bluuebunny/crossref_metadata_2025_split huggingface dataset:
Isolation and Characterization of an Arterivirus Defective Interfering RNA Genome
Authors: Molenkamp Richard , Rozier Babette C. D. , Greve Sophie , Spaan Willy J. M. , Snijder Eric J. 
Published: July 2002

ABSTRACT

Equine arteritis virus
(EAV), the type member of the family
Arteriviridae
, is a single-stranded RNA virus with a positive-stranded genome of approximately 13 kb. EAV uses a discontinuous transcription mechanism to produce a nested set of six subgenomic mRNAs from which its structural genes are expressed. We have generated the first documented arterivirus defective interfering (DI) RNAs by serial undiluted passaging of a wild-type EAV stock in BHK-21 cells. A cDNA copy of the smallest DI RNA (5.6 kb) was cloned. Upon transfection into EAV-infected BHK-21 cells, transcripts derived from this clone (pEDI) were replicated and packaged. Sequencing of pEDI revealed that the DI RNA was composed of three segments of the EAV genome (nucleotides 1 to 1057, 1388 to 1684, and 8530 to 12704) which were fused in frame with respect to the replicase reading frame. Remarkably, this DI RNA has retained all of the sequences encoding the structural proteins. By insertion of the chloramphenicol acetyltransferase reporter gene in the DI RNA genome, we were able to delimitate the sequences required for replication/DI-based transcription and packaging of EAV DI RNAs and to reduce the maximal size of a replication-competent EAV DI RNA to approximately 3 kb.
Uncoating Kinetics of Hepatitis A Virus Virions and Provirions
Authors: Bishop Naomi E. , Anderson David A. 
Published: July 2002

ABSTRACT
When the growth kinetics of immature hepatitis A virus provirions and mature virions were monitored, distinct eclipse phases were noted for both types of particles. Strikingly, uncoating of virions occurred around 4 h postinfection, while uncoating of provirions occurred predominantly between 8 and 10 h postinfection. It is proposed that the heterogenous mixture of infectious hepatitis A virus particles (virions and provirions) typically present in inocula is responsible for the normally asynchronous nature of hepatitis A virus uncoating kinetics.
A Murine Leukemia Virus (MuLV) Long Terminal Repeat Derived from Rhesus Macaques in the Context of a Lentivirus Vector and MuLV
gag
Sequence Results in High-Level Gene Expression in Human T Lymphocytes
Authors: Kung Sam K. P. , An Dong Sung , Chen Irvin S. Y. 
Published: July 2002

ABSTRACT

We constructed human immunodeficiency virus type 1 (HIV-1) vectors that will allow higher levels of gene expression in T cells. Gene expression under the control of an internal cytomegalovirus (CMV) immediate-early promoter in a self-inactivating lentiviral vector (CSCG) is 4- to 15-fold lower in T-cell lines (SUPT1 and CEMX174) than in non-lymphoid-cell lines (HeLa and 293T). This is in contrast to a Moloney murine leukemia virus (MoMLV)-based retrovirus vector (SRαLEGFP). We therefore replaced the internal CMV promoter of CSCG with three different murine oncoretroviral long terminal repeat (LTR) promoters—murine sarcoma virus (MSV), MoMLV (MLV), and the LTR (termed Rh-MLV) that is derived from the ampho-mink cell focus-forming (AMP/MCF) retrovirus in the serum of one rhesus macaque monkey that developed T-cell lymphoma following autologous transplantation of enriched bone marrow stem cells transduced with a retrovirus vector preparation containing replication-competent viruses (E. F. Vanin, M. Kaloss, C. Broscius, and A. W. Nienhuis, J. Virol. 68:4241–4250, 1994). We found that the combination of Rh-MLV LTR and a partial
gag
sequence of MoMLV (Δ
gag
871–1612
) in CS-Rh-MLV-E gave the highest level of enhanced green fluorescent protein (EGFP) gene expression compared with MLV, MSV LTR, phosphoglycerate kinase, and CMV promoters in T-cell lines, as well as activated primary T cells. Interestingly, there was a further two- to threefold increase in EGFP expression (thus, 10-fold-higher expression than with CMV) when the Rh-MLV promoter and Δ
gag
871–1612
were used in a self-inactivating-vector setting that has a further deletion in the U3 region of the HIV-1 LTR. These hybrid vectors should prove useful in gene therapy applications for T cells.
Simian Immunodeficiency Virus Containing Mutations in N-Terminal Tyrosine Residues and in the PxxP Motif in Nef Replicates Efficiently in Rhesus Macaques
Authors: Carl Silke , Iafrate A. John , Lang Sabine M. , Stolte Nicole , Stahl-Hennig Christiane , Mätz-Rensing Kerstin , Fuchs Dietmar , Skowronski Jacek , Kirchhoff Frank 
Published: July 2002

