bluuebunny/crossref_metadata_2025_split huggingface dataset:
Initiation of Genomic Plus-Strand RNA Synthesis from DNA and RNA Templates by a Viral RNA-Dependent RNA Polymerase
Authors: Sivakumaran K. , Kao C. Cheng 
Published: December 2019

ABSTRACT
In contrast to the synthesis of minus-strand genomic and plus-strand subgenomic RNAs, the requirements for brome mosaic virus (BMV) genomic plus-strand RNA synthesis in vitro have not been previously reported. Therefore, little is known about the biochemical requirements for directing genomic plus-strand synthesis. Using DNA templates to characterize the requirements for RNA-dependent RNA polymerase template recognition, we found that initiation from the 3′ end of a template requires one nucleotide 3′ of the initiation nucleotide. The addition of a nontemplated nucleotide at the 3′ end of minus-strand BMV RNAs led to initiation of genomic plus-strand RNA in vitro. Genomic plus-strand initiation was specific since cucumber mosaic virus minus-strand RNA templates were unable to direct efficient synthesis under the same conditions. In addition, mutational analysis of the minus-strand template revealed that the −1 nontemplated nucleotide, along with the +1 cytidylate and +2 adenylate, is important for RNA-dependent RNA polymerase interaction. Furthermore, genomic plus-strand RNA synthesis is affected by sequences 5′ of the initiation site.
Evolution of the Hepatitis C Virus Second Envelope Protein Hypervariable Region in Chronically Infected Patients Receiving Alpha Interferon Therapy
Authors: Pawlotsky Jean-Michel , Germanidis Georgios , Frainais Pierre-Olivier , Bouvier Magali , Soulier Alexandre , Pellerin Muriel , Dhumeaux Daniel 
Published: December 2019

ABSTRACT
Sustained hepatitis C virus (HCV) RNA clearance is achieved in 8 to 12% of patients with chronic HCV infection treated with alpha interferon (IFN-α) at the approved dose of 3 MU three times a week for 6 months and in about 25% of those receiving this treatment for 12 months. We used single-strand conformation polymorphism analysis combined with cloning and sequencing strategies to characterize the genetic evolution of HCV second envelope gene hypervariable region 1 (HVR1) quasispecies during and after IFN therapy in patients who failed to clear HCV RNA. Sustained HCV RNA clearance was achieved in 6% of patients. Profound changes in HVR1 quasispecies major variants were estimated to occur in 70% of the patients during and after therapy. These changes were evolutionary and were characterized by shifts in the virus population, related to selection and subsequent diversification of minor pretreatment variants. The quasispecies changes appeared to be induced by changes in the host environment likely resulting from the IFN-induced enhancement and post-IFN attenuation of neutralizing and possibly cytotoxic responses against HVR1. The remaining patients had no apparent changes in HVR1 quasispecies major variants, suggesting selection of major pretreatment variants, but some changes were observed in other genomic regions. We conclude that IFN-α administration and withdrawal profoundly alters the nature of circulating HCV quasispecies, owing to profound changes in virus-host interactions, in patients in whom sustained HCV RNA clearance fails to occur. These changes are associated with profound alterations of the natural outcome of HCV-related liver disease, raising the hypothesis of a causal relationship.
The Epstein-Barr Virus BZLF1 Protein Interacts Physically and Functionally with the Histone Acetylase CREB-Binding Protein
Authors: Adamson Amy L. , Kenney Shannon 
Published: December 2019

