bluuebunny/crossref_metadata_2025_split huggingface dataset:
Monoclonal Antibody 667 Recognizes the Variable Region A Motif of the Ecotropic Retrovirus CasBrE Envelope Glycoprotein and Inhibits Env Binding to the Viral Receptor
Authors: Dreja Hanna , Gros Laurent , Villard Sylvie , Bachrach Estanislao , Oates Anna , Granier Claude , Chardes Thierry , Mani Jean-Claude , Piechaczyk Marc , Pelegrin Mireia 
Published: September 2003

ABSTRACTMonoclonal antibody (MAb) 667 is a neutralizing mouse monoclonal antibody recognizing the envelope glycoprotein (Env) of the ecotropic neurotropic murine retrovirus CasBrE but not that of other murine retroviruses. Since 667 can be used for preclinical studies of antiviral gene therapy as well as for studying the early events of retroviral infection, we have cloned its cDNAs and molecularly characterized it in detail. Spot technique-based experiments showed that 667 recognizes a linear epitope of 12 amino acids located in the variable region A of the receptor binding domain. Alanine scanning experiments showed that six amino acids within the epitope are critical for MAb binding. One of them, D57, is not present in any other murine retroviral Env, which suggests a critical role for this residue in the selectivity of 667. MAb 667 heavy- and light-chain cDNAs were functionally characterized by transient transfection into Cos-7 cells. Enzyme-linked immunosorbent assays and Biacore studies showed that the specificities as well as the antigen-binding thermodynamic and kinetic properties of the recombinant 667 MAb (r667) produced by Cos-7 cells and those of the parental hybridoma-produced MAb (h667) were similar. However, h667 was shown to contain contaminating retroviral and/or retrovirus-like particles which interfere with both viral binding and neutralization experiments. These contaminants could successfully be removed by a stringent purification protocol. Importantly, this purified 667 could completely prevent retrovirus binding to target cells and was as efficient as the r667 MAb produced by transfected Cos-7 cells in neutralization assays. In conclusion, this study shows that the primary mechanism of virus neutralization by MAb 667 is the blocking of the retroviral receptor binding domain of CasBrE Env. In addition, the findings of this study constitute a warning against the direct use of hybridoma cell culture supernatants for studying the initial events of retroviral cell infection as well as for carrying out in vivo neutralization experiments and suggest that either recombinant antibodies or highly purified antibodies are preferable for these purposes.
Serial Recombination during Circulation of Type 1 Wild-Vaccine Recombinant Polioviruses in China
Authors: Liu Hong-Mei , Zheng Du-Ping , Zhang Li-Bi , Oberste M. Steven , Kew Olen M. , Pallansch Mark A. 
Published: September 2003

ABSTRACT
Type 1 wild-vaccine recombinant polioviruses sharing a 367-nucleotide (nt) block of Sabin 1-derived sequence spanning the VP1 and 2A genes circulated widely in China from 1991 to 1993. We surveyed the sequence relationships among 34 wild-vaccine recombinants by comparing six genomic intervals: the conserved 5′-untranslated region (5′-UTR) (nt 186 to 639), the hypervariable portion of the 5′-UTR (nt 640 to 742), the VP4 and partial VP2 genes (nt 743 to 1176), the VP1 gene (nt 2480 to 3385), the 2A gene (nt 3386 to 3832), and the partial 3D gene (nt 6011 to 6544). The 5′-UTR, capsid (VP4-VP2 and VP1), and 2A sequence intervals had similar phylogenies. By contrast, the partial 3D sequences could be distributed into five divergent genetic classes. Most (25 of 34) of the wild-vaccine recombinant isolates showed no evidence of additional recombination beyond the initial wild-Sabin recombination event. Eight isolates from 1992 to 1993, however, appear to be derived from three independent additional recombination events, and one 1993 isolate was derived from two consecutive events. Complete genomic sequences of a representative isolate for each 3D sequence class demonstrated that these exchanges had occurred in the 2B, 2C, and 3D genes. The 3D gene sequences were not closely related to those of the Sabin strains or 53 diverse contemporary wild poliovirus isolates from China, but all were related to the 3D genes of species C enteroviruses. The appearance within approximately 2.5 years of five recombinant classes derived from a single ancestral infection illustrates the rapid emergence of new recombinants among circulating wild polioviruses.
Strategic Attack on Host Cell Gene Expression during Adenovirus Infection
Authors: Zhao Hongxing , Granberg Fredrik , Elfineh Ludmila , Pettersson Ulf , Svensson Catharina 
Published: September 2003