ABSTRACTSIVmac Nef contains two N-terminal tyrosines that were proposed to be part of an SH2-ligand domain and/or a tyrosine-based endocytosis signal and a putative SH3-ligand domain (P104xxP107). In the present study, we investigated the effects of combined mutations in these tyrosine and proline residues on simian immunodeficiency virus (SIV) Nef interactions with the cellular signal transduction and endocytic machinery. We found that mutation of Y28F, Y39F, P104A, and P107A (FFAA-Nef) had little effect on Nef functions such as the association with the cellular tyrosine kinase Src, downregulation of cell surface expression of CD4 and class I major histocompatibility complex, and enhancement of virion infectivity. However, mutations in the PxxP sequence reduced the ability of Nef to stimulate viral replication in primary lymphocytes. Three macaques infected with the SIVmac239 FFAA-Nef variant showed high viral loads during the acute phase of infection. Reversions in the mutated prolines were observed between 12 and 20 weeks postinfection. Importantly, reversion of A107→P, which restored the ability of Nef to coprecipitate a 62-kDa phosphoprotein in in vitro kinase assays, did not precede the development of a high viral load. The Y28/Y39→F28/F39substitutions did not revert. In conclusion, mutations in both the tyrosine residues and the putative SH3 ligand domain apparently do not disrupt major aspects of SIV Nef function in vivo.
Genetic Evidence of a Role for ATM in Functional Interaction between Human T-Cell Leukemia Virus Type 1 Tax and p53
Authors: Van Phillip L. , Yim Kyong-Wook , Jin Dong-Yan , Dapolito George , Kurimasa Akihiro , Jeang Kuan-Teh 
Published: July 2002

ABSTRACT
Recent evidence from several investigators suggest that the human T-cell leukemia virus type 1 Tax oncoprotein represses the transcriptional activity of the tumor suppressor protein, p53. An examination of published findings reveals serious controversy as to the mechanism(s) utilized by Tax to inhibit p53 activity and whether the same mechanism is used by Tax in adherent and suspension cells. Here, we have investigated Tax-p53 interaction simultaneously in adherent epithelial (HeLa and Saos) and suspension T-lymphocyte (Jurkat) cells. Our results indicate that Tax activity through the CREB/CREB-binding protein (CBP), but not NF-κB, pathway is needed to repress the transcriptional activity of p53 in all tested cell lines. However, we did find that while CBP binding by Tax is necessary, it is not sufficient for inhibiting p53 function. Based on knockout cell studies, we correlated a strong genetic requirement for the ATM, but not protein kinase-dependent DNA, protein in conferring a Tax-p53-repressive phenotype.
Transport of Human Immunodeficiency Virus Type 1 Pseudoviruses across the Blood-Brain Barrier: Role of Envelope Proteins and Adsorptive Endocytosis
Authors: Banks William A. , Freed Eric O. , Wolf Kathleen M. , Robinson Sandra M. , Franko Mark , Kumar Vijaya B. 
Published: July 2002

ABSTRACTBlood-borne human immunodeficiency virus type 1 (HIV-1) crosses the blood-brain barrier (BBB) to induce brain dysfunction. How HIV-1 crosses the BBB is unclear. Most work has focused on the ability of infected immune cells to cross the BBB, with less attention devoted to the study of free virus. Since the HIV-1 coat glycoprotein gp120 can cross the BBB, we postulated that gp120 might be key in determining whether free virus can cross the BBB. We used radioactive virions which do (Env+) or do not (Env−) bear the envelope proteins to characterize the ability of HIV-1 to be taken up by the murine BBB. In vivo and in vitro studies showed that the envelope proteins are key to the uptake of free virus and that uptake was enhanced by wheat germ agglutinin, strongly suggesting that the envelope proteins induce viral adsorptive endocytosis and transcytosis in brain endothelia. Capillary depletion showed that Env+virus completely crossed the vascular BBB to enter the parenchyma of the brain. Virus also entered the cerebrospinal fluid, suggesting passage across the choroid plexus as well. About 0.22% of the intravenously injected dose was taken up per g of brain. In vitro studies showed that postinternalization membrane cohesion (membrane binding not reversed with acid wash or cell lysis) was a regulated event. Intact virus was recovered from the brain endothelial cytosol and was effluxed from the endothelial cells. These results show that free HIV-1 can cross the BBB by an event related to adsorptive endocytosis and mediated by the envelope proteins.
Requirement of Interaction of Nectin-1α/HveC with Afadin for Efficient Cell-Cell Spread of Herpes Simplex Virus Type 1
Authors: Sakisaka Toshiaki , Taniguchi Tomokuni , Nakanishi Hiroyuki , Takahashi Kenichi , Miyahara Masako , Ikeda Wataru , Yokoyama Shigekazu , Peng Ying-Feng , Yamanishi Koichi , Takai Yoshimi 
Published: July 2002