ABSTRACT
The Epstein-Barr virus (EBV) immediate-early protein BZLF1 (Z) is a key regulator of the EBV latent-to-lytic switch. Z is a transcriptional activator which induces EBV early gene expression. We demonstrate here that Z interacts with CREB-binding protein (CBP), a histone acetylase and transcriptional coactivator. This interaction requires the amino-terminal region of CBP as well as the transactivation and leucine zipper domains of Z. We show that CBP enhances Z-mediated transactivation of EBV early promoters, in reporter gene assays and in the context of the endogenous genome. We also demonstrate that Z decreases CREB transactivation function and that this inhibitory effect is reversed by overexpression of CBP. We show that Z also interacts directly with CREB. However, mutational analysis indicates that Z inhibition of CREB activity requires the direct interaction between Z and CBP but not the direct interaction between Z and CREB. We propose that Z interacts with CBP to enhance viral early gene transcription. In addition, the Z-CBP interaction may control host cellular transcription factor activity through competition for limiting amounts of cellular CBP.
Mapping of the Feline Calicivirus Proteinase Responsible for Autocatalytic Processing of the Nonstructural Polyprotein and Identification of a Stable Proteinase-Polymerase Precursor Protein
Authors: Sosnovtseva Svetlana A. , Sosnovtsev Stanislav V. , Green Kim Y. 
Published: December 2019

ABSTRACT

Expression of the region of the feline calicivirus (FCV) ORF1 encoded by nucleotides 3233 to 4054 in an in vitro rabbit reticulocyte system resulted in synthesis of an active proteinase that specifically processes the viral nonstructural polyprotein. Site-directed mutagenesis of the cysteine (Cys
1193
) residue in the putative active site of the proteinase abolished autocatalytic cleavage as well as cleavage of the viral capsid precursor, suggesting that this “3C-like” proteinase plays an important role in proteolytic processing during viral replication. Expression of the region encoding the C-terminal portion of the FCV ORF1 (amino acids 942 to 1761) in bacteria allowed direct N-terminal sequence analysis of the virus-specific polypeptides produced in this system. The results of these analyses indicate that the proteinase cleaves at amino acid residues E
960
-A
961
, E
1071
-S
1072
, E
1345
-T
1346
, and E
1419
-G
1420
; however, the cleavage efficiency is varied. The E
1071
-S
1072
cleavage site defined the N terminus of a 692-amino-acid protein that contains sequences with similarity to the picornavirus 3C proteinase and 3D polymerase domains. Immunoprecipitation of radiolabeled proteins from FCV-infected feline kidney cells with serum raised against the FCV ORF1 C-terminal region showed that this “3CD-like” proteinase-polymerase precursor protein is apparently stable and accumulates in cells during infection.
Patterns of Chemokine Receptor Fusion Cofactor Utilization by Human Immunodeficiency Virus Type 1 Variants from the Lungs and Blood
Authors: Singh Anjali , Besson Gideon , Mobasher Azin , Collman Ronald G. 
Published: December 2019

ABSTRACT

Human immunodeficiency virus type 1 (HIV-1) infection is highly compartmentalized, with distinct viral genotypes being found in the lungs, brain, and other organs compared with blood. CCR5 and CXCR4 are the principal HIV-1 coreceptors, and a number of other molecules support entry in vitro but their roles in vivo are uncertain. To address the relationship between tissue compartmentalization and the selective use of entry coreceptors, we generated functional
env
clones from primary isolates derived from the lungs and blood of three infected individuals and analyzed their use of the principal, secondary, orphan, and virus-encoded coreceptors for fusion. All Env proteins from lung viruses used CCR5 but not CXCR4, while those from blood viruses used CCR5 or CXCR4 or both. The orphan receptor APJ was widely used for fusion by Env proteins from both blood and lung viruses, but none used the cytomegalovirus-encoded receptor US28. Fusion mediated by the secondary coreceptors CCR2b, CCR3, CCR8, and CX
3
CR1 and orphan receptors GPR1, GPR15, and STRL33 was variable and heterogeneous, with relatively broad utilization by
env
clones from isolates of one subject but limited use by
env
clones from the other two subjects. However, there was no clear distinction between blood and lung viruses in secondary or orphan coreceptor fusion patterns. In contrast to fusion, none of the secondary or orphan receptors enabled efficient productive infection. These results confirm, at the level of cofactor utilization, previous observations that HIV-1 populations in the lungs and blood are biologically distinct and demonstrate diversity within lung-derived as well as blood-derived quasispecies. However, the heterogeneity in coreceptor utilization among clones from each isolate and the lack of clear distinction between lung- and blood-derived Env proteins argue against selective coreceptor utilization as a major determinant of compartmentalization.
Human Immunodeficiency Virus Type 1 Intergroup (M/O) Recombination in Cameroon
Authors: Takehisa Jun , Zekeng Léopold , Ido Eiji , Yamaguchi-Kabata Yumi , Mboudjeka Innocent , Harada Yosuke , Miura Tomoyuki , Kaptué Lazare , Hayami Masanori 
Published: December 2019

