bluuebunny/crossref_metadata_2025_split huggingface dataset:
An Endoplasmic Reticulum-Targeting Signal Sequence Enhances the Immunogenicity of an Immunorecessive Simian Virus 40 Large T Antigen Cytotoxic T-Lymphocyte Epitope
Authors: Fu Tong-Ming , Mylin Lawrence M. , Schell Todd D. , Bacik Igor , Russ Gustav , Yewdell Jonathan W. , Bennink Jack R. , Tevethia Satvir S. 
Published: December 2019

ABSTRACTAn immunological hierarchy among three H-2Db-restricted cytotoxic T lymphocyte (CTL) determinants in simian virus 40 (SV40) large T antigen (Tag) was described previously: determinants I and II/III are immunodominant, whereas determinant V is immunorecessive. To assess the immunogenicity of each determinant individually and define mechanisms that contribute to the immunorecessive nature of determinant V, we constructed a panel of recombinant vaccinia viruses (rVVs) expressing minigenes encoding these determinants in various polypeptide contexts. We found the following. (i) Immunization of mice with an rVV encoding full-length SV40 Tag resulted in priming for CTL responses to determinants I and II/III but not determinant V. (ii) rVVs encoding peptide I or II/III in the cytosol or targeted to the endoplasmic reticulum (ER) were highly antigenic and immunogenic. (iii) rVVs encoding peptide V minigenes were antigenic and immunogenic if the peptide was targeted to the ER, expressed in the cytosol with short flanking sequences, or expressed from within a self-protein, murine dihydrofolate reductase. (iv) Presentation of the nonflanked peptide V (preceded by a Met codon only) could be enhanced by using a potent inhibitor of the proteasome. (v) H-2Db–epitope V peptide complexes decayed more rapidly than complexes containing epitope I or II/III peptides. In brefeldin A blocking experiments, functional epitope V complexes were detected longer on targets expressing ER-targeted epitope V than on targets expressing forms of epitope V dependent on the transporter associated with antigen processing. Therefore, limited formation of relatively unstable cell surface H-2Dbcomplexes most likely contributes to the immunorecessive nature of epitope V within SV40 Tag. Increasing the delivery of epitope V peptide to the major histocompatibility complex class I presentation pathway by ER targeting dramatically enhanced the immunogenicity of epitope V.
mRNA Instability Elements in the Human Papillomavirus Type 16 L2 Coding Region
Authors: Sokolowski Marcus , Tan Wei , Jellne Marianne , Schwartz Stefan 
Published: December 2019

ABSTRACT

Human papillomavirus capsid proteins L1 and L2 are detected only in terminally differentiated cells, indicating that expression of the L1 and L2 genes is blocked in dividing cells. The results presented here establish that the human papillomavirus type 16 L2 coding region contains
cis
-acting inhibitory sequences. When placed downstream of a reporter gene, the human papillomavirus type 16 L2 sequence reduced both mRNA and protein levels in an orientation-dependent manner. Deletion analysis revealed that the L2 sequence contains two
cis
-acting inhibitory RNA regions. We identified an inhibitory region in the 5′-most 845 nucleotides of L2 that acted by reducing cytoplasmic mRNA stability and a second, weaker inhibitory region in the 3′ end of L2. In contrast, human papillomavirus type 1 L1 and L2 genes did not encode strong inhibitory sequences. This result is consistent with observations of high virus production in human papillomavirus type 1-infected tissue, whereas only low levels of human papillomavirus type 16 virions are detectable in infected epithelium. The presence of inhibitory sequences in the L1 and L2 mRNAs may aid the virus in avoiding the host immunosurveillance and in establishing persistent infections.
Human MxA Protein Confers Resistance to Semliki Forest Virus and Inhibits the Amplification of a Semliki Forest Virus-Based Replicon in the Absence of Viral Structural Proteins
Authors: Landis Heinrich , Simon-Jödicke Angela , Klöti Andreas , Di Paolo Claudio , Schnorr Jens-Jörg , Schneider-Schaulies Sibylle , Hefti Hans Peter , Pavlovic Jovan 
Published: December 2019