ABSTRACT

We recently found a novel cell-cell adhesion system at cadherin-based adherens junctions (AJs), consisting at least of nectin, a Ca
2+
-independent homophilic immunoglobulin-like adhesion molecule, and afadin, an actin filament-binding protein that connects nectin to the actin cytoskeleton. Nectin is associated with cadherin through afadin and α-catenin. The cadherin-catenin system increases the concentration of nectin at AJs in an afadin-dependent manner. Nectin constitutes a family consisting of three members: nectin-1, -2, and -3. Nectin-1 serves as an entry and cell-cell spread mediator of herpes simplex virus type 1 (HSV-1). We studied here a role of the interaction of nectin-1α with afadin in entry and/or cell-cell spread of HSV-1. By the use of cadherin-deficient L cells overexpressing the full length of nectin-1α capable of interacting with afadin and L cells overexpressing a truncated form of nectin-1α incapable of interacting with afadin, we found that the interaction of nectin-1α with afadin increased the efficiency of cell-cell spread, but not entry, of HSV-1. This interaction did not affect the binding to nectin-1α of glycoprotein D, a viral component mediating entry of HSV-1 into host cells. Furthermore, the cadherin-catenin system increased the efficiency of cell-cell spread of HSV-1, although it also increased the efficiency of entry of HSV-1. It is likely that efficient cell-cell spread of HSV-1 is caused by afadin-dependent concentrated localization of nectin-1α at cadherin-based AJs.
Genotypic Correlates of Phenotypic Resistance to Efavirenz in Virus Isolates from Patients Failing Nonnucleoside Reverse Transcriptase Inhibitor Therapy
Authors: Bacheler Lee , Jeffrey Susan , Hanna George , D'Aquila Richard , Wallace Lany , Logue Kelly , Cordova Beverly , Hertogs Kurt , Larder Brendan , Buckery Renay , Baker David , Gallagher Karen , Scarnati Helen , Tritch Radonna , Rizzo Chris 
Published: July 2002

ABSTRACT
Efavirenz (also known as DMP 266 or SUSTIVA) is a potent nonnucleoside inhibitor of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) activity and of HIV-1 replication in vitro and in vivo. Most patients on efavirenz-containing regimens have sustained antiviral responses; however, rebounds in plasma viral load have been observed in some patients in association with the emergence of mutant strains of HIV-1. Virus isolates from the peripheral blood mononuclear cells (PBMCs) of patients with such treatment failures, as well as recombinant viruses incorporating viral sequences derived from patient plasma, show reduced in vitro susceptibility to efavirenz in association with mutations in the RT gene encoding K103N, Y188L, or G190S/E substitutions. Patterns of RT gene mutations and in vitro susceptibility were similar in plasma virus and in viruses isolated from PBMCs. Variant strains of HIV-1 constructed by site-directed mutagenesis confirmed the role of K103N, G190S, and Y188L substitutions in reduced susceptibility to efavirenz. Further, certain secondary mutations (V106I, V108I, Y181C, Y188H, P225H, and F227L) conferred little resistance to efavirenz as single mutations but enhanced the level of resistance of viruses carrying these mutations in combination with K103N or Y188L. Viruses with K103N or Y188L mutations, regardless of the initial selecting nonnucleoside RT inhibitor (NNRTI), exhibited cross-resistance to all of the presently available NNRTIs (efavirenz, nevirapine, and delavirdine). Some virus isolates from nevirapine or delavirdine treatment failures that lacked K103N or Y188L mutations remained susceptible to efavirenz in vitro, although the clinical significance of this finding is presently unclear.
The NH
2
-Terminal Domain of the Human T-Cell Leukemia Virus Type 1 Capsid Protein Is Involved in Particle Formation
Authors: Rayne Fabienne , Bouamr Fadila , Lalanne Jacqueline , Mamoun Robert Z. 
Published: July 2002

