bluuebunny/crossref_metadata_2025_split huggingface dataset:
Membrane-Associated Heparan Sulfate Proteoglycan Is a Receptor for Adeno-Associated Virus Type 2 Virions
Authors: Summerford Candace , Samulski Richard Jude 
Published: December 2019

ABSTRACTThe human parvovirus adeno-associated virus (AAV) infects a broad range of cell types, including human, nonhuman primate, canine, murine, and avian. Although little is known about the initial events of virus infection, AAV is currently being developed as a vector for human gene therapy. Using defined mutant CHO cell lines and standard biochemical assays, we demonstrate that heparan sulfate proteoglycans mediate both AAV attachment to and infection of target cells. Competition experiments using heparin, a soluble receptor analog, demonstrated dose-dependent inhibition of AAV attachment and infection. Enzymatic removal of heparan but not chondroitin sulfate moieties from the cell surface greatly reduced AAV attachment and infectivity. Finally, mutant cell lines that do not produce heparan sulfate proteoglycans were significantly impaired for both AAV binding and infection. This is the first report that proteoglycan has a role in cellular attachment of a parvovirus. Together, these results demonstrate that membrane-associated heparan sulfate proteoglycan serves as the viral receptor for AAV type 2, and provide an explanation for the broad host range of AAV. Identification of heparan sulfate proteoglycan as a viral receptor should facilitate development of new reagents for virus purification and provide critical information on the use of AAV as a gene therapy vector.
Immunization of Pigs with a Particle-Mediated DNA Vaccine to Influenza A Virus Protects against Challenge with Homologous Virus
Authors: Macklin Michael D. , McCabe Dennis , McGregor Martha W. , Neumann Veronica , Meyer Todd , Callan Robert , Hinshaw Virginia S. , Swain William F. 
Published: December 2019

ABSTRACTParticle-mediated delivery of a DNA expression vector encoding the hemagglutinin (HA) of an H1N1 influenza virus (A/Swine/Indiana/1726/88) to porcine epidermis elicits a humoral immune response and accelerates the clearance of virus in pigs following a homotypic challenge. Mucosal administration of the HA expression plasmid elicits an immune response that is qualitatively different than that elicited by the epidermal vaccination in terms of inhibition of the initial virus infection. In contrast, delivery of a plasmid encoding an influenza virus nucleoprotein from A/PR/8/34 (H1N1) to the epidermis elicits a strong humoral response but no detectable protection in terms of nasal virus shed. The efficacy of the HA DNA vaccine was compared with that of a commercially available inactivated whole-virus vaccine as well as with the level of immunity afforded by previous infection. The HA DNA and inactivated viral vaccines elicited similar protection in that initial infection was not prevented, but subsequent amplification of the infection is limited, resulting in early clearance of the virus. Convalescent animals which recovered from exposure to virulent swine influenza virus were completely resistant to infection when challenged. The porcine influenza A virus system is a relevant preclinical model for humans in terms of both disease and gene transfer to the epidermis and thus provides a basis for advancing the development of DNA-based vaccines.
Foamy Virus Particle Formation
Authors: Fischer Nicole , Heinkelein Martin , Lindemann Dirk , Enssle Jörg , Baum Christopher , Werder Evelyn , Zentgraf Hanswalter , Müller Justus G. , Rethwilm Axel 
Published: December 2019

ABSTRACT

Subgenomic expression plasmids for the so-called human foamy virus (HFV) structural gag, gag/pol, and env genes were constructed and used to analyze foamy virus particle formation by electron microscopy. Expression of an R-U5-
gag-pol
construct under control of the human cytomegalovirus immediate-early enhancer-promoter resulted in the formation of viral cores with a homogeneous size of approximately 50 nm located in the cytoplasm. Upon coexpression of an envelope construct, particles were observed budding into cytoplasmic vesicles and from the plasma membrane. Expression of the Gag protein precursor pr74 alone led to aberrantly formed viral particles of heterogeneous size and with open cores. Normal-shaped cores were seen after transfection of a construct expressing the p70

gag

cleavage product, indicating that p70

gag

is able to assemble into capsids. Coexpression of p70

gag

and Env resulted in budding virions, ruling out a requirement of the reverse transcriptase for capsid or virion formation. In sharp contrast to other retroviruses, the HFV cores did not spontaneously bud from cellular membranes. Radiochemical labeling followed by protein gel electrophoresis also revealed the intracellular retention of Env-deprived HFV capsids.
Use of Murine CXCR-4 as a Second Receptor by Some T-Cell-Tropic Human Immunodeficiency Viruses
Authors: Parolin Cristina , Borsetti Alessandra , Choe Hyeryun , Farzan Michael , Kolchinsky Peter , Heesen Michael , Ma Qing , Gerard Craig , Palú Giorgio , Dorf Martin E. , Springer Timothy , Sodroski Joseph 
Published: December 2019