ABSTRACTMx proteins form a small family of interferon (IFN)-induced GTPases with potent antiviral activity against various negative-strand RNA viruses. To examine the antiviral spectrum of human MxA in homologous cells, we stably transfected HEp-2 cells with a plasmid directing the expression of MxA cDNA. HEp-2 cells are permissive for many viruses and are unable to express endogenous MxA in response to IFN. Experimental infection with various RNA and DNA viruses revealed that MxA-expressing HEp-2 cells were protected not only against influenza virus and vesicular stomatitis virus (VSV) but also against Semliki Forest virus (SFV), a togavirus with a single-stranded RNA genome of positive polarity. In MxA-transfected cells, viral yields were reduced up to 1,700-fold, and the degree of inhibition correlated well with the expression level of MxA. Furthermore, expression of MxA prevented the accumulation of 49S RNA and 26S RNA, indicating that SFV was inhibited early in its replication cycle. Very similar results were obtained with MxA-transfected cells of the human monocytic cell line U937. The results demonstrate that the antiviral spectrum of MxA is not restricted to negative-strand RNA viruses but also includes SFV, which contains an RNA genome of positive polarity. To test whether MxA protein exerts its inhibitory activity against SFV in the absence of viral structural proteins, we took advantage of a recombinant vector based on the SFV replicon. The vector contains only the coding sequence for the viral nonstructural proteins and the bacterial LacZ gene, which was cloned in place of the viral structural genes. Upon transfection of vector-derived recombinant RNA, expression of the β-galactosidase reporter gene was strongly reduced in the presence of MxA. This finding indicates that viral components other than the structural proteins are the target of MxA action.
A27L Protein Mediates Vaccinia Virus Interaction with Cell Surface Heparan Sulfate
Authors: Chung Che-Sheng , Hsiao Jye-Chian , Chang Yuan-Shau , Chang Wen 
Published: December 2019

ABSTRACT
Vaccinia virus has a wide host range and infects mammalian cells of many different species. This suggests that the cell surface receptors for vaccinia virus are ubiquitously expressed and highly conserved. Alternatively, different receptors are used for vaccinia virus infection of different cell types. Here we report that vaccinia virus binds to heparan sulfate, a glycosaminoglycan (GAG) side chain of cell surface proteoglycans, during virus infection. Soluble heparin specifically inhibits vaccinia virus binding to cells, whereas other GAGs such as condroitin sulfate or dermantan sulfate have no effect. Heparin also blocks infections by cowpox virus, rabbitpox virus, myxoma virus, and Shope fibroma virus, suggesting that cell surface heparan sulfate could be a general mediator of the entry of poxviruses. The biochemical nature of the heparin-blocking effect was investigated. Heparin analogs that have acetyl groups instead of sulfate groups also abolish the inhibitory effect, suggesting that the negative charges on GAGs are important for virus infection. Furthermore, BSC40 cells treated with sodium chlorate to produce undersulfated GAGs are more refractory to vaccinia virus infection. Taken together, the data support the notion that cell surface heparan sulfate is important for vaccinia virus infection. Using heparin-Sepharose beads, we showed that vaccinia virus virions bind to heparin in vitro. In addition, we demonstrated that the recombinant A27L gene product binds to the heparin beads in vitro. This recombinant protein was further shown to bind to cells, and such interaction could be specifically inhibited by soluble heparin. All the data together indicated that A27L protein could be an attachment protein that mediates vaccinia virus binding to cell surface heparan sulfate during viral infection.
CD4
+
-T-Cell and CD20
+
-B-Cell Changes Predict Rapid Disease Progression after Simian-Human Immunodeficiency Virus Infection in Macaques
Authors: Steger Krista K. , Dykhuizen Marta , Mitchen Jacque L. , Hinds Paul W. , Preuninger Brenda L. , Wallace Marianne , Thomson James , Montefiori David C. , Lu Yichen , Pauza C. David 
Published: December 2019