ABSTRACT
The human immunodeficiency virus type 1 (HIV-1) and human T-cell leukemia virus type 1 (HTLV-1) capsid proteins (CA) display similar structures formed by two independently folded N-terminal (NTD) and C-terminal (CTD) domains. To characterize the functions harbored by the HTLV-1 CA domains in particle formation, 12 sites scattered throughout the protein were mutated. The effects of the mutations on Gag membrane binding, proteolytic processing, and virus-like particle secretion were analyzed. It appears that the NTD is the major partner of indirect or direct Gag-Gag interactions. In particular, most of the NTD mutations impaired virion morphogenesis, and no mutation located in the NTD could be fully rescued by coexpression of wild-type Gag. In contrast, the CTD seems not to be involved in Gag-Gag interactions. Nevertheless, an unknown function required for particle formation is located in the CTD. Thus, despite an overall structural similarity between the HIV-1 and HTLV-1 CA proteins, their NTDs and CTDs exhibit different functions.
Nasal-Associated Lymphoid Tissue Is a Site of Long-Term Virus-Specific Antibody Production following Respiratory Virus Infection of Mice
Authors: Liang Bin , Hyland Lisa , Hou Sam 
Published: July 2002

ABSTRACT
Nasal immunoglobulin A provides an initial defense against inhaled respiratory pathogens. However, it is not known whether the nasal-associated lymphoid tissues (NALT) are able to mount an effective long-lasting pathogen-specific immune response, nor is it known whether functional differences exist between the organized NALT (O-NALT) and the diffuse NALT lining the nasal passages (D-NALT). Here we show that although both the O-NALT and the D-NALT are capable of producing virus-specific antibody in response to influenza virus infection, the frequency of specific antibody-forming cells in the D-NALT is much greater than the frequency observed in the O-NALT. Furthermore, we show that the D-NALT but not the O-NALT is the site of long-term virus-specific humoral immunity which lasts for the life of the animal. These results indicate that the D-NALT is not only the major effector site of the NALT but also the site of local long-term specific antibody production.
Characterization of Nuclear RNases That Cleave Hepatitis B Virus RNA near the La Protein Binding Site
Authors: Heise Tilman , Guidotti Luca G. , Chisari Francis V. 
Published: July 2002

ABSTRACTHepatitis B virus (HBV) RNA is downregulated by inflammatory cytokines induced in the liver by adoptively transferred HBV-specific cytotoxic T lymphocytes (CTLs) and during murine cytomegalovirus (MCMV) infections of the livers of HBV transgenic mice. The disappearance of HBV RNA is tightly associated with the cytokine-induced proteolytic cleavage of a previously defined HBV RNA-binding protein known as La autoantigen. La binds to a predicted stem-loop structure at the 5′ end of the posttranscriptional regulatory element of HBV RNA between nucleotides 1243 and 1333. In the present study, we searched for nuclear RNase activities that might be involved in HBV RNA decay. Nuclear extracts derived from control livers and CTL-injected and MCMV-infected livers were analyzed for the ability to cleave HBV RNA. Endonucleolytic activity that cleaved HBV RNA at positions 1269 to 1270 and 1271 to 1272, immediately 5′ of the stem-loop bound by the La protein (positions 1272 to 1293), was detected. Furthermore, we provide evidence that the cytokine-dependent downregulation of HBV RNA following MCMV infection is temporally associated with the upregulation of the endonucleolytic activity herein described. Collectively, these results suggest a model in which the steady-state HBV RNA content is controlled by the stabilizing influence of La and the destabilizing influence of nuclear RNase activities.
Endoplasmic Reticulum andcis-Golgi Localization of Human T-Lymphotropic Virus Type 1 p12I: Association with Calreticulin and Calnexin
Authors: Ding Wei , Albrecht Björn , Luo Rushu , Zhang Weiqing , Stanley James R. L. , Newbound Garret C. , Lairmore Michael D. 
Published: July 2002