ABSTRACTThe human CXCR-4 molecule serves as a second receptor for primary, T-cell-tropic, and laboratory-adapted human immunodeficiency virus type 1 (HIV-1) isolates. Here we show that murine CXCR-4 can support the entry of some of these HIV-1 isolates. Differences between mouse and human CXCR-4 in the ability to function as an HIV-1 receptor are determined by sequences in the second extracellular loop of the CXCR-4 protein.
Non-AUG Translation Initiation in a Plant RNA Virus: a Forty-Amino-Acid Extension Is Added to the N Terminus of the Soil-Borne Wheat Mosaic Virus Capsid Protein
Authors: Shirako Yukio 
Published: December 2019

ABSTRACTRNA 2 of soil-borne wheat mosaic virus (SBWMV), the type species of the genusFurovirus, encodes a protein previously hypothesized to be initiated at an in-frame non-AUG codon upstream of the AUG initiation codon (nucleotide positions 334 to 336) for the 19-kDa capsid protein. Site-directed mutagenesis and in vitro transcription and translation analysis indicated that CUG (nucleotides 214 to 216) is the initiation codon for a protein with a calculated molecular mass of 25 kDa composed of a 40-amino-acid extension to the N terminus of the 19-kDa capsid protein. A stable deletion mutant, which was isolated after extensive passages of a wild-type SBWMV, contained a mixture of two deleted RNA 2’s, only one of which coded for the 25-kDa protein. The amino acid sequence of the N-terminal extension was moderately conserved and the CUG initiation codon was preserved among three SBWMV isolates from Japan and the United States. This amino acid sequence conservation, as well as the retention of expression of the 25-kDa protein in the stable deletion mutant, suggests that the 25-kDa protein is functional in the life cycle of SBWMV. This is the first report of a non-AUG translation initiation in a plant RNA virus genome.
African Swine Fever Virus Infection in the Argasid Host,
Ornithodoros porcinus porcinus
Authors: Kleiboeker S. B. , Burrage T. G. , Scoles G. A. , Fish D. , Rock D. L. 
Published: December 2019

ABSTRACT

The pathogenesis of African swine fever virus (ASFV) infection in
Ornithodoros porcinus porcinus
was examined in nymphal ticks infected with the ASFV isolate Chiredzi/83/1. At times postinfection (p.i.) ranging from 6 h to 290 days, ticks or dissected tick tissues were titrated for virus and examined ultrastructurally for evidence of virus replication. The ASFV infection rate in ticks was 100% in these experiments, and virus infection was not associated with a significant increase in tick mortality. Initial ASFV replication occurred in phagocytic digestive cells of the midgut epithelium. Subsequent infection and replication of ASFV in undifferentiated midgut cells was observed at 15 days p.i. Generalization of virus infection from midgut to other tick tissues required 2 to 3 weeks and most likely involved virus movement across the basal lamina of the midgut into the hemocoel. Secondary sites of virus replication included hemocytes (type I and II), connective tissue, coxal gland, salivary gland, and reproductive tissue. Virus replication was not observed in the nervous tissue of the synganglion, Malpighian tubules, and muscle. Persistent infection, characterized by active virus replication, was observed for all involved tick tissues. After 91 days p.i., viral titers in salivary gland and reproductive tissue were consistently the highest detected. Successful tick-to-pig transmission of ASFV at 48 days p.i. correlated with high viral titers in salivary and coxal gland tissue and their secretions. A similar pattern of virus infection and persistence in
O. porcinus porcinus
was observed for three additional ASFV tick isolates in their associated ticks.
Eukaryotic Elongation Factor 1δ Is Hyperphosphorylated by the Protein Kinase Encoded by the U
L
13 Gene of Herpes Simplex Virus 1
Authors: Kawaguchi Yasushi , Van Sant Charles , Roizman Bernard 
Published: December 2019