ABSTRACTHere we describe, for the first time, recombinants between two highly divergent major groups of human immunodeficiency virus type 1 (HIV-1), M and O, within a Cameroonian woman infected with three different HIV-1 strains, a group O virus, a subtype D virus, and a recently reported IBNG (A/G)-like recombinant virus. Using nested extra-long PCR amplification, we sequenced from thepolregion to theenvregion including accessory genes of the viral genome obtained from the patient’s uncultured peripheral blood mononuclear cells and examined the phylogenetic position of each gene. Compared with sequential blood samples obtained in 1995 and 1996, there were multiple segmental exchanges between three HIV-1 strains (O, D, and IBNG) and all the recombinants appeared to be derived from a common M/O ancestor. Importantly, recombination between groups M and O occurred, even though the homology between these two groups is 69, 76, 68, and 55% in thegag,pol,vif-vpr, andenvregions, respectively. Recombination between strains with such distant lineages may contribute substantially to generating new HIV-1 variants.
Three-Dimensional Structure of Aleutian Mink Disease Parvovirus: Implications for Disease Pathogenicity
Authors: McKenna Robert , Olson Norman H. , Chipman Paul R. , Baker Timothy S. , Booth Tim F. , Christensen Jesper , Aasted Bent , Fox James M. , Bloom Marshall E. , Wolfinbarger James B. , Agbandje-McKenna Mavis 
Published: December 2019

ABSTRACT

The three-dimensional structure of expressed VP2 capsids of Aleutian mink disease parvovirus strain G (ADV
G-VP2
) has been determined to 22 Å resolution by cryo-electron microscopy and image reconstruction techniques. A structure-based sequence alignment of the VP2 capsid protein of canine parvovirus (CPV) provided a means to construct an atomic model of the ADV
G-VP2
capsid. The ADV
G-VP2
reconstruction reveals a capsid structure with a mean external radius of 128 Å and several surface features similar to those found in human parvovirus B19 (B19), CPV, feline panleukopenia virus (FPV), and minute virus of mice (MVM). Dimple-like depressions occur at the icosahedral twofold axes, canyon-like regions encircle the fivefold axes, and spike-like protrusions decorate the threefold axes. These spikes are not present in B19, and they are more prominent in ADV compared to the other parvoviruses owing to the presence of loop insertions which create mounds near the threefold axes. Cylindrical channels along the fivefold axes of CPV, FPV, and MVM, which are surrounded by five symmetry-related β-ribbons, are closed in ADV
G-VP2
and B19. Immunoreactive peptides made from segments of the ADV
G-VP2
capsid protein map to residues in the mound structures. In vitro tissue tropism and in vivo pathogenic properties of ADV map to residues at the threefold axes and to the wall of the dimples.
Adenovirus-Mediated Gene Expression In Vivo Is Enhanced by the Antiapoptotic Bcl-2 Gene
Authors: Bilbao Guadalupe , Contreras Juan L. , Zhang Huang-Ge , Pike M. Joyce , Overturf Ken , Mikheeva Galina , Krasnykh Victor , Curiel David T. 
Published: December 2019