ABSTRACT

Simian-human immunodeficiency virus 89.6PD (SHIV
89.6PD
) was pathogenic after intrarectal inoculation of rhesus macaques. Infection was achieved with a minimum of 2,500 tissue culture infectious doses of cell-free virus stock, and there was no evidence for transient viremia in animals receiving subinfectious doses by the intrarectal route. Some animals experienced rapid progression of disease characterized by loss of greater than 90% of circulating CD4
+
T cells, sustained decreases in CD20
+
B cells, failure to elicit virus-binding antibodies in plasma, and high levels of antigenemia. Slower-progressing animals had moderate but varying losses of CD4
+
T cells; showed increases in circulating CD20
+
B cells; mounted vigorous responses to antibodies in plasma, including neutralizing antibodies; and had low or undetectable levels of antigenemia. Rapid progression led to death within 30 weeks after intrarectal inoculation. Plasma antigenemia at 2 weeks after inoculation (
P
≤ 0.002), B- and T-cell losses (
P
≤ 0.013), and failure to seroconvert (
P
≤ 0.005) were correlated statistically with rapid progression. Correlations were evident by 2 to 4 weeks after intrarectal SHIV inoculation, indicating that early events in the host-pathogen interaction determined the clinical outcome.
The Spike Protein of Murine Coronavirus Mouse Hepatitis Virus Strain A59 Is Not Cleaved in Primary Glial Cells and Primary Hepatocytes
Authors: Hingley Susan T. , Leparc-Goffart Isabelle , Weiss Susan R. 
Published: December 2019

ABSTRACT
Mouse hepatitis virus strain A59 (MHV-A59) produces meningoencephalitis and severe hepatitis during acute infection. Infection of primary cells derived from the central nervous system (CNS) and liver was examined to analyze the interaction of virus with individual cell types derived from the two principal sites of viral replication in vivo. In glial cell cultures derived from C57BL/6 mice, MHV-A59 produces a productive but nonlytic infection, with no evidence of cell-to-cell fusion. In contrast, in continuously cultured cells, this virus produces a lytic infection with extensive formation of syncytia. The observation of few and delayed syncytia following MHV-A59 infection of hepatocytes more closely resembles infection of glial cells than that of continuously cultured cell lines. For MHV-A59, lack of syncytium formation correlates with lack of cleavage of the fusion glycoprotein, or spike (S) protein. The absence of cell-to-cell fusion following infection of both primary cell types prompted us to examine the cleavage of the spike protein. Cleavage of S protein was below the level of detection by Western blot analysis in MHV-A59-infected hepatocytes and glial cells. Furthermore, no cleavage of this protein was detected in liver homogenates from C57BL/6 mice infected with MHV-A59. Thus, cleavage of the spike protein does not seem to be essential for entry and spread of the virus in vivo, as well as for replication in vitro.
Genetic Diversity and Tissue Compartmentalization of the Hepatitis C Virus Genome in Blood Mononuclear Cells, Liver, and Serum from Chronic Hepatitis C Patients
Authors: Navas Sonia , Martín Julio , Quiroga Juan Antonio , Castillo Inmaculada , Carreño Vicente 
Published: December 2019

ABSTRACT

The degree of genetic variability in the hypervariable region 1 of hepatitis C virus (HCV) was analyzed by cloning and sequencing HCV genomes obtained in paired samples of serum, liver tissue, and peripheral blood mononuclear cells (PBMC) from four chronic hepatitis C patients. Genetic variability in serum was higher than in liver tissue or PBMC at the level of complexity (the number of different sequences obtained from each type of tissue) as well as at the level of genetic distance between all pairs of sequences within each tissue (compared by the Student
t
test;
P
< 0.001 for two patients and
P
< 0.01 for another). The spectrum of viral genomes differed among the three types of tissue, as shown by segregation of sequences according to their tissue of origin in phylogenetic analysis and by statistical analysis of mean genetic distances observed between sequences obtained from different tissues (
P
< 0.001), but sequences from liver tissue and PBMC were more closely related to each other than to those from serum.
Mutations in thetatGene Are Responsible for Human Immunodeficiency Virus Type 1 Postintegration Latency in the U1 Cell Line
Authors: Emiliani Stephane , Fischle Wolfgang , Ott Melanie , Van Lint Carine , Amella Carol Ann , Verdin Eric 
Published: December 2019

