bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of the Cytolytic T-Lymphocyte Response to a Candidate Vaccine Strain of Equine Herpesvirus 1 in CBA Mice
Authors: Smith Patrick M. , Zhang Yunfei , Jennings Stephen R. , O’Callaghan Dennis J. 
Published: December 2019

ABSTRACTThe cytolytic T-lymphocyte (CTL) response to respiratory infection with equine herpesvirus 1 (EHV-1) in CBA (H-2k) mice was investigated. Intranasal (i.n.) inoculation of mice with the attenuated EHV-1 strain KyA resulted in the generation of a primary virus-specific CTL response in the draining mediastinal lymph nodes 5 days following infection. EHV-1-specific CTL could be restimulated from the spleen up to 26 weeks after the resolution of infection, indicating that a long-lived memory CTL population was generated. Depletion of CD8+T cells by treatment with antibody and complement prior to assay eliminated CTL activity from both primary and memory populations, indicating that cytolytic activity in this model was mediated by class I major histocompatibility complex-restricted, CD8+T cells. A single i.n. inoculation with KyA induced protective immunity against infection with the pathogenic EHV-1 strain, RacL11. The adoptive transfer of splenocytes from KyA-immune donors into sublethally irradiated recipients resulted in a greater than 250-fold reduction in RacL11 in the lung. The elimination of both CD4+and CD8+T cells from the transferred cells abrogated clearance of RacL11, while the selective depletion of either subpopulation alone had little effect. These results suggested that both lymphocyte subpopulations contribute to viral clearance, with either subpopulation alone being sufficient.
Nonreciprocal Pseudotyping: Murine Leukemia Virus Proteins Cannot Efficiently Package Spleen Necrosis Virus-Based Vector RNA
Authors: Certo Jeanine L. , Shook Betsy F. , Yin Philip D. , Snider John T. , Hu Wei-Shau 
Published: December 2019

ABSTRACTIt has been documented that spleen necrosis virus (SNV) can package murine leukemia virus (MLV) RNA efficiently and propagate MLV vectors to the same titers as it propagates SNV-based vectors. Although the SNV packaging signal (E) and MLV packaging signal (Ψ) have little sequence homology, similar double-hairpin RNA structures were predicted and supported by experimental evidence. To test whether SNV RNA can be packaged by MLV proteins, we modified an SNV vector to be expressed in an MLV-based murine helper cell line. Surprisingly, we found that MLV proteins could not support the replication of SNV vectors. The decrease in titer was approximately 2,000- to 20,000-fold in one round of retroviral replication. RNA analysis revealed that SNV RNA was not efficiently packaged by MLV proteins. RNA hybridization of the cellular and viral RNAs indicated that SNV RNA was packaged at least 25-fold less efficiently than MLV RNA, which was the sensitivity limit of the hybridization assay. The contrast between the MLV and SNV packaging specificity is striking. SNV proteins can recognize both SNV E and MLV Ψ, but MLV can recognize only MLV Ψ. This is the first demonstration of two retroviruses with nonreciprocal packaging specificities.
Virion-Targeted Viral Inactivation of Human Immunodeficiency Virus Type 1 by Using Vpr Fusion Proteins
Authors: Kobinger Gary P. , Borsetti Alessandra , Nie Zilin , Mercier Johanne , Daniel Nesrine , Göttlinger Heinrich G. , Cohen Éric A. 
Published: December 2019

ABSTRACT

Inactivation of progeny virions with chimeric virion-associated proteins represents a novel therapeutic approach against human immunodeficiency virus (HIV) replication. The HIV type 1 (HIV-1) Vpr gene product, which is packaged into virions, is an attractive candidate for such a strategy. In this study, we developed Vpr-based fusion proteins that could be specifically targeted into mature HIV-1 virions to affect their structural organization and/or functional integrity. Two Vpr fusion proteins were constructed by fusing to the first 88 amino acids of HIV-1 Vpr the chloramphenicol acetyltransferase enzyme (VprCAT) or the last 18 C-terminal amino acids of the HIV-1 Vpu protein (VprIE). These Vpr fusion proteins were initially designed to quantify their efficiency of incorporation into HIV-1 virions when produced in
cis
from the provirus. Subsequently, CD4
+
Jurkat T-cell lines constitutively expressing the VprCAT or the VprIE fusion protein were generated with retroviral vectors. Expression of the VprCAT or the VprIE fusion protein in CD4
+
Jurkat T cells did not interfere with cellular viability or growth but conferred substantial resistance to HIV replication. The resistance to HIV replication was more pronounced in Jurkat-VprIE cells than in Jurkat-VprCAT cells. Moreover, the antiviral effect mediated by VprIE was dependent on an intact p6

