pinecone/core-2020-05-10-deduplication huggingface dataset:
Immunogenic and Antigenic Dominance of a Nonneutralizing Epitope over a Highly Conserved Neutralizing Epitope in the gp41 Envelope Glycoprotein of Human Immunodeficiency Virus Type 1: Its Deletion Leads to a Strong Neutralizing Response

AbstractThe Kennedy peptide, 731PRGPDRPEGIEEEGGERDRDRS752, from the cytoplasmic domain of the gp41 transmembrane envelope glycoprotein of HIV-1 contains a conformationally dependent neutralizing epitope (ERDRD) and a linear nonneutralizing epitope (IEEE). No recognized murine T cell epitope is present. The peptide usually stimulates virus-specific antibody, but this is not always neutralizing. Here we show that IEEE (or possibly IEEE plus adjacent sequence) is immunogenically and antigenically dominant over the ERDRD neutralizing epitope. Thus rabbits immunized in a variety of routes, doses, and adjuvants with a chimeric cowpea mosaic virus (CPMV) expressing the Kennedy peptide on its surface (CPMV-HIV/1) synthesized IEEE-specific serum antibody but no ERDRD-specific or HIV-1-neutralizing antibody. To test if this resulted from immunodominance or from a hole in the antibody repertoire, we immunized rabbits with chimera CPMV-HIV/29, which expresses the GERDRDR part of the Kennedy sequence. This chimera readily stimulated ERDRD-specific, neutralizing antibody. In mice the situation was less extreme, but individual animals with low neutralizing titers had a high ratio of IEEE-specific:ERDRD-specific antibody. Data are consistent with immunodominance of IEEE over ERDRD in the Kennedy peptide. IEEE-specific antibody was also antigenically dominant and prevented ERDRD-specific antibody from binding to its epitope and from neutralizing HIV-1. It may be that HIV-1 has evolved a nonneutralizing immunodominant epitope that allows it to possess a neutralizing epitope without suffering the consequences, and this idea is supported by the covariance of both epitope sequences. To our knowledge this is the first example of a defined sequence that controls the activity of an adjacent epitope
Efficient Rescue of Infectious Bursal Disease Virus from Cloned cDNA: Evidence for Involvement of the 3′-Terminal Sequence in Genome Replication

AbstractTo study the mechanism of replication of infectious bursal disease virus (IBDV), and to determine factors on the IBDV RNA which are involved in viral replication, we used cloned full-length cDNA of both the A- and B-segments to generate infectious IBDV. Infectious IBDV was rescued from plasmids that contained full-length IBDV cDNA behind a T7 promoter, by transfecting these plasmids into cells which were infected with a recombinant Fowlpox virus that expressed T7 RNA polymerase. By using the cDNA transfection system we evaluated the effect of the length of the 3′ terminus of the A-segment plus strand of IBDV. Although wild-type IBDV predominantly contains four cytosines at the 3′ terminus, no difference in virus yield was found when virus was rescued from cDNAs containing three to six adjacent cytosines. When the 3′ terminus was shorter than three cytosines the efficiency to generate infectious IBDV from cDNA was reduced, but IBDV could still be recovered reproducibly. The rescued viruses from cDNAs containing 3′-terminal deletions appeared to have a restored 3′-terminal sequence. The missing nucleotides are probably restored by using complementary bases of a stem-loop structure as template
Poliovirus Protease 3Cpro Kills Cells by Apoptosis

AbstractThe tetracycline-based Tet-Off expression system has been used to analyze the effects of poliovirus protease 3Cpro on human cells. Stable HeLa cell clones that express this poliovirus protease under the control of an inducible, tightly regulated promoter were obtained. Tetracycline removal induces synthesis of 3C protease, followed by drastic morphological alterations and cellular death. Degradation of cellular DNA in nucleosomes and generation of apoptotic bodies are observed from the second day after 3Cpro induction. The cleavage of poly(ADP-ribose) polymerase, an enzyme involved in DNA repair, occurs after induction of 3Cpro, indicating caspase activation by this poliovirus protease. The 3Cpro-induced apoptosis is blocked by the caspase inhibitor z-VAD-fmk. Our findings suggest that the protease 3C is responsible for triggering apoptosis in poliovirus-infected cells by a mechanism that involves caspase activation
Large-Scale Monitoring of Host Cell Gene Expression during HIV-1 Infection Using cDNA Microarrays