ABSTRACT

The translation elongation factor 1δ (EF-1δ) consists of two forms, a hypophosphorylated form (apparent
M
r
, 38,000) and a hyperphosphorylated form (apparent
M
r
, 40,000). Earlier Y. Kawaguchi, R. Bruni, and B. Roizman (J. Virol. 71:1019–1024, 1997) reported that whereas mock-infected cells accumulate the hypophosphorylated form, the hyperphosphorylated form of EF-1δ accumulates in cells infected with herpes simplex virus 1. We now report that the accumulation of the hyperphosphorylated EF-1δ is due to phosphorylation by U
L
13 protein kinase based on the following observations. (i) The relative amounts of hypo- and hyperphosphorylated EF-1δ in Vero cells infected with mutant virus lacking the U
L
13 gene could not be differentiated from those of mock-infected cells. In contrast, the hyperphosphorylated EF-1δ was the predominant form in Vero cells infected with wild-type viruses, a recombinant virus in which the deleted U
L
13 sequences were restored, or with a virus lacking the U
S
3 gene, which also encodes a protein kinase. (ii) The absence of the hyperphosphorylated EF-1δ in cells infected with the U
L
13 deletion mutant was not due to failure of posttranslational modification of infected-cell protein 22 (ICP22)/U
S
1.5 or of interaction with ICP0, inasmuch as preferential accumulation of hyperphosphorylated EF-1δ was observed in cells infected with viruses from which the genes encoding ICP22/U
S
1.5 or ICP0 had been deleted. (iii) Both forms of EF-1δ were labeled by
32
P
i
in vivo, but the prevalence of the hyperphosphorylated EF-1δ was dependent on the presence of the U
L
13 protein. (iv) EF-1δ immunoprecipitated from uninfected Vero cells was phosphorylated by U
L
13 precipitated by the anti-U
L
13 antibody from lysates of wild-type virus-infected cells, but not by complexes formed by the interaction of the U
L
13 antibody with lysates of cells infected with a mutant lacking the U
L
13 gene. This is the first evidence that a viral protein kinase targets a cellular protein. Together with evidence that ICP0 also interacts with EF-1δ reported in the paper cited above, these data indicate that herpes simplex virus 1 has evolved a complex strategy for optimization of infected-cell protein synthesis.
Neutralization Sensitivity of Human Immunodeficiency Virus Type 1 Primary Isolates to Antibodies and CD4-Based Reagents Is Independent of Coreceptor Usage
Authors: Trkola Alexandra , Ketas Tom , KewalRamani Vineet N. , Endorf Fred , Binley James M. , Katinger Hermann , Robinson Jim , Littman Dan R. , Moore John P. 
Published: December 2019

ABSTRACT

We have investigated whether the identity of the coreceptor (CCR5, CXCR4, or both) used by primary human immunodeficiency virus type 1 (HIV-1) isolates to enter CD4
+
cells influences the sensitivity of these isolates to neutralization by monoclonal antibodies and CD4-based agents. Coreceptor usage was not an important determinant of neutralization titer for primary isolates in peripheral blood mononuclear cells. We also studied whether dualtropic primary isolates (able to use both CCR5 and CXCR4) were differentially sensitive to neutralization by the same antibodies when entering U87MG-CD4 cells stably expressing either CCR5 or CXCR4. Again, we found that the coreceptor used by a virus did not greatly affect its neutralization sensitivity. Similar results were obtained for CCR5- or CXCR4-expressing HOS cell lines engineered to express green fluorescent protein as a reporter of HIV-1 entry. Neutralizing antibodies are therefore unlikely to be the major selection pressure which drives the phenotypic evolution (change in coreceptor usage) of HIV-1 that can occur in vivo. In addition, the increase in neutralization sensitivity found when primary isolates adapt to growth in transformed cell lines in vitro has little to do with alterations in coreceptor usage.
The Genome Sequence of Herpes Simplex Virus Type 2
Authors: Dolan Aidan , Jamieson Fiona E. , Cunningham Charles , Barnett Barbara C. , McGeoch Duncan J. 
Published: December 2019

ABSTRACTThe genomic DNA sequence of herpes simplex virus type 2 (HSV-2) strain HG52 was determined as 154,746 bp with a G+C content of 70.4%. A total of 74 genes encoding distinct proteins was identified; three of these were each present in two copies, within major repeat elements of the genome. The HSV-2 gene set corresponds closely with that of HSV-1, and the HSV-2 sequence prompted several local revisions to the published HSV-1 sequence (D. J. McGeoch, M. A. Dalrymple, A. J. Davison, A. Dolan, M. C. Frame, D. McNab, L. J. Perry, J. E. Scott, and P. Taylor, J. Gen. Virol. 69:1531–1574, 1988). No compelling evidence for the existence of any additional protein-coding genes in HSV-2 was identified.
In Vitro and In Vivo Biology of Recombinant Adenovirus Vectors with E1, E1/E2A, or E1/E4 Deleted
Authors: Lusky M. , Christ M. , Rittner K. , Dieterle A. , Dreyer D. , Mourot B. , Schultz H. , Stoeckel F. , Pavirani A. , Mehtali M. 
Published: December 2019