gag

domain, indicating that the impairment of HIV-1 replication required the specific incorporation of Vpr fusion protein into virions. Gene expression, assembly, or release was not affected upon expression of these Vpr fusion proteins. Indeed, the VprIE and VprCAT fusion proteins were shown to affect the infectivity of progeny virus, since HIV virions containing the VprCAT or the VprIE fusion proteins were, respectively, 2 to 3 times and 10 to 30 times less infectious than the wild-type virus. Overall, this study demonstrated the successful transfer of resistance to HIV replication in tissue cultures by use of Vpr-based antiviral genes.
Modulation of Viral Immunoinflammatory Responses with Cytokine DNA Administered by Different Routes
Authors: Chun Sangjun , Daheshia Massoud , Kuklin Nelly A. , Rouse Barry T. 
Published: December 2019

ABSTRACTThe efficacy of plasmid DNA encoding cytokine administered by different routes, systemic or surface exposure, was evaluated and compared for their modulating effects on subsequent lesions caused by infection with herpes simplex virus (HSV). Systemic or topical administration of both interleukin-4 (IL-4) and IL-10 DNA but not IL-2 DNA caused a long-lasting suppression of HSV-specific delayed-type hypersensitivity response. IL-4 or IL-10 DNA preadministration also modulated the expression of immunoinflammatory lesions associated with corneal infection of HSV. Suppression of ocular lesions required that the DNA be administered to the nasal mucosa or ocular surfaces and was not evident after intramuscular administration. The modulating effect of IL-10 DNA was most evident after topical ocular administration, whereas the effects of IL-4 DNA given by both routes appeared to be equal. Preexposure of IL-4 DNA, but not IL-10 DNA, resulted in a significant change in Th subset balance following HSV infection. Our results indicate that the modulating effect of IL-4 or IL-10 DNA may proceed by different mechanisms. Furthermore, our results suggest that surface administration of cytokine DNA is a convenient means of modulating immunoinflammatory lesions.
Epstein-Barr Virus Uses Different Complexes of Glycoproteins gH and gL To Infect B Lymphocytes and Epithelial Cells
Authors: Wang Xi , Kenyon William J. , Li Qingxue , Müllberg Jürgen , Hutt-Fletcher Lindsey M. 
Published: December 2019

ABSTRACTThe Epstein-Barr virus (EBV) gH-gL complex includes a third glycoprotein, gp42. gp42 binds to HLA class II on the surfaces of B lymphocytes, and this interaction is essential for infection of the B cell. We report here that, in contrast, gp42 is dispensable for infection of epithelial cell line SVKCR2. A soluble form of gp42, gp42.Fc, can, however, inhibit infection of both cell types. Soluble gp42 can interact with EBV gH and gL and can rescue the ability of virus lacking gp42 to transform B cells, suggesting that a gH-gL-gp42.Fc complex can be formed by extrinsic addition of the soluble protein. Truncated forms of gp42.Fc that retain the ability to bind HLA class II but that cannot interact with gH and gL still inhibit B-cell infection by wild-type virus but cannot inhibit infection of SVKCR2 cells or rescue the ability of recombinant gp42-negative virus to transform B cells. An analysis of wild-type virions indicates the presence of more gH and gL than gp42. To explain these results, we describe a model in which wild-type EBV virions are proposed to contain two types of gH-gL complexes, one that includes gp42 and one that does not. We further propose that these two forms of the complex have mutually exclusive abilities to mediate the infection of B cells and epithelial cells. Conversion of one to the other concurrently alters the ability of virus to infect each cell type. The model also suggests that epithelial cells may express a molecule that serves the same cofactor function for this cell type as HLA class II does for B cells and that the gH-gL complex interacts directly with this putative epithelial cofactor.
The Duck Hepatitis B Virus Polymerase Is Activated by Its RNA Packaging Signal, ɛ
Authors: Tavis John E. , Massey Brandon , Gong Yunhao 
Published: December 2019