ABSTRACTAn adenovirus vector encoding the human Bcl-2 gene (hBcl-2) was derived. In vivo expression of hBcl-2 in murine livers enhanced and prolonged adenovirus-mediated gene expression. Furthermore, in the hBcl-2-treated group a significant reduction in the apoptosis induced by the adenovirus vector was observed. Thus, the cytoprotection of the vector-infected cells with antiapoptotic genes appears promising for successful in vivo gene therapy.
Systematic Excision of Vector Sequences from the BAC-Cloned Herpesvirus Genome during Virus Reconstitution
Authors: Wagner Markus , Jonjić Stipan , Koszinowski Ulrich H. , Messerle Martin 
Published: December 2019

ABSTRACT
Recently the mouse cytomegalovirus (MCMV) genome was cloned as an infectious bacterial artificial chromosome (BAC) (M. Messerle, I. Crnković, W. Hammerschmidt, H. Ziegler, and U. H. Koszinowski, Proc. Natl. Acad. Sci. USA 94:14759–14763, 1997). The virus obtained from this construct is attenuated in vivo due to deletion of viral sequences and insertion of the BAC vector. We reconstituted the full-length MCMV genome and flanked the BAC vector with identical viral sequences. This new construct represents a versatile basis for construction of MCMV mutants since virus generated from the construct loses the bacterial sequences and acquires wild-type properties.
Tandemization of a Subregion of the Enhancer Sequences from SRS 19-6 Murine Leukemia Virus Associated with T-Lymphoid but Not Other Leukemias
Authors: Granger Steven W. , Bundy Linda M. , Fan Hung 
Published: December 2019

ABSTRACTMost simple retroviruses induce tumors of a single cell type when infected into susceptible hosts. The SRS 19-6 murine leukemia virus (MuLV), which originated in mainland China, induces leukemias of multiple cellular origins. Indeed, infected mice often harbor more than one tumor type. Since the enhancers of many MuLVs are major determinants of tumor specificity, we tested the role of the SRS 19-6 MuLV enhancers in its broad disease specificity. The enhancer elements of the Moloney MuLV (M-MuLV) were replaced by the 170-bp enhancers of SRS 19-6 MuLV, yielding the recombinants ΔMo+SRS+and ΔMo+SRS−M-MuLV. M-MuLV normally induces T-lymphoid tumors in all infected mice. Surprisingly, when neonatal mice were inoculated with ΔMo+SRS+or ΔMo+SRS−M-MuLV, all tumors were of T-lymphoid origin, typical of M-MuLV rather than SRS 19-6 MuLV. Thus, the SRS 19-6 MuLV enhancers did not confer the broad disease specificity of SRS 19-6 MuLV to M-MuLV. However, all tumors contained ΔMo+SRS M-MuLV proviruses with common enhancer alterations. These alterations consisted of tandem multimerization of a subregion of the SRS 19-6 enhancers, encompassing the conserved LVb and core sites and adjacent sequences. Moreover, when tumors induced by the parental SRS 19-6 MuLV were analyzed, most of the T-lymphoid tumors had similar enhancer alterations in the same region whereas tumors of other lineages retained the parental SRS 19-6 MuLV enhancers. These results emphasize the importance of a subregion of the SRS 19-6 MuLV enhancer in induction of T-cell lymphoma. The relevant sequences were consistent with crucial sequences for T-cell lymphomagenesis identified for other MuLVs such as M-MuLV and SL3-3 MuLV. These results also suggest that other regions of the SRS 19-6 MuLV genome contribute to its broad leukemogenic spectrum.
Inhibition of Epstein-Barr Virus Replication by a Benzimidazole
l
-Riboside: Novel Antiviral Mechanism of 5,6-Dichloro-2-(Isopropylamino)-1-β-
l
-Ribofuranosyl-1H-Benzimidazole
Authors: Zacny Valerie L. , Gershburg Eduard , Davis Michelle G. , Biron Karen K. , Pagano Joseph S. 
Published: December 2019