ABSTRACT
Isogenic, E3-deleted adenovirus vectors defective in E1, E1 and E2A, or E1 and E4 were generated in complementation cell lines expressing E1, E1 and E2A, or E1 and E4 and characterized in vitro and in vivo. In the absence of complementation, deletion of both E1 and E2A completely abolished expression of early and late viral genes, while deletion of E1 and E4 impaired expression of viral genes, although at a lower level than the E1/E2A deletion. The in vivo persistence of these three types of vectors was monitored in selected strains of mice with viral genomes devoid of transgenes to exclude any interference by immunogenic transgene-encoded products. Our studies showed no significant differences among the vectors in the short-term maintenance and long-term (4-month) persistence of viral DNA in liver and lung cells of immunocompetent and immunodeficient mice. Furthermore, all vectors induced similar antibody responses and comparable levels of adenovirus-specific cytotoxic T lymphocytes. These results suggest that in the absence of transgenes, the progressive deletion of the adenovirus genome does not extend the in vivo persistence of the transduced cells and does not reduce the antivirus immune response. In addition, our data confirm that, in the absence of transgene expression, mouse cellular immunity to viral antigens plays a minor role in the progressive elimination of the virus genome.
p53 and RPA Are Sequestered in Viral Replication Centers in the Nuclei of Cells Infected with Human Cytomegalovirus
Authors: Fortunato Elizabeth A. , Spector Deborah H. 
Published: December 2019

ABSTRACT
Previously, we reported that human cytomegalovirus (HCMV) infection of fibroblasts markedly affects p53 and other regulatory proteins and inhibits transit through the cell cycle (F. M. Jault, J.-M. Jault, F. Ruchti, E. A. Fortunato, C. Clark, J. Corbeil, D. D. Richman, and D. H. Spector, J. Virol. 69:6697–6704, 1995). Although the p53 steady-state levels are elevated throughout the infection, evidence suggests that the ability of p53 to transactivate some of its downstream targets is compromised. To elucidate the mechanisms governing the accumulation of p53, we examined the synthesis, stability, and localization of the protein in HCMV-infected fibroblasts. Synthesis of p53 was not increased in the infected cells during the first 24 h postinfection. In fact, pulse-chase experiments revealed that synthesis of p53 in infected fibroblasts was lower than in mock-infected cells. However, after an initial decay, the p53 was stabilized. In addition, beginning at approximately 30 h postinfection, p53 was localized to discrete foci within the nuclei of infected cells. The morphology of these foci suggested that they were replication centers. We confirmed that these are sites of DNA replication by demonstrating both incorporation of bromodeoxyuridine and localization of UL44 (the viral polymerase processivity factor) into these centers. The single-stranded DNA binding protein RPA was also sequestered. In contrast, Rb and HCMV IE1 72 remained distributed throughout the infected cell nuclei, indicating specific targeting of certain proteins. Taken together, our results provide two alternative mechanisms to account for the increased steady-state levels of p53 observed in HCMV-infected fibroblasts.
A Retention Signal Necessary and Sufficient for Endoplasmic Reticulum Localization Maps to the Transmembrane Domain of Hepatitis C Virus Glycoprotein E2
Authors: Cocquerel Laurence , Meunier Jean-Christophe , Pillez André , Wychowski Czeslaw , Dubuisson Jean 
Published: December 2019