ABSTRACT

The ɛ stem-loop at the 5′ end of the pregenomic RNA of the hepatitis B viruses is both the primary element of the RNA packaging signal and the origin of reverse transcription. We have previously presented evidence for a third essential role for ɛ, that of an essential cofactor in the maturation of the viral polymerase (J. E. Tavis and D. Ganem, J. Virol. 70:5741–5750, 1996). In this case, binding of ɛ to the polymerase is proposed to induce a physical alteration to the polymerase that is needed for it to develop enzymatic activity. Three lines of evidence employing duck hepatitis B virus supporting this hypothesis are presented here. First, an unusual DNA polymerase activity employing exogenous RNAs (the
trans
reaction) that was originally discovered with recombinant duck hepatitis B virus polymerase expressed in
Saccharomyces cerevisiae
yeasts was shown to be an authentic property of the viral polymerase. The
trans
reaction was found to be template-dependent reverse transcription of the exogenous RNA. The
trans
reaction occurred independently of the hepadnavirus protein-priming mechanism, yet it was still strongly stimulated by ɛ. This directly demonstrates a role for ɛ in activation of the polymerase. Second, the reverse transcriptase domain of the polymerase was shown to be physically altered following binding to ɛ, as would be expected if the alteration was required for maturation of the polymerase to an enzymatically active form. Finally, analysis of 15 mutations throughout the duck hepatitis B virus polymerase demonstrated that the ɛ-dependent alteration to the polymerase was a prerequisite for DNA priming, reverse transcription, and the
trans
reaction.
Role of the SH3-Ligand Domain of Simian Immunodeficiency Virus Nef in Interaction with Nef-Associated Kinase and Simian AIDS in Rhesus Macaques
Authors: Khan Imran H. , Sawai Earl T. , Antonio Erwin , Weber Claudia J. , Mandell Carol P. , Montbriand Phillip , Luciw Paul A. 
Published: December 2019

ABSTRACT

The
nef
gene of the human and simian immunodeficiency viruses (HIV and SIV) is dispensable for viral replication in T-cell lines; however, it is essential for high virus loads and progression to simian AIDS (SAIDS) in SIV-infected adult rhesus macaques. Nef proteins from HIV type 1 (HIV-1), HIV-2, and SIV contain a proline-Xaa-Xaa-proline (PxxP) motif. The region of Nef with this motif is similar to the Src homology region 3 (SH3) ligand domain found in many cell signaling proteins. In virus-infected lymphoid cells, Nef interacts with a cellular serine/threonine kinase, designated Nef-associated kinase (NAK). In this study, analysis of viral clones containing point mutations in the
nef
gene of the pathogenic clone SIVmac239 revealed that several strictly conserved residues in the PxxP region were essential for Nef-NAK interaction. The results of this analysis of Nef mutations in in vitro kinase assays indicated that the PxxP region in SIV Nef was strikingly similar to the consensus sequence for SH3 ligand domains possessing the minus orientation. To test the significance of the PxxP motif of Nef for viral pathogenesis, each proline was mutated to an alanine to produce the viral clone SIVmac239-P
104
A/P
107
A. This clone, expressing Nef that does not associate with NAK, was inoculated into seven juvenile rhesus macaques. In vitro kinase assays were performed on virus recovered from each animal; the ability of Nef to associate with NAK was restored in five of these animals as early as 8 weeks after infection. Analysis of
nef
genes from these viruses revealed patterns of genotypic reversion in the mutated PxxP motif. These revertant genotypes, which included a second-site suppressor mutation, restored the ability of Nef to interact with NAK. Additionally, the proportion of revertant viruses increased progressively during the course of infection in these animals, and two of these animals developed fatal SAIDS. Taken together, these results demonstrated that in vivo selection for the ability of SIV Nef to associate with NAK was correlated with the induction of SAIDS. Accordingly, these studies implicate a role for the conserved SH3 ligand domain for Nef function in virally induced immunodeficiency.
Genes Required for Replication of the 15.5-Kilobase RNA Genome of a Plant Closterovirus
Authors: Peremyslov Valery V. , Hagiwara Yuka , Dolja Valerian V. 
Published: December 2019