AbstractHuman immunodeficiency virus type 1 (HIV-1) infection alters the expression of host cell genes at both the mRNA and protein levels. To obtain a more comprehensive view of the global effects of HIV infection of CD4-positive T-cells at the mRNA level, we performed cDNA microarray analysis on approximately 1500 cellular cDNAs at 2 and 3 days postinfection (p.i.) with HIV-1. Host cell gene expression changed little at 2 days p.i., but at 3 days p.i. 20 cellular genes were identified as differentially expressed. Genes involved in T-cell signaling, subcellular trafficking, and transcriptional regulation, as well as several uncharacterized genes, were among those whose mRNAs were differentially regulated. These results support the hypothesis that HIV-1 infection alters expression of a broad array of cellular genes and provides a framework for future functional studies on the differentially expressed mRNA products
Insertional Mutagenesis of AAV2 Capsid and the Production of Recombinant Virus

AbstractThe structural genes of adeno-associated virus serotype 2 (AAV2) have been altered by linker insertional mutagenesis in order to define critical components of virion assembly and infectivity. An in-frame restriction site linker was inserted across the capsid coding domain of a recombinant plasmid. After complementation in vivo, recombinant AAV2 viruses were generated and assayed for capsid production, packaging, transduction, heparin agarose binding, and morphology. Three classes of capsid mutants where identified. Class I mutants expressed structural proteins but were defective in virion assembly. Class II mutants generated intact virions that protected the viral genome from DNase, but failed to infect target cells. The majority of these mutants bound the heparin affinity matrix, suggesting that attachment to the AAV primary receptor was not rate limiting. One class II mutant, H2634, assembled virions and bound heparin using only Vp3, indicating that this subunit is responsible for mediating AAV receptor attachment. Finally, class III mutants assembled virions, encapsidated DNA, and infected target cells. Infectivity of these mutants ranged from 5 to 100% of that of the wild-type, demonstrating for the first time the ability to alter capsid proteins without interfering with infectivity. These AAV virions with altered capsid subunits will provide critical templates for manipulating AAV vectors for cell-specific gene delivery in vivo. In summary, the AAV capsid variants described here will facilitate further study of virus assembly, entry, and infection, as well as advance the development of this versatile vector system
The Vaccinia Virus 39-kDa Protein Forms a Stable Complex with the p4a/4a Major Core Protein Early in Morphogenesis

AbstractThe vaccinia virus (VV) 39-kDa protein, the product of the A4L gene, is a highly antigenic protein of the viral core. Pulse-chase and immunoprecipitation experiments have shown that the 39-kDa protein interacts with p4a (encoded by the A10L gene), the precursor of the most abundant virion protein. This interaction is maintained with the processed 4a form that arises during virion maturation. The controlled disruption of mature viral particles showed that the 39-kDa and 4a proteins are tightly bound within the virion. Immunoelectron microscopy showed that both proteins first localize within the cytoplasm and later accumulate inside the viral factories, reaching these locations via a mechanism apparently unrelated to cellular membranes. Double labeling experiments showed a colocalization of both proteins in all virus-induced structures
Rapid and Efficient Cell-to-Cell Transmission of Human Immunodeficiency Virus Infection from Monocyte-Derived Macrophages to Peripheral Blood Lymphocytes

AbstractMacrophages are considered of central importance in cell-to-cell transmission of human immunodeficiency virus (HIV) infection in vivo. In this report, we describe a novel cell-to-cell transmission model using HIV-infected monocyte-derived macrophages (MDMs) as donor cells and peripheral blood lymphocytes (PBLs) as recipients. Virus was transmitted during a 2-h coincubation period from intracellular or tightly cell-associated viral stores in adherent infected MDMs to nonadherent CD3+ PBLs. Transmission required cell contact, but syncytia formation was not observed. HIV cell-to-cell transmission occurred in both allogeneic and autologous systems, and replication was higher in phytohemagglutinin (PHA)-stimulated than unstimulated recipient PBLs. In contrast, transmission of infection by cell-free virus was barely detectable without PHA stimulation of recipients, suggesting the cell–cell interaction may have provided stimuli to recipient cells in the cell-to-cell system. Viral DNA levels increased 5–24 h postmixing, and this increase was inhibited by pretreatment of cells with the reverse transcription inhibitor azidothymidine, indicating de novo reverse transcription was involved. Cell-to-cell transmission was more efficient than infection with cell-free virus released from donor MDMs, or 0.1 TCID50/cell cell-free viral challenge. This model provides a system to further investigate the mechanisms and characteristics of HIV cell-to-cell transmission between relevant primary cells that may be analogous to this important mode of virus spread in vivo
Protection of Macaques against a SHIV with a Homologous HIV-1 Env and a Pathogenic SHIV-89.6P with a Heterologous Env by Vaccination with Multiple Gene-Deleted SHIVs