ABSTRACTPrevious reports have demonstrated that the U1 cell line, a model for postintegration latency, is defective at the level of Tat function and can be rescued by exogenously provided Tat protein. Sequence analysis oftatcDNAs from the U1 cell line identified two distinct forms of Tat, in agreement with the fact that this cell line contains two integrated human immunodeficiency (HIV) proviruses. One Tat cDNA lacked an ATG initiation codon, while the other contained an H-to-L mutation at amino acid 13 (H13→L). BothtatcDNAs were defective in terms of transcriptional activation of long terminal repeat-luciferase reporter gene in transient-transfection experiments. Introduction of the H13→L mutation in a wild-typetatbackground caused a severe reduction in transcriptional activation. Introduction of the same mutation in an infectious HIV molecular clone caused a severely defective phenotype which could be rescued when the HIV proviral DNA was transfected in a Jurkat cell line stably expressing the Tat protein (Jurkat-Tat) or in Jurkat cells treated with tumor necrosis factor alpha. Infectious virus stocks generated in Jurkat-Tat cells were used to infect Jurkat cells and exhibited severely impaired growth which could also be rescued by infecting Jurkat-Tat cells. These observations definetatmutations as a mechanism for HIV postintegration latency.
Protective Immune Response to Foot-and-Mouth Disease Virus with VP1 Expressed in Transgenic Plants
Authors: Carrillo C. , Wigdorovitz A. , Oliveros J. C. , Zamorano P. I. , Sadir A. M. , Gómez N. , Salinas J. , Escribano J. M. , Borca M. V. 
Published: December 2019

ABSTRACT

It has been reported recently that genes encoding antigens of bacterial and viral pathogens can be expressed in plants in a form in which they retain native immunogenic properties. The structural protein VP1 of foot-and-mouth disease virus (FMDV), which has frequently been shown to contain critical epitopes, has been expressed in different vectors and shown to induce virus-neutralizing antibodies and protection in experimental and natural hosts. Here we report the production of transformed plants (
Arabidopsis thaliana
) expressing VP1. Mice immunized with leaf plant extracts elicited specific antibody responses to synthetic peptides representing amino acid residues 135 to 160 of VP1, to VP1 itself, and to intact FMDV particles. Additionally, all of the immunized mice were protected against challenge with virulent FMDV. To our knowledge, this is the first study showing protection against a viral disease by immunization with an antigen expressed in a transgenic plant.
Production and Characterization of Improved Adenovirus Vectors with the E1, E2b, and E3 Genes Deleted
Authors: Amalfitano Andrea , Hauser Michael A. , Hu Huimin , Serra Delila , Begy Catherine R. , Chamberlain Jeffrey S. 
Published: December 2019

ABSTRACTAdenovirus (Ad)-based vectors have great potential for use in the gene therapy of multiple diseases, both genetic and nongenetic. While capable of transducing both dividing and quiescent cells efficiently, Ad vectors have been limited by a number of problems. Most Ad vectors are engineered such that a transgene replaces the Ad E1a, E1b, and E3 genes; subsequently the replication-defective vector can be propagated only in human 293 cells that supply the deleted E1 gene functions intrans. Unfortunately, the use of high titers of E1-deleted vectors has been repeatedly demonstrated to result in low-level expression of viral genes still resident in the vector. In addition, the generation of replication-competent Ad (RCA) by recombination events with the E1 sequences residing in 293 cells further limits the usefulness of E1-deleted Ad vectors. We addressed these problems by isolating new Ad vectors deleted for the E1, E3, and the E2b gene functions. The new vectors can be readily grown to high titers and have several improvements, including an increased carrying capacity and a theoretically decreased risk for generating RCA. We have also demonstrated that the further block to Ad vector replication afforded by the deletion of both the E1 and E2b genes significantly diminished Ad late gene expression in comparison to a conventional E1-deleted vector, without destabilization of the modified vector genome. The results suggested that these modified vectors may be very useful both for in vitro and in vivo gene therapy applications.
In Vitro Infection and Type-Restricted Antibody-Mediated Neutralization of Authentic Human Papillomavirus Type 16
Authors: White Wendy I. , Wilson Susan D. , Bonnez William , Rose Robert C. , Koenig Scott , Suzich Joann A. 
Published: December 2019

