pinecone/core-2020-05-10-deduplication huggingface dataset:
The Role of Gamma Interferon in Immune Resistance to Vaginal Infection by Herpes Simplex Virus Type 2 in Mice

AbstractWe investigated the role of interferon gamma (IFN-γ) in a mouse model of immunity to vaginal infection by herpes simplex virus type 2 (HSV-2). Within 8 h after immune mice were challenged intravaginally with HSV-2, IFN-γ concentrations in vaginal secretions reached levels that can be antiviral in vitro. This rapid synthesis of IFN-γ occurred in immune-challenged mice but not in nonimmune-challenged mice, indicating that it required memory T cells. Immunostaining and in situ hybridization revealed that the IFN-γ was synthesized by cells whose morphological appearance suggested that they were lymphocytes and macrophage-like cells in the mucosa. The presence of IFN-γ in vaginal secretions was correlated with upregulation of MHC class II antigens in the epithelium and with vigorous (30-fold) recruitment of T and B lymphocytes into the vagina. In vivo administration of anti-IFN-γ to immune mice 17 h before virus challenge blocked the subsequent appearance of IFN-γ in vaginal secretions, blocked upregulation of class II antigens, blocked adherence of T cells to endothelium and their recruitment into the vagina, and markedly reduced immunity against reinfection of the vaginal epithelium
Domains of Rinderpest Virus Phosphoprotein Involved in Interaction with Itself and the Nucleocapsid Protein

AbstractThe yeast two-hybrid system was used to identify domains involved in specific in vivo interactions between the Rinderpest virus (RPV) phosphoprotein (P) and nucleocapsid protein (N). N and P genes were cloned in both the yeast GAL4 DNA-binding and GAL4 activation domain vectors, which enabled analysis of self and interprotein interactions. Mapping of the domain of P protein involved in its association with itself revealed that the COOH-terminal 32 amino acids (316–347) that forms a part of the highly conserved coiled coil region is important for interaction. In addition, just the coiled coil region of RPV P protein fused to the DNA-binding domain and activation domain of GAL4 was found to be sufficient to bring about activation of the β-galactosidase reporter. Similarly, mapping of the domains of P protein involved in its interaction with N protein revealed that NH2-terminal 59 amino acids and COOH-terminal 32 amino acids (316–347) involved in P–P interaction are simultaneously required for association with N protein. Interestingly, a P protein mutant with just the NH2-terminal 59 amino acids and the coiled coil domain with all other P protein regions deleted retained its ability to interact with N protein. Furthermore, we were able to show N and P protein interaction in vitro using recombinant N and P proteins expressed in Escherichia coli, demonstrating the existence of direct physical interaction between the two proteins
Internal Initiation of Translation of Bovine Viral Diarrhea Virus RNA

AbstractInitiation of translation on the bovine viral diarrhea virus (BVDV) internal ribosomal entry site (IRES) was reconstituted in vitro from purified translation components to the stage of 48S ribosomal initiation complex formation. Ribosomal binding and positioning on this mRNA to form a 48S complex did not require the initiation factors eIF4A, eIF4B, or eIF4F, and translation of this mRNA was resistant to inhibition by a trans-dominant eIF4A mutant that inhibited cap-mediated initiation of translation. The BVDV IRES contains elements that are bound independently by ribosomal 40S subunits and by eukaryotic initiation factor (eIF) 3, as well as determinants that mediate direct attachment of 43S ribosomal complexes to the initiation codon
Mutational Analyses of the Putative Calcium Binding Site and Hinge of the Turnip Crinkle Virus Coat Protein

AbstractThe turnip crinkle carmovirus (TCV) coat protein (CP) is folded into R (RNA-binding), S (shell), and P (protruding) domains. The S domain is an eight-stranded β barrel common to the coat protein subunits of most RNA viruses. A five-amino-acid hinge connects the S and P domains. In assembled particles, each pair of CP subunits is thought to bind a single calcium ion through interactions with three residues of one subunit and two residues of a neighboring subunit. These five residues comprise the putative calcium-binding site (CBS). The putative CBS and hinge are adjacent to one another. Mutations were introduced into the putative CBS or hinge in an effort to further determine the biological functions of TCV CP. One putative CBS mutant, TCV-M32, exhibited wild-type cell-to-cell movement but failed to move systemically in Nicotiana benthamiana, and particles were not detected. Another putative CBS mutant, TCV-M23, exhibited deficient cell-to-cell movement but particles accumulated in isolated protoplasts. Two other putative CBS mutants, TCV-M22 and -M33, showed wild-type cell-to-cell and systemic movement but elicited mild systemic symptoms that were somewhat delayed. All of the hinge mutants exhibited wild-type movement but some elicited non-wild-type symptoms. Point mutations in the putative CBS or hinge appear to alter virus–ion interactions, secondary structure, or particle conformation, thereby affecting interactions between the CP and plant hosts
Functional Characterization of the HveA Homolog Specified by African Green Monkey Kidney Cells with a Herpes Simplex Virus Expressing the Green Fluorescence Protein