ABSTRACT
The hepatitis C virus (HCV) genome encodes two envelope glycoproteins (E1 and E2). These glycoproteins interact to form a noncovalent heterodimeric complex which is retained in the endoplasmic reticulum (ER). To identify whether E1 and/or E2 contains an ER-targeting signal potentially involved in ER retention of the E1-E2 complex, these proteins were expressed alone and their intracellular localization was studied. Due to misfolding of E1 in the absence of E2, no conclusion on the localization of its native form could be drawn from the expression of E1 alone. E2 expressed in the absence of E1 was shown to be retained in the ER similarly to E1-E2 complex. Chimeric proteins in which E2 domains were exchanged with corresponding domains of a protein normally transported to the plasma membrane (CD4) were constructed to identify the sequence responsible for its ER retention. The transmembrane domain (TMD) of E2 (C-terminal 29 amino acids) was shown to be sufficient for retention of the ectodomain of CD4 in the ER compartment. Replacement of the E2 TMD by the anchor signal of CD4 or a glycosyl phosphatidylinositol (GPI) moiety led to its expression on the cell surface. In addition, replacement of the E2 TMD by the anchor signal of CD4 or a GPI moiety abolished the formation of E1-E2 complexes. Together, these results suggest that, besides having a role as a membrane anchor, the TMD of E2 is involved in both complex formation and intracellular localization.
A 2.9-Kilobase Noncoding Nuclear RNA Functions in the Establishment of Persistent Hz-1 Viral Infection
Authors: Chao Yu-Chan , Lee Song-Tay , Chang Ming-Chuan , Chen Hong-Hwa , Chen Shih-Shun , Wu Tzong-Yuan , Liu Fu-Hwa , Hsu Err-Lieh , Hou Roger F. 
Published: December 2019

ABSTRACT

Differential viral gene expression during both productive and persistent infections of Hz-1 virus in insect cells was elucidated. Despite more than 100 viral transcripts being expressed during productive viral infection, massive viral gene shutoff was observed during viral persistency, leaving the 2.9-kb persistence-associated transcript 1 (PAT1) as the only detectable viral RNA. Persistence-associated gene 1 (
pag1
), which encodes PAT1, was cloned and found to contain no significant open reading frames. PAT1 is not associated with the cellular translation machinery and is located exclusively in the nucleus. Further experiments showed that PAT1 is functional in the establishment of persistent Hz-1 viral infection in the cells. All the evidence collectively indicates that PAT1 is a novel nuclear transcript of viral origin. Our results showed that although PAT1 and
XIST
RNA, a mammalian X-inactive specific transcript, are transcribed by different genes, they have interesting similarities.
Distinct Pathways for Tumor Necrosis Factor Alpha and Ceramides in Human Cytomegalovirus Infection
Authors: Allan-Yorke Justine , Record Michel , de Préval Claude , Davrinche Christian , Davignon Jean-Luc 
Published: December 2019

ABSTRACTHuman cytomegalovirus (HCMV) infection can be fatal to immunocompromised individuals. We have previously reported that gamma interferon and tumor necrosis factor alpha (TNF-α) synergistically inhibit HCMV replication in vitro. Ceramides have been described as second messengers induced by TNF-α. To investigate the mechanisms involved in the inhibition of HCMV by TNF-α, in the present study we have analyzed ceramide production by U373 MG astrocytoma cells and the effects of TNF-α versus ceramides on HCMV replication. Our results show that U373 MG cells did not produce ceramides upon incubation with TNF-α. Moreover, long-chain ceramides induced by treatment with exogenous bacterial sphingomyelinase inhibited HCMV replication in synergy with TNF-α. Surprisingly, short-chain permeant C6-ceramide increased viral replication. Our results show that the anti-HCMV activity of TNF-α is independent of ceramides. In addition, our results suggest that TNF-α and endogenous long-chain ceramides use separate pathways of cell signalling to inhibit HCMV replication, while permeant C6-ceramide appears to activate a third pathway leading to an opposite effect.
Anchorage-Independent Transcription of the Cyclin A Gene Induced by the E7 Oncoprotein of Human Papillomavirus Type 16
Authors: Schulze Almut , Mannhardt Boris , Zerfass-Thome Karin , Zwerschke Werner , Jansen-Dürr Pidder 
Published: December 2019

ABSTRACT
To develop an experimental model for E7-mediated anchorage-independent growth, we studied the ability of E7-expressing NIH 3T3 subclones to enter S phase when they were cultured in suspension. We found that expression of E7 prevents the inhibition of cyclin E-associated kinase and also triggers activation of cyclin A gene expression in suspension cells. A point mutation in the amino terminus of E7 prevented E7-driven rescue of cyclin E-associated kinase activity in suspension cells; however, cells with this mutation retained some ability to activate cyclin A gene expression and promote S-phase entry. Activation of cyclin A gene expression by E7 was correlated with an increased binding of free E2F to a regulatory element in the cyclin A promoter which mediates both repression of cyclin A upon loss of adhesion and its reactivation by E7. Surprisingly, expression of E7 led to a nuclear accumulation of one species of free E2F, namely, an E2F-4–DP-1 heterodimer, that is exclusively cytoplasmic in the absence of E7. Taken together, the data reported here indicate that several different E7-dependent changes of cellular-growth-regulating pathways can cooperate to allow adhesion-independent entry into S phase.