ABSTRACT
A full-length cDNA clone of beet yellows closterovirus (BYV) was engineered and used to map functions involved in the replication of the viral RNA genome and subgenomic RNA formation. Among 10 open reading frames (ORFs) present in BYV, ORFs 1a and 1b suffice for RNA replication and transcription. The proteins encoded in these ORFs harbor putative methyltransferase, RNA helicase, and RNA polymerase domains common to Sindbis virus-like viruses and a large interdomain region that is unique to closteroviruses. The papain-like leader proteinase (L-Pro) encoded in the 5′-proximal region of ORF 1a was found to have a dual function in genome amplification. First, the autocatalytic cleavage between L-Pro and the remainder of the ORF 1a product was essential for replication of RNA. Second, an additional L-Pro function that was separable from proteolytic activity was required for efficient RNA accumulation. The deletion of a large, ∼5.6-kb, 3′-terminal region coding for a 6-kDa hydrophobic protein, an HSP70 homolog, a 64-kDa protein, minor and major capsid proteins, a 20-kDa protein, and a 21-kDa protein (p21) resulted in replication-competent RNA. However, examination of mutants with replacements of start codons in each of these seven 3′-terminal ORFs revealed that p21 functions as an enhancer of genome amplification. The intriguing analogies between the genome organization and replicational requirements of plant closteroviruses and animal coronavirus-like viruses are discussed.
Replication Defect of Moloney Murine Leukemia Virus with a Mutant Reverse Transcriptase That Can Incorporate Ribonucleotides and Deoxyribonucleotides
Authors: Gao Guangxia , Goff Stephen P. 
Published: December 2019

ABSTRACT
Reverse transcriptase (RT) plays a critical role in retrovirus replication, directing the synthesis of a double- stranded DNA copy of the viral RNA genome. We have previously described a mutant RT of the Moloney murine leukemia virus in which F155 was replaced by valine, and we demonstrated that this substitution allowed the enzyme to incorporate ribonucleotides to form RNA while still retaining its normal ability to incorporate deoxyribonucleotides to form DNA. When introduced into the viral genome, this mutation rendered the virus incapable of replication. Characterization of the mutant virus revealed that the enzyme was still active and able to synthesize minus-strand strong stop DNA and some longer products but failed to make full-length minus-strand DNA. We propose that the failure of the enzyme to complete DNA synthesis in vivo resulted from its ability to incorporate ribonucleotides into the products, which served as inhibitors for DNA synthesis. We also tested seven other amino acid residues for their abilities to substitute for F155 in virus replication; of these, only tyrosine could support virus replication. In an attempt to select for second-site suppressor mutations, the F155V mutant was subjected to random mutagenesis and was used as a parent for the isolation of revertant viruses. Two independent revertants were found to have changed the valine residue at position 155 back to the wild- type phenylalanine. These results suggest that an aromatic ring at this position is important for virus replication.
Functional Replacement of the Intracellular Region of the Notch1 Receptor by Epstein-Barr Virus Nuclear Antigen 2
Authors: Sakai Takashi , Taniguchi Yoshihito , Tamura Kumiko , Minoguchi Shigeru , Fukuhara Takataro , Strobl Lothar J. , Zimber-Strobl Ursula , Bornkamm George W. , Honjo Tasuku 
Published: December 2019

ABSTRACT

The intracellular region (RAMIC) of the mouse Notch1 receptor interacts with RBP-J/CBF-1, which binds to the DNA sequence CGTGGGAA and suppresses differentiation by transcriptional activation of genes regulated by RBP-J. Epstein-Barr virus nuclear antigen 2 (EBNA2) is essential for immortalization of human B cells by the virus. EBNA2 is a pleiotropic activator of viral and cellular genes and is targeted to DNA at least in part by interacting with RBP-J. We found that EBNA2 and the Notch1 RAMIC compete for binding to RBP-J, indicating that their interaction sites on RBP-J overlap at least partially. EBNA2 and Notch1 RAMIC transactivated the same set of viral and host promoters, i.e., the EBNA2 response element of the Epstein-Barr virus
TP1
and the
HES-1
promoter. Furthermore, EBNA2 functionally replaced the Notch1 RAMIC by suppressing differentiation of C2C12 myoblast progenitor cells.
U
L
27.5 Is a Novel γ
2
Gene Antisense to the Herpes Simplex Virus 1 Gene Encoding Glycoprotein B
Authors: Chang Yijan E. , Menotti Laura , Filatov Felix , Campadelli-Fiume Gabriella , Roizman Bernard 
Published: December 2019