AbstractWe cloned the gene specified by African monkey kidney cells (Vero) that codes for the homolog of the herpes virus entry mediator (HveA) specified by HeLa cells. The primary sequence of the monkey HveA (HveAs) differed significantly from HveA. Single amino acid differences were distributed throughout the amino and carboxyl terminal portions of the HveAs in comparison with the HveA, whereas certain regions were highly conserved. The predicted membrane spanning domains of the two receptors differed substantially due to insertions and deletions of short amino acid sequences. The ability of HveAs to mediate HSV virus entry was tested in a series of experiments using the recombinant virus KOS/EGFP, which constitutively expressed the enhanced green fluorescence protein (EGFP) and Chinese hamster ovary cells (CHO) transformed with the HveAs gene. The KOS/EGFP virus was constructed by inserting an EGFP gene cassette within the intergenic region between the UL53 (gK) and UL54 (ICP27) genes. The KOS/EGFP virus formed viral plaques and replicated as well as the wild-type KOS virus. HveAs-transformed CHO cells constitutively expressing HveAs mediated herpesvirus entry efficiently, whereas cells transformed with the HveAs gene in the noncoding orientation did not mediate virus entry. A genetically engineered protein composed of the amino-terminal portion of the HveAs protein fused to the heavy chain of mouse IgG immunoglobulin as well as mouse antibodies raised against HveAs blocked virus entry into HveAs-transformed CHO cells. Thus, HveAs is the functional homolog of HveA
Histidine Codons Appended to the Gene Encoding the RPO22 Subunit of Vaccinia Virus RNA Polymerase Facilitate the Isolation and Purification of Functional Enzyme and Associated Proteins from Virus-Infected Cells

AbstractVaccinia virus encodes a eukaryotic-like RNA polymerase composed of two large and six small subunit protein species. A replication-competent virus with 10 histidine codons added to the single endogenous J4R open reading frame was constructed. The altered migration of the 22-kDa subunit of RNA polymerase on SDS–polyacrylamide gel electrophoresis confirmed that J4R encoded the RPO22 subunit and that the mutant virus was genetically stable. The histidine-tagged RNA polymerase bound quantitatively to metal-affinity resins and was eluted in an active form upon addition of imidazole. Glycerol gradient sedimentation of the eluted fraction indicated that most of the RPO22 in infected cells is associated with RNA polymerase. Using stringent washing conditions, metal-affinity chromatography resulted in a several hundred-fold increase in RNA-polymerase-specific activity, and substantially pure enzyme was obtained with an additional conventional chromatography step. When mild conditions were used for washing the metal-affinity resin, the vaccinia virus-encoded capping enzyme, early transcription factor, and nucleoside triphosphate phosphohydrolase I specifically co-eluted with the tagged RNA polymerase, consistent with their physical association. The ability to selectively bind RNA polymerase to an affinity column provided a simple and rapid method of concentrating and purifying active enzyme and protein complexes
Latent Varicella-Zoster Virus in Human Dorsal Root Ganglia

AbstractTo understand further the molecular events underlying the process of Varicella-zoster virus (VZV) latency in human ganglionic tissues, in situ hybridisation (ISH) for VZV RNA and DNA, and PCR in situ amplification for VZV DNA were used in human dorsal root ganglia from 12 individuals (3 normal and 9 who had died with AIDS). The results showed that (a) two separate regions of the VZV genome, represented by genes 4 and 40, were detected in neurons in two normal and three AIDS ganglia, (b) evidence of transcription of VZV genes 4, 21, 29, and 63 was found in normal and AIDS cases, and (c) VZV DNA and RNA for the same gene (gene 29) was detected in neurons in serial tissue sections in three cases. Thus more than one region of the VZV genome is present in neurons during VZV ganglionic latency, and the presence of both a VZV gene and its corresponding RNA transcript can be shown to occur in the same localised region of DRG tissue
Transcript Mapping and Transregulatory Behavior of Varicella-Zoster Virus Gene 21, A Latency-Associated Gene