ABSTRACT
Human papillomavirus type 16 (HPV-16) is strongly associated with the development of cervical cancer. Studies of model systems with animal papillomaviruses have demonstrated the importance of neutralizing antibodies in preventing papillomavirus-associated disease. The assessment of neutralizing antibody responses against HPV-16, previously hampered by the lack of a viral source, was enabled by the recent propagation of an HPV-16 stock in xenografted severe combined immunodeficiency (SCID) mice. HPV-16 infection of an immortalized human keratinocyte cell line was demonstrated by detection of an HPV-16-specific spliced mRNA amplified by reverse transcriptase PCR. Infection was blocked by preincubation of the virus with antiserum generated against HPV-16 virus-like particles (VLPs) composed of the major capsid protein, L1. To examine potential cross-neutralizing activity among the different genital HPV types, rabbit antisera to L1 VLPs corresponding to HPV-6, -11, -18, -31, -33, -35, -39, and -45 were assayed for the ability to block the HPV-16 infection of cultured cells. Antiserum raised against HPV-33 L1 VLPs was the only heterologous antiserum which inhibited HPV-16 infection. Thus, a neutralization assay for HPV-16 may help to characterize the components required to compose a broadly efficacious genital HPV vaccine.
Attenuation of the Vaccine Oka Strain of Varicella-Zoster Virus and Role of Glycoprotein C in Alphaherpesvirus Virulence Demonstrated in the SCID-hu Mouse
Authors: Moffat Jennifer F. , Zerboni Leigh , Kinchington Paul R. , Grose Charles , Kaneshima Hideto , Arvin Ann M. 
Published: December 2019

ABSTRACT

The SCID-hu mouse implanted with human fetal tissue is a novel model for investigating human viral pathogenesis. Infection of human skin implants was used to investigate the basis for the clinical attenuation of the varicella-zoster virus (VZV) strain, V-Oka, from which the newly licensed vaccine is made. The pathogenicity of V-Oka was compared with that of its parent, P-Oka, another low-passage clinical isolate, strain Schenke (VZV-S), and VZV-Ellen, a standard laboratory strain. The role of glycoprotein C (gC) in infectivity for human skin was assessed by using gC-negative mutants of V-Oka and VZV-Ellen. Whereas all of these VZV strains replicated well in tissue culture, only low-passage clinical isolates were fully virulent in skin, as shown by infectious virus yields and analysis of implant tissues for VZV DNA and viral protein synthesis. The infectivity of V-Oka in skin was impaired compared to that of P-Oka, providing the first evidence of a virologic basis for the clinical attenuation of V-Oka. The infectivity of V-Oka was further diminished in the absence of gC expression. All strains except gC-Ellen retained some capacity to replicate in human skin, but cell-free virus was recovered only from implants infected with P-Oka or VZV-S. Although VZV is closely related to herpes simplex virus type 1 (HSV-1) genetically, experiments in the SCID-hu model revealed differences in tropism for human cells that correlated with differences in VZV and HSV-1 disease. VZV caused extensive infection of epidermal and dermal skin cells, while HSV-1 produced small, superficial lesions restricted to the epidermis. As in VZV, gC expression was a determinant for viral replication in skin. VZV infects human CD4
+
and CD8
+
T cells in thymus/liver implants, but HSV-1 was detected only in epithelial cells, with no evidence of lymphotropism. These SCID-hu mouse experiments show that the clinical attenuation of the varicella vaccine can be attributed to decreased replication of V-Oka in skin and that tissue culture passage alone reduces the ability of VZV to infect human skin in vivo. Furthermore, gC, which is dispensable for replication in tissue culture, plays a critical role in the virulence of the human alphaherpesviruses VZV and HSV-1 for human skin.
Determinants of Human Immunodeficiency Virus Type 1 Resistance to gp41-Derived Inhibitory Peptides
Authors: Rimsky Laurence T. , Shugars Diane C. , Matthews Thomas J. 
Published: December 2019

ABSTRACT

A synthetic peptide, DP178, containing amino acids 127 to 162 of the human immunodeficiency virus type 1 (HIV-1) gp41 Env glycoprotein, is a potent inhibitor of virus infection and virus mediated cell-to-cell fusion (C. Wild, T. Greenwell, and T. Matthews, AIDS Res. Hum. Retroviruses 9:1051–1053, 1993). In an effort to understand the mechanism of action of this peptide, we derived resistant variants of HIV-1
IIIB
and NL4-3 by serial virus passage in the presence of increasing doses of the peptide. Sequence analysis of the resistant isolates suggested that a contiguous 3-amino-acid sequence within the amino-terminal heptad repeat motif of gp41 was associated with resistance. Site-directed mutagenesis studies confirmed this observation and indicated that changes in two of these three residues were necessary for development of the resistant phenotype. Direct binding of DP178 to recombinant protein and synthetic peptide analogs containing the wild-type and mutant heptad repeat sequences revealed a strong correlation between DP178 binding and the biological sensitivity of the corresponding virus isolates to DP178. The results are discussed from the standpoints of the mechanism of action of DP178 and recent crystallographic information for a core structure of the gp41 ectodomain.