ABSTRACT

An antibody made against the herpes simplex virus 1 U
S
5 gene predicted to encode glycoprotein J was found to react strongly with two proteins, one with an apparent
M
r
of 23,000 and mapping in the S component and one with a herpes simplex virus protein with an apparent
M
r
of 43,000. The antibody also reacted with herpes simplex virus type 2 proteins forming several bands with apparent
M
r
s ranging from 43,000 to 50,000. Mapping studies based on intertypic recombinants, analyses of deletion mutants, and ultimately, reaction of the antibody with a chimeric protein expressed by in-frame fusion of the glutathione
S
-transferase gene to an open reading frame antisense to the gene encoding glycoprotein B led to the definitive identification of the new open reading frame, designated U
L
27.5. Sequence analyses indicate the conservation of a short amino acid sequence common to U
S
5 and U
L
27.5. The coding sequence of the herpes simplex virus U
L
27.5 open reading frame is strongly homologous to the sequence encoding the carboxyl terminus of the herpes simplex virus 2 U
L
27.5 sequence. However, both open reading frames could encode proteins predicted to be significantly larger than the mature U
L
27.5 proteins accumulating in the infected cells, indicating that these are either processed posttranslationally or synthesized from alternate, nonmethionine-initiating codons. The U
L
27.5 gene expression is blocked by phosphonoacetate, indicating that it is a γ
2
gene. The product accumulated predominantly in the cytoplasm. U
L
27.5 is the third open reading frame found to map totally antisense to another gene and suggests that additional genes mapping antisense to known genes may exist.
T-Cell Response to Woodchuck Hepatitis Virus (WHV) Antigens during Acute Self-Limited WHV Infection and Convalescence and after Viral Challenge
Authors: Menne Stephan , Maschke Jan , Lu Mengji , Grosse-Wilde Hans , Roggendorf Michael 
Published: December 2019

ABSTRACTThe infection of woodchucks with woodchuck hepatitis virus (WHV) provides an experimental model to study early immune responses during hepadnavirus infection that cannot be tested in patients. The T-cell response of experimentally WHV-infected woodchucks to WHsAg, rWHcAg, and WHcAg peptides was monitored by observing 5-bromo-2′-deoxyuridine and [2-3H]adenine incorporation. The first T-cell responses were directed against WHsAg 3 weeks after infection; these were followed by responses to rWHcAg including the immunodominant T-cell epitope of WHcAg (amino acids 97 to 110). Maximal proliferative responses were detected when the animals seroconvered to anti-WHs and anti-WHc (week 6). A decrease in the T-cell response to viral antigens coincided with clearance of viral DNA. Polyclonal rWHcAg-specific T-cell lines were established 6, 12, 18, and 24 weeks postinfection, and their responses to WHcAg peptides were assessed. Five to seven peptides including the immunodominant epitope were recognized throughout the observation period (6 months). At 12 months after infection, T-cell responses to antigens and peptides were not detected. Reactivation of T-cell responses to viral antigens and peptides occurred within 7 days after challenge of animals with WHV. These results demonstrate that a fast and vigorous T-cell response to WHsAg, rWHcAg, and amino acids 97 to 110 of the WHcAg occurs within 3 weeks after WHV infection. The peak of this response was associated with viral clearance and may be crucial for recovery from infection. One year after infection, no proliferation of T cells in response to antigens was observed; however, the WHV-specific T-cell response was reactivated after challenge of woodchucks with WHV and may be responsible for protection against WHV reinfection.
Evolution of Hypervariable Region 1 of Hepatitis C Virus in Primary Infection
Authors: Manzin Aldo , Solforosi Laura , Petrelli Enzo , Macarri Giampiero , Tosone Grazia , Piazza Marcello , Clementi Massimo 
Published: December 2019

ABSTRACT

The hypervariable region 1 (HVR-1) of the putative envelope encoding E2 region of hepatitis C virus (HCV) RNA was analyzed in sequential samples from three patients with acute type C hepatitis infected from different sources to address (i) the dynamics of intrahost HCV variability during the primary infection and (ii) the role of host selective pressure in driving viral genetic evolution. HVR-1 sequences from 20 clones per each point in time were analyzed after amplification, cloning, and purification of plasmid DNA from single colonies of transformed cells. The intrasample evolutionary analysis (nonsynonymous mutations per nonsynonymous site [

K
a

], synonymous mutations per synonymous site [

K
s

],

K
a

/

K
s

ratio, and genetic distances [gd]) documented low gd in early samples (ranging from 2.11 to 7.79%) and a further decrease after seroconversion (from 0 to 4.80%), suggesting that primary HCV infection is an oligoclonal event, and found different levels and dynamics of host pressure in the three cases. The intersample analysis (pairwise comparisons of intrapatient sequences; r

K
a

, r

K
s

, r

K
a

/r

K
s

ratio, and gd) confirmed the individual features of HCV genetic evolution in the three subjects and pointed to the relative contribution of either neutral evolution or selective forces in driving viral variability, documenting that adaptation of HCV for persistence in vivo follows different routes, probably representing the molecular counterpart of the viral fitness for individual environments.