ABSTRACT
To understand the interaction between the virus and its host, we used three sources of cDNA microarrays to examine the expression of 12,309 unique genes at 6 h postinfection of HeLa cells with high multiplicities of adenovirus type 2. Seventy-six genes with significantly changed expression ratios were identified, suggesting that adenovirus only modulates expression of a limited set of cellular genes. Quantitative real-time PCR analyses on selected genes were performed to confirm the microarray results. Significantly, a pronounced transcriptional activation by the promiscuous E1A-289R transcriptional activator was not apparent. Instead, promoter sequences in 45% of the upregulated genes harbored a potential E2F binding site, suggesting that the ability of the amino-terminal domain of E1A to regulate E2F-dependent transcription may be a major pathway for regulation of cellular gene expression. CDC25A was the only upregulated gene directly involved in cell cycle control. In contrast, several genes implicated in cell growth arrest were repressed. The transforming growth factor beta superfamily was specifically affected in the expression of both the upstream ligand and an intracellular regulator. In agreement with previous reports, adenovirus also targeted the innate immune response by downregulating several cytokines, including CLL2, CXCL1, and interleukin-6. Finally, stress response genes encoding GADD45B, ATF3, and TP53AP1 were upregulated. Importantly, we also found a novel countermeasure—activation of the apoptosis inhibitor survivin.
Tomato Mosaic Virus Replication Protein Suppresses Virus-Targeted Posttranscriptional Gene Silencing
Authors: Kubota Kenji , Tsuda Shinya , Tamai Atsushi , Meshi Tetsuo 
Published: September 2003

ABSTRACT

Posttranscriptional gene silencing (PTGS), a homology-dependent RNA degradation system, has a role in defending against virus infection in plants, but plant viruses encode a suppressor to combat PTGS. Using transgenic tobacco in which the expression of green fluorescent protein (GFP) is posttranscriptionally silenced, we investigated a tomato mosaic virus (ToMV)-encoded PTGS suppressor. Infection with wild-type ToMV (L strain) interrupted GFP silencing in tobacco, coincident with visible symptoms, whereas some attenuated strains of ToMV (L
11
and L
11
A strains) failed to suppress GFP silencing. Analyses of recombinant viruses containing the L and L
11
A strains revealed that a single base change in the replicase gene, which causes an amino acid substitution, is responsible for the symptomless and suppressor-defective phenotypes of the attenuated strains. An agroinfiltration assay indicated that the 130K replication protein acts as a PTGS suppressor. Small interfering RNAs (siRNAs) of 21 to 25 nucleotides accumulated during ToMV infection, suggesting that the major target of the ToMV-encoded suppressor is downstream from the production of siRNAs in the PTGS pathway. Analysis with GFP-tagged recombinant viruses revealed that the suppressor inhibits the establishment of the ToMV-targeted PTGS system in the inoculated leaves but does not detectably suppress the activity of the preexisting, sequence-specific PTGS machinery there. Taken together, these results indicate that it is likely that the ToMV-encoded suppressor, the 130K replication protein, blocks the utilization of silencing-associated small RNAs, so that a homology-dependent RNA degradation machinery is not newly formed.
Human T-Cell Lymphotropic Virus Type 1 p12
I
Enhances Interleukin-2 Production during T-Cell Activation
Authors: Ding Wei , Kim Seung-Jae , Nair Amrithraj M. , Michael Bindhu , Boris-Lawrie Kathleen , Tripp Adam , Feuer Gerold , Lairmore Michael D. 
Published: September 2003