AbstractTo evaluate the potential of SHIVs as anti-HIV-1 live vaccines, we constructed two gene-deleted SHIVs, designated SHIV-drn and SHIV-dxrn. The former lacks vpr/nef and the latter lacks vpx/vpr/nef. Four macaques that had been vaccinated with SHIV-drn were challenged with SHIV-NM-3rN, which has an HIV-1 Env that is the same as that of SHIV-drn. No challenge virus was detected by DNA PCR in, or recovered from, two of the macaques. In the other two, challenge virus was detected once and twice, respectively. Plasma viral loads were much lower than those in unvaccinated controls. Another four macaques were vaccinated with SHIV-dxrn. These macaques showed resistance but less than that of SHIV-drn-vaccinated macaques. When the two SHIV-drn-vaccinated macaques were challenged with pathogenic SHIV-89.6P, which has an HIV-1 Env that is antigenically different from that of SHIV-drn, replication of the challenge virus was restricted, and the usual decrease in the number of CD4+ cells was prevented. In this protection, it is noteworthy that protection involved not only neutralizing antibodies and killer cell activity, but also other unknown specific and nonspecific immunity elicited by the infection
Neuropathogenicity and Sensitivity to Antibody Neutralization of Lactate Dehydrogenase-Elevating Virus Are Determined by Polylactosaminoglycan Chains on the Primary Envelope Glycoprotein

AbstractCommon strains of lactate dehydrogenase-elevating virus (LDV, an arterivirus), such as LDV-P and LDV-vx, are highly resistant to antibody neutralization and invariably establish a viremic, persistent, yet asymptomatic, infection in mice. Other LDV strains, LDV-C and LDV-v, have been identified that, in contrast, are highly susceptible to antibody neutralization and are incapable of a high viremic persistent infection, but at the same time have gained the ability to cause paralytic disease in immunosuppressed C58 and AKR mice. Our present results further indicate that these phenotypic differences represent linked properties that correlate with the number of N-glycosylation sites associated with the single neutralization epitope on the short ectodomain of the primary envelope glycoprotein, VP-3P. The VP-3P ectodomains of LDV-P/vx possess three N-glycosylation sites, whereas those of LDV-C/v lack the two N-terminal sites. We have now isolated four independent neutralization escape variants of neuropathogenic LDV-C and LDV-v on the basis of their ability to establish a high viremic persistent infection in mice. The VP-3P ectodomains of all four variants had specifically regained two N-glycosylation sites concomitant with decreased immunogenicity of the neutralization eptitope and decreased sensitivity to antibody neutralization as well as loss of neuropathogenicity
SIV/HIV nef Recombinant Virus (SHIVnef) Produces Simian AIDS in Rhesus Macaques