ABSTRACTHuman T-lymphotropic virus type 1 (HTLV-1) is a complex retrovirus encoding regulatory and accessory genes in four open reading frames (ORF I to IV) of the pX region. We have demonstrated an important role of pX ORF I expression, which encodes p12I, in establishment of HTLV-1 infection in a rabbit model and for optimal viral infectivity in quiescent primary lymphocytes. These data indicated that p12Imay enhance lymphocyte activation and thereby promote virus infection. To further define the role of p12Iin cell activation, we characterized the subcellular localization of p12Iin transfected 293T cells and HeLa-Tat cells by multiple methods, including immunofluorescence confocal microscopy, electron microscopy, and subcellular fractionation. Herein, we demonstrate that p12Iaccumulates in the endoplasmic reticulum (ER) andcis-Golgi apparatus. The location of p12Iwas unchanged following treatments with both cycloheximide (blocking de novo protein synthesis) and brefeldin A (disrupting ER-to-Golgi protein transport), indicating that the protein is retained in the ER andcis-Golgi. Moreover, using coimmunoprecipitation assays, we identify the direct binding of p12Iwith both calreticulin and calnexin, resident ER proteins which regulate calcium storage. Our results indicate that p12Idirectly binds key regulatory proteins involved in calcium-mediated cell signaling and suggest a role of p12Iin the establishment of HTLV-1 infection by activation of host cells.
Identification of a Putative Coreceptor on Vero Cells That Participates in Dengue 4 Virus Infection
Authors: Martı́nez-Barragán José de Jesús , del Angel Rosa M. 
Published: July 2002

ABSTRACTDengue virus infects target cells by attaching to a cell surface receptor through the envelope (E) glycoprotein, located on the surface of the viral membrane. On Vero and BHK cells, heparan sulfate (HS) moieties of proteoglycans are the receptors for dengue virus; however, additional proteins have also been described as putative dengue virus receptors on C6/36, HL60, and BM cells. HS can also act as a receptor for other types of viruses or as an attachment molecule for viruses that require additional host cell molecules to allow viral penetration. In this study we searched for molecules other than HS that could participate in dengue virus infection of Vero cells. Labeled dengue 4 virus bound with high affinity to two molecules of 74 and 44 kDa. Binding of dengue virus to the 74-kDa molecule was susceptible to protease and sodium periodate treatment and resistant to heparinase treatments. Lectins such as concanavalin A and wheat germ agglutinin prevented dengue virus binding to both the 74- and the 44-kDa protein in overlay assays, while phytohemagglutinin P did not affect binding, suggesting that carbohydrate residues (α-mannose orN-acetylglucosamine) are important in virus binding to host cells. Protease susceptibility, biotin labeling, and immunofluorescence with a polyclonal antibody raised against the 74-kDa protein consistently identified the protein on the surfaces of Vero cells. Moreover, the antibody against the 74-kDa protein was able to inhibit dengue virus infection. These data suggest that HS might serve as a primary receptor, probably concentrating virus particles on the surfaces of Vero cells, and then other molecules, such as the 74-kDa protein, might participate as coreceptors in viral penetration. The 74-kDa protein possibly constitutes part of a putative receptor complex for dengue virus infection of Vero cells.
Chimeric Nectin1-Poliovirus Receptor Molecules Identify a Nectin1 Region Functional in Herpes Simplex Virus Entry
Authors: Cocchi Francesca , Lopez Marc , Dubreuil Patrice , Campadelli Fiume Gabriella , Menotti Laura 
Published: July 2002

ABSTRACT
Human nectin1 (hNectin1), an adhesion molecule belonging to the nectin family of the immunoglobulin superfamily, mediates entry of herpes simplex virus (HSV) into cells. The hNectin1 domain that mediates virus entry into cells and also binds glycoprotein D (gD) has been localized to the first N-terminal V-type domain. The poliovirus receptor (PVR) is a structural homolog to nectins, but it cannot function as an HSV entry receptor. hNectin1-PVR chimeras were constructed to functionally locate the site on hNectin1 involved in HSV entry (HSV entry site). The epitope recognized by monoclonal antibody (MAb) R1.302, which is able to block HSV entry, was also located. The chimeric receptors were designed to preserve the overall structure of the V domain. The HSV entry activity mapped entirely to the hNectin1 portion located between residues 64 and 94 (64-94), likely to encode the C, C′, and C" β-strands and intervening loops. In turn, this site consisted of two portions: one with low-level basal activity for HSV entry (77-94), and one immediately upstream (residues 64 to 76) which greatly enhanced the HSV entry activity of the downstream region. The gD-binding site mapped substantially to the same site, whereas the MAb R1.302 epitope also required a further downstream portion (95-102). The involvement of the 64-76 portion is at difference with previous indirect mapping results that were based on competitive binding studies (C. Krummenacher et al., J. Virol. 74:10863–10872, 2000). The A, A′, B, D, E, F, and G β-strands and intervening loops did not appear to play any role in HSV entry. According to the predicted three-dimensional structure of PVR, the C C′ C" site is located peripherally in the V domain and very likely represents an accessible portion at the cell surface.