ABSTRACT

Human T-cell lymphotropic virus type 1 (HTLV-1) causes adult T-cell leukemia/lymphoma (ATLL) and a variety of lymphoproliferative disorders. The early virus-cell interactions that determine a productive infection remain unclear. However, it is well recognized that T-cell activation is required for effective retroviral integration into the host cell genome and subsequent viral replication. The HTLV-1 pX open reading frame I encoding protein, p12
I
, is critical for the virus to establish persistent infection in vivo and for infection in quiescent primary lymphocytes in vitro. p12
I
localizes in the endoplasmic reticulum (ER) and cis-Golgi apparatus, increases intracellular calcium and activates nuclear factor of activated T cells (NFAT)-mediated transcription. To clarify the function of p12
I
, we tested the production of IL-2 from Jurkat T cells and peripheral blood mononuclear cells (PBMC) expressing p12
I
. Lentiviral vector expressed p12
I
in Jurkat T cells enhanced interleukin-2 (IL-2) production in a calcium pathway-dependent manner during T-cell receptor (TCR) stimulation. Expression of p12
I
also induced higher NFAT-mediated reporter gene activities during TCR stimulation in Jurkat T cells. In contrast, p12 expression in PBMC elicited increased IL-2 production in the presence of phorbal ester stimulation, but not during TCR stimulation. Finally, the requirement of ER localization for p12
I
-mediated NFAT activation was demonstrated and two positive regions and two negative regions in p12
I
were identified for the activation of this transcription factor by using p12
I
truncation mutants. These results are the first to indicate that HTLV-1, an etiologic agent associated with lymphoproliferative diseases, uses a conserved accessory protein to induce T-cell activation, an antecedent to efficient viral infection.
Interacting Domains of the HN and F Proteins of Newcastle Disease Virus
Authors: Gravel Kathryn A. , Morrison Trudy G. 
Published: September 2003

ABSTRACT

The activation of most paramyxovirus fusion proteins (F proteins) requires not only cleavage of F
0
to F
1
and F
2
but also coexpression of the homologous attachment protein, hemagglutinin-neuraminidase (HN) or hemagglutinin (H). The type specificity requirement for HN or H protein coexpression strongly suggests that an interaction between HN and F proteins is required for fusion, and studies of chimeric HN proteins have implicated the membrane-proximal ectodomain in this interaction. Using biotin-labeled peptides with sequences of the Newcastle disease virus (NDV) F protein heptad repeat 2 (HR2) domain, we detected a specific interaction with amino acids 124 to 152 from the NDV HN protein. Biotin-labeled HR2 peptides bound to glutathione
S
-transferase (GST) fusion proteins containing these HN protein sequences but not to GST or to GST containing HN protein sequences corresponding to amino acids 49 to 118. To verify the functional significance of the interaction, two point mutations in the HN protein gene, I133L and L140A, were made individually by site-specific mutagenesis to produce two mutant proteins. These mutations inhibited the fusion promotion activities of the proteins without significantly affecting their surface expression, attachment activities, or neuraminidase activities. Furthermore, these changes in the sequence of amino acids 124 to 152 in the GST-HN fusion protein that bound HR2 peptides affected the binding of the peptides. These results are consistent with the hypothesis that HN protein binds to the F protein HR2 domain, an interaction important for the fusion promotion activity of the HN protein.
Replication of Chimeric Human Immunodeficiency Virus Type 1 (HIV-1) Containing HIV-2 Integrase (IN): Naturally Selected Mutations in IN Augment DNA Synthesis
Authors: Padow Marcus , Lai Lilin , Deivanayagam Champion , DeLucas Lawrence J. , Weiss Robert B. , Dunn Diane M. , Wu Xiaoyun , Kappes John C. 
Published: September 2003

ABSTRACT

The human immunodeficiency virus type 1 (HIV-1) integrase (IN) protein augments the initiation of reverse transcription. Chimeric HIV-1 containing HIV-2
IN
(SG3
IN2
) is severely impaired in virus infectivity and DNA synthesis. To analyze the nature of this defect, we infected T cells with the chimeric SG3
IN2
virus and by continuous passage in cell culture selected for virus with improved replication properties. Viruses from two different time points were chosen for further analysis, an early culture-adapted virus (CF-65) that exhibited an intermediate level of infectivity, and a later-passaged virus (CF-131) that was significantly more infectious. Sequence analysis of multiple clones derived from the CF-65 virus culture demonstrated a diversity of mutations in the reverse transcriptase (RT) and a common V204I IN mutation. Analysis of clones derived from the CF-131 virus indicated the selection of two additional IN mutations, Q96H and K127E, and a fixed V179I RT mutation. By cloning
RT
and/or
IN
sequences back into the original SG3
IN2
chimeric virus, we demonstrated that mutations in both RT and IN contributed to the improvement in viral fitness. The effect of the HIV-2IN(IN
2
) mutations on virus DNA synthesis was analyzed by packaging IN
2
mutants into HIV-1 as Vpr-IN
2
fusion proteins. This analysis revealed that the Q96H, K127E and V204I mutations increased the infectivity of the chimeric virus by augmenting the initiation of viral cDNA synthesis in infected cells. The Q96H and K127E mutations are present in adjacent helical structures on the surface of the IN protein and together account for most of the increase observed in DNA synthesis. Our findings provide evidence that the IN protein augments the initiation of reverse transcription through specific interactions with other viral components comprising the initiation complex. Moreover, they implicate specific regions on the surface of IN that may help to elucidate mechanisms by which the HIV-1 IN protein augments the initiation of HIV-1 reverse transcription in vivo.
Junonia coenia
Densovirus-Based Vectors for Stable Transgene Expression in Sf9 Cells: Influence of the Densovirus Sequences on Genomic Integration
Authors: Bossin Hervé , Fournier Philippe , Royer Corinne , Barry Patrick , Cérutti Pierre , Gimenez Sylvie , Couble Pierre , Bergoin Max 
Published: September 2003