AbstractThe simian immunodeficiency virus (SIV) nef gene is an important determinant of viral load and acquired immunodeficiency syndrome (AIDS) in macaques. A role(s) for the HIV-1 nef gene in infection and pathogenesis was investigated by constructing recombinant viruses in which the nef gene of the pathogenic molecular clone SIVmac239 nef was replaced with either HIV-1sf2nef or HIV-1sf33nef. These chimeras, designated SHIV-2nef and SHIV-33nef, expressed HIV-1 Nef protein and replicated efficiently in cultures of rhesus macaque lymphoid cells. In two SHIV-2nef-infected juvenile rhesus macaques and in one of two SHIV-33nef-infected juvenile macaques, virus loads remained at low levels in both peripheral blood and lymph nodes in acute and chronic phases of infection (for >83 weeks). In striking contrast, the second SHIV-33nef-infected macaque showed high virus loads during the chronic stage of infection (after 24 weeks). CD4+ T-cell numbers declined dramatically in this latter animal, which developed simian AIDS (SAIDS) at 47–53 weeks after inoculation; virus was recovered at necropsy at 53 weeks and designated SHIV-33Anef. Sequence analysis of the HIV-1sf33nef gene in SHIV-33Anef revealed four consistent amino acid changes acquired during passage in vivo. Interestingly, one of these consensus mutations generated a tyr-x-x-leu (Y-X-X-L) motif in the HIV-1sf33 Nef protein. This motif is characteristic of certain endocytic targeting sequences and also resembles a src-homology region-2 (SH-2) motif found in many cellular signaling proteins. Four additional macaques infected with SHIV-33Anef contained high virus loads, and three of these animals progressed to fatal SAIDS. Several of the consensus amino acid changes in Nef, including Y-X-X-L motif, were retained in these recipient animals exhibiting high virus load and disease. In summary, these findings indicate that the SHIV-33Anef chimera is pathogenic in rhesus macaques and that this approach, i.e., construction of chimeric viruses, will be important for analyzing the function(s) of HIV-1 nef genes in immunodeficiency in vivo, testing antiviral therapies aimed at inhibiting AIDS, and investigating adaptation of this HIV-1 accessory gene to the macaque host
Characterization of Recombination Events Leading to the Production of an Ecotropic Replication-Competent Retrovirus in a GP+envAM12-Derived Producer Cell Line

AbstractReplication-competent retrovirus (RCR) was identified in a GP+envAM12-derived producer cell, containing the MFG-S-Neo retroviral vector, using a marker rescue assay. Studies were undertaken to determine the origin and structure of this RCR. Receptor interference assays demonstrated that the virus was pseudotyped with an ecotropic envelope. Molecular analysis demonstrated the presence of a MoMLV ecotropic env recombinant where the neomycin resistance gene of the MFG-S-Neo vector was replaced by MoMLV ecotropic env. Additional recombinants linking the retroviral pol gene to neo and the neo gene to MoMLV env were also identified. A full-length MoMLV retroviral genome was detected by nested PCR in the contaminated amphotropic producer cells and in cells infected with its supernatant. Unexpectedly, this was also present in the GP+E86 packaging cells together with a previously undescribed envelope construct possessing a full 5′ and 3′ LTR, although these cells were consistently negative for the presence of RCR. These anomalies in the GP+E86 packaging cell line result in increased homology with the MFG-S-Neo vector, leading to an increased risk for the production of RCR. Our findings point to a need for increased vigilance when using these packaging lines to generate replication-defective retrovirus
The Primate Lentivirus-Encoded Nef Protein Can Regulate Several Steps of the Viral Replication Cycle

AbstractThe primate lentiviruses encode a protein, Nef, which is required for efficient viral replication in their host. Several biological activities of nef identified in vitro may contribute to this requirement in vivo, including receptor modulation and interference with cellular signaling pathways. We show that HIV- and SIV-encoded Nef can enhance virus production within a single viral replication cycle, not only by increasing viral infectivity, as previously reported, but also by acting through the efficiency of viral transcription and of viral release
Epstein–Barr Virus Nuclear Antigen-1 Binds to Nuclear Transporter Karyopherin α1/NPI-1 in Addition to Karyopherin α2/Rch1

AbstractWe searched for cellular proteins that interact with Epstein–Barr (EBV) virus nuclear antigen-1, which is a latent EBV origin-binding protein detected in all EBV latently infected cells and essential for maintenance of the latent EBV genome, by a yeast two-hybrid screening of a B lymphocyte cDNA library in this study. Interaction of polypeptides synthesized from three selected cDNA clones with EBNA-1 proteins was confirmed in vitro using their glutathione-S-transferase-fusion polypeptides and by coimmunoprecipitation analyses of B cell extracts with anti-EBNA-1 monoclonal antibodies and monospecific antibodies against cellular proteins of interest. We report the following: (i) Karyopherin α (karyopherin α1, hSRP1, and NPI-1), an adaptor subunit of nuclear localization signal receptors, which direct proteins to the nuclear pore, interacted with EBNA-1. (ii) EBNA-1 proteins endogenous in the B cell line Raji of Burkitt lymphoma origin bound to another adaptor protein, karyopherin α2 (hSRP1α, hRch1), interactions of which to recombinant EBNA-1 polypeptides were previously reported. (iii) Nearly 90% of all the cDNA clones examined was p32 (SF2-associated P32, p32/TAP, and gC1q-R), and endogenous EBNA-1 proteins in the Raji cells bound to p32, a potential of which to affect localization of EBNA-1 in transfected Vero cells has been recently suggested. These results suggest that EBNA-1, which has the unique NLS containing Lys–Arg and overlapping with one of the phosphorylation domains, is recognized and transported to the nuclei by these two distinct karyopherin α proteins, which are differentially expressed in different cell types, implying a regulatory localization system for EBNA-1
A Mutation Correcting the DNA Interaction Defects of a Mutant Phage λ Terminase, gpNu1 K35A Terminase