AbstractGene 21 is one of at least four genes transcribed during latent infection of varicella-zoster virus (VZV) in human ganglia. It may encode a nucleocapsid protein, but its function in lytic and latent infection is not clear. To characterize further the structure and the function of the gene 21 open reading frame (ORF 21), precise localization of its transcripts and their termini was determined by using Northern analysis, S1 nuclease or RNase protection, and primer extension assays. One abundant 3.5-kb transcript that spans ORF 21 was identified. A predominant transcription start site was defined at −78 nucleotide (nt) relative to the ORF 21 translation start codon ATG, and two potential TATA elements were identified at 26 and 83 nt upstream of the 5′ end of gene 21 transcripts. Transcription was found to terminate 210 nt beyond the ORF 21 translation stop codon and immediately before the start codon of ORF 22. In transient expression assays, the ORF 21 showed no significant transregulatory activity on promoters of diverse kinetic classes. The ORF 21 promoter, however, was transactivated strongly by VZV infection or by ORF 62
A Mechanism of Resistance to HIV-1 Entry: Inefficient Interactions of CXCR4 with CD4 and gp120 in Macrophages

AbstractTo test the hypothesis that inefficient interactions of CXCR4 with CD4 and gp120 could affect HIV-1 entry, we incubated macrophages, monocytes, and lymphocytes with gp120 and coimmunoprecipitated CD4 by using anti-CXCR4 antibodies. CD4 was efficiently coimmunoprecipitated in lymphocytes and monocytes but not in macrophages. Overexpression of CD4 in macrophages resulted in detection of CD4–CXCR4 and gp120–CD4–CXCR4 complexes in parallel with the restoration of macrophage fusion susceptibility. These results suggest a mechanism of resistance to entry of some X4 HIV-1 strains into macrophages and a method for dissection of the initial stages of HIV entry
Nitric Oxide Down-Regulates Epstein–Barr Virus Reactivation in Epithelial Cell Lines

AbstractNitric oxide (NO), a mediator of biological functions, has an antimicrobial activity against a variety of pathogens including viruses. In this study, we found that a constitutive, low level of inducible NO synthase (iNOS) mRNA was expressed in the EBV-infected gastric tissue-derived GT38 and GT39 cell lines, by analysis with the reverse transcription–polymerase chain reaction (RT–PCR) and Southern blotting. Treatment of these cells with a specific NOS inhibitor, NG-monomethyl-l-arginine (l-NMMA), induced the immediate-early, EBV transactivator gene BZLF1 protein ZEBRA, suggesting a significant increase in EBV reactivation by l-NMMA. Northern blotting demonstrated that BZLF1 and BRLF1 transcripts were also induced by 12-O-tetradecanoylphorbol-13 acetate (TPA). Meanwhile, constitutive expression of iNOS mRNA was inhibited by TPA. l-NMMA also enhanced TPA-induced expression of the BZLF1 gene. On the other hand, a NO donor, S-nitroso-N-acetylpenicillamine (SNAP), which releases NO in an aqueous solution, inhibited the TPA-induced BZLF1 gene expression in a dose-dependent manner at both mRNA and protein levels. These results demonstrated that NO is a regulatory factor in maintaining virus latency via inhibiting EBV reactivation in the infected epithelial cells
Role of the Cytoplasmic Tail of gE in Antibody-Induced Redistribution of Viral Glycoproteins Expressed on Pseudorabies-Virus-Infected Cells

AbstractPseudorabies virus (PrV) glycoprotein gE is a nonessential glycoprotein involved in virulence and spread of the virus. It also has an important, yet unknown, function during antibody-induced capping of viral glycoproteins on the plasma membrane of PrV-infected swine kidney cells. In the present study, it was shown, by the use of a PrV strain expressing a truncated gE glycoprotein, that the cytoplasmic tail of gE is of significant importance for viral glycoprotein capping to occur. In addition, using PrV strains carrying point mutations in the cytoplasmic tail of gE, it was demonstrated that two tyrosine-based motifs are very important for correct functioning of gE during viral glycoprotein capping. Furthermore it was shown that genistein and tyrphostin, two tyrosine kinase activity inhibitors, inhibit viral glycoprotein capping in a concentration-dependent manner. In conclusion, it can be stated that efficient antibody-induced viral glycoprotein capping requires the presence of two YxxL sequences in the cytoplasmic tail of glycoprotein gE, as well as the activation of a tyrosine phosphorylation signal transduction pathway
Isolation of a Human Endogenous Retroviral HERV-H Element with an Open env Reading Frame