ABSTRACT

The invertebrate parvovirus
Junonia coenia
densovirus (JcDNV) shares similarities with terminal hairpins and nonstructural (NS) protein activities of adeno-associated virus (AAV) despite their evolutionary divergence (B. Dumas, M. Jourdan, A. M. Pascaud, and M. Bergoin, Virology, 191:202-222, 1992, and C. Ding, M. Urabe, M. Bergoin, and R. M. Kotin, J. Virol. 76:338-345, 2002). We demonstrate here that persistent transgene expression in insect cells results from stable integration of transfected JcDNV-derived vectors into the host genome. To assess the integrative properties of JcDNV vectors, the green fluorescent protein (GFP)
gfp
marker gene was fused in frame into the major open reading frame (ORF1) of the viral sequence under the control of the P9 capsid protein promoter. In addition, the influence of the nonstructural proteins on the posttransfection maintenance of the vectors was examined by interruption of one or all three NS ORFs. Following transfection of Sf9 cells with each of the JcDNV constructs, clones showing persistent GFP expression were isolated. Structural analyses revealed that the majority of the JcDNV plasmid sequence was integrated into the genome of the fluorescent clones. Integration was observed whether or not NS proteins were expressed. However, the presence of NS genes in the constructs greatly influenced the number of integrated copies and their distribution in the host genome. Disruption of NS genes expression resulted in integration of head-to-tail concatemers at multiple sites within the genome. Further analyses demonstrated that the
cis
JcDNV 5′ inverted terminal repeat region was the primary site of recombination. Sequence analyses of integration junctions showed rearrangements of both flanking and internal sequences for most integrations. These findings demonstrate that JcDNV vectors integrate into insect cells in a manner similar to AAV plasmids in mammalian cells.
Identification of a Heparin-Binding Motif on Adeno-Associated Virus Type 2 Capsids
Authors: Kern A. , Schmidt K. , Leder C. , Müller O. J. , Wobus C. E. , Bettinger K. , Von der Lieth C. W. , King J. A. , Kleinschmidt J. A. 
Published: September 2003

ABSTRACT
Infection of cells with adeno-associated virus (AAV) type 2 (AAV-2) is mediated by binding to heparan sulfate proteoglycan and can be competed by heparin. Mutational analysis of AAV-2 capsid proteins showed that a group of basic amino acids (arginines 484, 487, 585, and 588 and lysine 532) contribute to heparin and HeLa cell binding. These amino acids are positioned in three clusters at the threefold spike region of the AAV-2 capsid. According to the recently resolved atomic structure for AAV-2, arginines 484 and 487 and lysine 532 on one site and arginines 585 and 588 on the other site belong to different capsid protein subunits. These data suggest that the formation of the heparin-binding motifs depends on the correct assembly of VP trimers or even of capsids. In contrast, arginine 475, which also strongly reduces heparin binding as well as viral infectivity upon mutation to alanine, is located inside the capsid structure at the border of adjacent VP subunits and most likely influences heparin binding indirectly by disturbing correct subunit assembly. Computer simulation of heparin docking to the AAV-2 capsid suggests that heparin associates with the three basic clusters along a channel-like cavity flanked by the basic amino acids. With few exceptions, mutant infectivities correlated with their heparin- and cell-binding properties. The tissue distribution in mice of recombinant AAV-2 mutated in R484 and R585 indicated markedly reduced infection of the liver, compared to infection with wild-type recombinant AAV, but continued infection of the heart. These results suggest that although heparin binding influences the infectivity of AAV-2, it seems not to be necessary.
Bioluminescence Imaging Reveals Systemic Dissemination of Herpes Simplex Virus Type 1 in the Absence of Interferon Receptors
Authors: Luker Gary D. , Prior Julie L. , Song Jiling , Pica Christina M. , Leib David A. 
Published: September 2003