AbstractTerminase, the DNA packaging enzyme of bacteriophage λ, is a heteromultimer composed of gpNu1 (181 aa) and gpA (641 aa) subunits, encoded by the λ Nu1 and A genes, respectively. Similarity between the deduced amino acid sequences of gpNu1 and gpA and the nucleotide binding site consensus sequence suggests that each terminase subunit has an ATP reactive center. Terminase has been shown to have two distinct ATPase activities. The gpNu1 subunit has a low-affinity ATPase stimulated by nonspecific DNA and gpA has a high-affinity ATPase. In previous work, a mutant terminase, gpNu1 K35A holoterminase, had a mild defect in interactions with DNA, such that twofold increased DNA concentrations were required both for full stimulation of the low-affinity ATPase and for saturation of the cos cleavage reaction. In addition, the gpNu1 K35A terminase exhibited a post-cleavage defect in DNA packaging that accounted for the lethality of the Nu1 K35A mutation [Y. Hwang and M. Feiss (1997) Virology 231, 218–230]. In the work reported here, a mutation in the turn of the putative helix-turn-helix DNA binding domain has been isolated as a suppressor of the gpNu1 K35A change. This suppressor mutation causes the change A14V in gpNu1. A14V reverses the DNA-binding defects of gpNu1 K35A terminase, both for stimulation of the low-affinity ATPase and for saturation of the cos cleavage defect. A14V suppresses the post-cleavage DNA packaging defect caused by the gpNu1 K35A change
Possible Emergence of New Geminiviruses by Frequent Recombination

AbstractAlthough exchange of genetic information by recombination plays a role in the evolution of viruses, the extent to which it generates diversity is not clear. We analyzed genomes of geminiviruses for recombination using a new statistical procedure developed to detect gene conversions. Geminiviruses (family, Geminiviridae) are a group of plant viruses characterized by a genome of circular single-stranded DNA (∼2700 nucleotides in length) encapsidated in twinned quasi-isometric particles. Complete nucleotide sequences of geminiviruses were aligned, and recombination events were detected by searching pairs of viruses for sequences that are significantly more similar than expected based on random distribution of polymorphic sites. The analyses revealed that recombination is very frequent and occurs between species and within and across genera. Tests identified 420 statistically significant recombinant fragments distributed across the genome. The results suggest that recombination is a significant contributor to geminivirus evolution. The high rate of recombination may be contributing to the recent emergence of new geminivirus diseases
Identification of a Novel 0.7-kb Polyadenylated Transcript in the LAT Promoter Region of HSV-1 That Is Strain Specific and May Contribute to Virulence

AbstractHerpes Simplex virus expresses latency-associated transcripts (LATs) the function of which remains obscure despite increasing knowledge of their structure and expression. Upstream of the LAT coding region is a region of the genome that is poorly characterized although it lies in an area that is responsible for modulation of reactivation efficiency in two different animal models. Transcript mapping with strains 17, McKrae, KOS, and F has revealed strain differences in this region of the viral genome. Strain 17 and McKrae expressed a novel polyadenylated 0.7-kb transcript that is absent from KOS and F. This transcript is expressed in the LAT direction and has the kinetics of a true late gene during the lytic cycle of infection. A deletion mutant, 17ΔBsa, which does not express the 0.7-kb RNA, is less virulent than the parental strain 17. A rescuant with F sequence (17ΔBsa/RF) shows virulence similar to F, whereas a rescuant with strain 17 sequence (17ΔBsa/R17) is similar to strain 17. Virulence is altered by deletion or substitution in the region encoding the 0.7-kb transcript (BsaI-BsaI); however, reactivation in the mouse explant cocultivation assay or the adrenergically induced rabbit reactivation model remained unchanged. The importance of this region for virulence is discussed