AbstractAbout 100 elements of the human endogenous retroviral HERV-H family have full-length env genes potentially coding for Env proteins with sequences highly similar to the immunosuppressive peptide CKS-17 from the MLV transmembrane protein p15E. However, previously sequenced HERV-H env genes have contained stop codons or framehifts. To isolate elements with open env reading frames, we first tried to assess the diversity of HERV-H env genes by comparing PCR-generated env sequences from genomic DNA with published HERV-H sequences. A region at the beginning of env displayed a similarity of 84–98% among 15 different elements. We then used a probe from one of the PCR-generated clones, 98% similar to the consensus sequence in this region, to screen a human genomic lambda library. Three HERV-H elements displaying ca. 98% identity in the env gene were isolated and were shown to have integrated relatively recently, after the divergence of the orangutan and the african great ape lineages. One of these elements, HERV-H19, had a 1752-bp open env reading frame, producing a 77-kDa Env protein in in vitro translation reactions. This is the first demonstration of a coding competent member of the HERV-H family. These findings raise the possibility that HERV-H Env proteins may play a biological role in human cells
Immune Responses and Protection Obtained by Oral Immunization with Rotavirus VP4 and VP7 DNA Vaccines Encapsulated in Microparticles

AbstractProtective immune responses in mice were obtained after oral immunization with rotavirus DNA vaccines encapsulated in poly(lactide-co-glycolide) (PLG) microparticles. The DNA vaccines used encoded outer capsid proteins VP4 and VP7; proteins that are the basis for rotavirus serotyping and the generation of virus neutralizing antibodies. One dose of vaccine was given to BALB/c mice by oral gavage (75 μg DNA/mouse). Rotavirus-specific serum antibodies and intestinal IgA antibodies were detectable by 6 weeks postimmunization. After challenge with homologous murine rotavirus at 12 weeks postimmunization, fecal rotavirus antigen was reduced significantly in immunized mice compared with controls. Protective immunity also was generated by oral delivery of unencapsulated VP 7 DNA vaccine but to a lesser degree. These results demonstrate that the oral route is effective for generating protective immune responses with rotavirus DNA vaccines targeting neutralization antigens
Goat Milk Epithelial Cells Are Highly Permissive to CAEV Infection in Vitro

AbstractThe main route of small ruminant lentivirus dissemination is the ingestion of infected cells present in colostrum and milk from infected animals. However, whether only macrophages or other cell subtypes are involved in this transmission is unknown. We derived epithelial cell cultures, 100% cytokeratin positive, from milk of naturally infected and noninfected goats. One such culture, derived from a naturally infected goat, constitutively produced a high titer of virus in the absence of any cytopathic effect. The other cultures, negative for natural lentivirus infection, were tested for their susceptibility to infection with the CAEV-CO strain and a French field isolate CAEV-3112. We showed that milk epithelial cells are easily infected by either virus and produce viruses at titers as high as those obtained in permissive goat synovial membrane cells. The CAEV-CO strain replicated in milk epithelial cells in absence of any cytopathic effect, whereas the CAEV-3112 field isolate induced both cell fusion and cell lysis. Our results suggest that CAEV-infected milk epithelial cells of small ruminants may play an important role in virus transmission and pathogenesis
Neutralizing Antibody Responses and Evolution of Antigenic Variants in Monozygotic Twin Lambs Infected with Phenotypically Distinct Ovine Lentiviruses

AbstractOvine lentivirus (OvLV) isolates 85/34 (OvLV 34) and 84/28 (OvLV 28) were initially characterized as phenotypically distinct “rapid/high” and “slow/low” strains based on replication kinetics, syncytiogenesis, and cell lysis in vitro. In the present study, sera from OvLV-34- or OvLV-28-infected monozygotic twin lambs defined these virus strains as distinct neutralization serotypes. We also show that immune recognition of at least one OvLV neutralization epitope is influenced by genetic differences between lambs. Additional studies determined the neutralization phenotype of virus isolates from alveolar macrophages of OvLV-34- or OvLV-28-infected lambs, evaluated the role of neutralizing antibodies in selection and persistence of antigenic variants, and related the severity of OvLV-induced lymphoid interstitial pneumonia (LIP) to the evolution of neutralization variants. These studies demonstrate that (i) macrophage-associated OvLV neutralization variants can arise in the presence or the absence of neutralizing antibodies directed to inoculum viruses, (ii) OvLV variants persist in macrophages in the presence of serum neutralizing antibodies, and (iii) the emergence of OvLV variants is apparently unrelated to the severity of LIP
Herpes Simplex Virus Induces Intracellular Redistribution of E2F4 and Accumulation of E2F Pocket Protein Complexes