ABSTRACT
Herpes simplex virus type 1 (HSV-1) can produce disseminated, systemic infection in neonates and patients with AIDS or other immunocompromising diseases, resulting in significant morbidity and mortality in spite of antiviral therapy. Components of host immunity that normally limit HSV-1 to localized epithelial and neuronal infection remain incompletely defined. We used in vivo bioluminescence imaging to determine effects of type I and II interferons (IFNs) on replication and tropism of HSV-1 infection in mice with genetic deficiency of type I, type II, or both type I and II IFN receptors. Following footpad or ocular infection of mice lacking type I IFN receptors, HSV-1 spread to parenchymal organs, including lung, liver, spleen, and regional lymph nodes, but mice survived. Deletion of type I and II IFN receptors produced quantitatively greatest and most widespread dissemination of virus to visceral organs and the nervous system, and these mice invariably died after ocular or footpad infection. Type II receptor knockout and wild-type mice had comparable viral replication and localization, with no systemic spread of HSV-1 or lethality. Therefore, while isolated deficiency of type II IFN receptors did not affect pathogenesis, loss of these receptors in combination with genetic deletion of type I receptors had a profound effect on susceptibility to HSV-1. These data demonstrate different effects of type I and II IFNs in limiting systemic dissemination of HSV-1 and further validate the use of bioluminescence imaging for studies of viral pathogenesis.
General Strategy for Broadening Adenovirus Tropism
Authors: Fontana Laura , Nuzzo Maurizio , Urbanelli Lorena , Monaci Paolo 
Published: September 2003

ABSTRACTIn spite of its broad host range, adenovirus type 5 (Ad5) transduces a number of clinically relevant tissues and cell types inefficiently, mostly because of low expression of the coxsackievirus-adenovirus receptor (CAR). To improve gene transfer to such cells, we modified the Ad5 fiber knob to recognize novel receptors. We expressed a functional Ad5 fiber knob domain on the capsid of phage λ and employed this display system to construct a large collection of ligands in the HI loop of the Ad5 knob. Panning this library on the CAR-negative mouse fibroblast cell line NIH 3T3 resulted in the identification of three clones with increased binding to these cells. Adenoviruses incorporating these ligands in the fiber gene transduced NIH 3T3 cells 2 or 3 orders of magnitude better than the parent vector. The same nonnative tropism was revealed in other cell types, independently of CAR expression. These Ad5 derivatives proved capable of transducing mouse and human primary immature dendritic cells with up to 100-fold increased efficiency.
Role of PACS-1 in Trafficking of Human Cytomegalovirus Glycoprotein B and Virus Production
Authors: Crump Colin M. , Hung Chien-Hui , Thomas Laurel , Wan Lei , Thomas Gary 
Published: September 2003

ABSTRACT
The final envelopment of herpesviruses during assembly of new virions is thought to occur by the budding of core viral particles into a late secretory pathway organelle, the trans-Golgi network (TGN), or an associated endosomal compartment. Several herpesvirus envelope glycoproteins have been previously shown to localize to the TGN when expressed independently from other viral proteins. In at least some cases this TGN localization has been shown to be dependent on clusters of acidic residues within their cytoplasmic domains. Similar acidic cluster motifs are found in endogenous membrane proteins that also localize to the TGN. These acidic cluster motifs interact with PACS-1, a connector protein that is required for the trafficking of proteins containing such motifs from endosomes to the TGN. We show here that PACS-1 interacts with the cytoplasmic domain of the HCMV envelope glycoprotein B (gB) and that PACS-1 function is required for normal TGN localization of HCMV gB. Furthermore, inhibition of PACS-1 activity in infected cells leads to a decrease in HCMV titer, whereas an increase in expression of functional PACS-1 leads to an increase in HCMV titer, suggesting that PACS-1 is required for efficient production of HCMV.