of the human α-1-antitrypsin regulatory region (ATX mice) were found to be deficient in their ability to conduct unscheduled DNA synthesis in response to UV-induced DNA damage. In order to measure the impact of HBx expression on DNA repair in vivo, double-transgenic mice that express HBx and possess a bacteriophage lambda transgene were sacrificed at 30, 90, and 240 days of age. Mutation frequency was determined for high-molecular-weight liver DNA of ATX and control mice by functional analysis of the lambda transgene. Expression of HBx did not significantly increase the accumulation of spontaneous mutations. These results are consistent with previous studies of HBx transgenic mice in which no effect of HBx on liver histology was apparent. This new animal model provides a powerful system in which to investigate the in vivo cooperation between HBx expression and environmental carcinogens.
The
Autographa californica
Nuclear Polyhedrosis Virus
p143
Gene Encodes a DNA Helicase
Authors: McDougal Vivien V. , Guarino Linda A. 
Published: July 2002

ABSTRACT

The P143 protein of
Autographa californica
nuclear polyhedrosis virus is essential for replication of viral DNA. To determine the function of P143, the protein was purified to near homogeneity from recombinant baculovirus-infected cells that overexpress P143. ATPase activity copurified with P143 protein during purification and also during gel filtration at a high salt concentration. The ATPase activity did not require the presence of single-stranded DNA, but was stimulated fourfold by the addition of single-stranded DNA. The ATPase activity of P143 had a

K
m

of 60 μM and a turnover of 4.5 molecules of ATP hydrolyzed/s/molecule of enzyme, indicating moderate affinity for ATP and high catalytic efficiency. P143 unwound a 40-nucleotide primer in an ATP-dependent manner, indicating that the enzyme possesses in vitro DNA helicase activity. Based on this result, it seems likely that P143 functions as a helicase in viral DNA replication.