AbstractAccumulation of E2F-p107 and E2F-pRB DNA binding complexes occurred after herpes simplex virus infection of U2-OS cells. Accumulation of E2F-p107 also occurred by 4 h p.i. in C33 cells. This corresponded to a time when host DNA synthesis was reduced by 50%, and lagged by ≥1 h, the onset of viral DNA synthesis. To determine the basis for increased nuclear E2F complexes, we investigated the effects of virus infection on the intracellular distribution of the E2F-dependent DNA binding complexes and their protein constituents. Western blot analyses of whole cell extracts revealed that amounts of E2F4, E2F1, DP1, and p107 remained unchanged after infection of C33 cells. Analysis of cytoplasmic and nuclear fractions, however, revealed that cytoplasmic E2F4 decreased and nuclear E2F4 increased. This correlated with a loss of cytoplasmic E2F DNA-binding activity and a corresponding increase in nuclear DNA-binding activity. Concomitant with its redistribution, the apparent molecular weight of total and p107-associated E2F4 increased, at least partially as a result of protein phosphorylation. Increased nuclear E2F-pRB in U2-OS cells was accompanied by the conversion of pRB from a hyper- to a hypophosphorylated state. Infection of U2-OS cells with viral mutants indicated that viral protein IE ICP4 was necessary for the decrease in cytoplasmic E2F-p107, and that viral protein DE ICP8 was required for nuclear accumulation of p107-E2F. In contrast, ICP8 was not required for accumulation of E2F-pRB. These results indicate that the increase in E2F-p107 may be explained by the redistribution and modification of E2F4 and the increase in E2F-pRB by modification of pRB
The Transcriptional Activation Domain of VP16 Is Required for Efficient Infection and Establishment of Latency by HSV-1 in the Murine Peripheral and Central Nervous Systems

AbstractThe herpes simplex virus (HSV) transactivator VP16 is a structural component of the virion that activates immediate-early viral gene expression. The HSV-1 mutant in1814, which contains a 12-bp insertion that compromises the transcriptional function of VP16, replicated to a low level if at all in the trigeminal ganglia of mice (I. Steiner, J. G. Spivack, S. L. Deshmane, C. I. Ace, C. M. Preston, and N. W. Fraser (1990). J. Virol. 64, 1630–1638; Valyi-Nagy et al., unpublished data). However, in1814 did establish a latent infection in the ganglia after corneal inoculation from which it could be reactivated. In this study, several HSV-1 strains were constructed with deletions in the VP16 transcriptional activation domain. These viruses were viable in cell culture, although some were significantly reduced in their ability to initiate infection. A deletion mutant completely lacking the activation domain of VP16 (RP5) was unable to replicate to any detectable level or to efficiently establish latent infections in the peripheral and central nervous systems of immunocompetent mice. However, similar to in1814, RP5 formed a slowly progressing persistent infection in immunocompromised nude mice. Thus RP5 is severely neuroattenuated in the murine model of HSV infection. However, the activation domain of VP16 is not essential for replication in the nervous system, since we observed a slow progressive infection persisting in the absence of an immune response
Involvement of a p53-Dependent Pathway in Rubella Virus-Induced Apoptosis

AbstractIn light of the important role of apoptotic cell death in the pathogenesis of several viral infections, we asked whether the cytopathogenicity evoked by rubella virus (RV) might also involve apoptotic mechanisms. The To-336 strain of RV induced apoptosis in Vero and RK-13 cells, but not in fibroblast cell lines. UV-inactivated RV virions did not elicit the apoptotic response, indicating that productive infection is required for the induction of cell death. Both p53 and p21 protein levels were highly elevated in RV-infected Vero cells. The level of p21 mRNA was increased, while expression of the p53 gene was unaffected by RV infection. A dominant-negative p53 mutant (p53W248) conferred partial protection from RV-induced apoptosis. These data implicate a p53-dependent apoptotic pathway in the cytopathogenicity of RV, thereby suggesting a mechanism by which RV exerts its teratogenic effects