ABSTRACT

Although a number of antiviral drugs inhibit replication of Epstein-Barr virus (EBV) in cell culture, and acyclovir (ACV) suppresses replication in vivo, currently available drugs have not proven effective for treatment of EBV-associated diseases other than oral hairy leukoplakia. Benzimidazole riboside compounds represent a new class of antiviral compounds that are potent inhibitors of human cytomegalovirus (HCMV) replication but not of other herpesviruses. Here we characterize the effects of two compounds in this class against lytic replication of EBV induced in a Burkitt lymphoma cell line latently infected with EBV. We analyzed linear forms of EBV genomes, indicative of lytic replication, and episomal forms present in latently infected cells by terminal probe analysis followed by Southern blot hybridization as well as the high-molecular-weight unprocessed viral DNA by pulsed-field gel electrophoresis.
d
-Ribofuranosyl benzimidazole compounds that act as inhibitors of HCMV DNA maturation, including BDCRB (5,6-dichloro-2-bromo-1-β-
d
-ribofuranosyl-1H-benzimidazole), did not affect the accumulation of high-molecular-weight or monomeric forms of EBV DNA in the induced cells. In contrast, the generation of linear EBV DNA as well as precursor viral DNA was sensitive to the
l
-riboside 1263W94 [5,6-dichloro-2-(isopropylamino)-1-β-
l
-ribofuranosyl-1H-benzimidazole]. The 50% inhibitory concentration range for 1263W94 was 0.15 to 1.1 μM, compared with 10 μM for ACV. Thus, 1263W94 is a potent inhibitor of EBV. In addition, 1263W94 inhibited the phosphorylation and the accumulation of the essential EBV replicative cofactor, early antigen D.
CD4-Chemokine Receptor Hybrids in Human Immunodeficiency Virus Type 1 Infection
Authors: Klasse P. J. , Rosenkilde Mette M. , Signoret Nathalie , Pelchen-Matthews Annegret , Schwartz Thue W. , Marsh Mark 
Published: December 2019

ABSTRACTMost human immunodeficiency virus (HIV) strains require both CD4 and a chemokine receptor for entry into a host cell. In order to analyze how the HIV-1 envelope glycoprotein interacts with these cellular molecules, we constructed single-molecule hybrids of CD4 and chemokine receptors and expressed these constructs in the mink cell line Mv-1-lu. The two N-terminal (2D) or all four (4D) extracellular domains of CD4 were linked to the N terminus of the chemokine receptor CXCR4. The CD4(2D)CXCR4 hybrid mediated infection by HIV-1LAIto nearly the same extent as the wild-type molecules, whereas CD4(4D)CXCR4 was less efficient. Recombinant SULAIprotein competed more efficiently with the CXCR4-specific monoclonal antibody 12G5 for binding to CD4(2D)CXCR4 than for binding to CD4(4D)CXCR4. Stromal cell-derived factor 1 (SDF-1) blocked HIV-1LAIinfection of cells expressing CD4(2D)CXCR4 less efficiently than for cells expressing wild-type CXCR4 and CD4, whereas down-modulation of CXCR4 by SDF-1 was similar for hybrids and wild-type CXCR4. In contrast, the bicyclam AMD3100, a nonpeptide CXCR4 ligand that did not down-modulate the hybrids, blocked hybrid-mediated infection at least as potently as for wild-type CXCR4. Thus SDF-1, but not the smaller molecule AMD3100, may interfere at multiple points with the binding of the surface unit (SU)-CD4 complex to CXCR4, a mechanism that the covalent linkage of CD4 to CXCR4 impedes. Although the CD4-CXCR4 hybrids yielded enhanced SU interactions with the chemokine receptor moiety, this did not overcome the specific coreceptor requirement of different HIV-1 strains: the X4 virus HIV-1LAIand the X4R5 virus HIV-189.6, unlike the R5 strain HIV-1SF162, infected Mv-1-lu cells expressing the CD4(2D)CXCR4 hybrid, but none could use hybrids of CD4 and the chemokine receptor CCR2b, CCR5, or CXCR2. Thus single-molecule hybrid constructs that mimic receptor-coreceptor complexes can be used to dissect coreceptor function and its inhibition.
Translation Initiation Factor eIF4B Interacts with a Picornavirus Internal Ribosome Entry Site in both 48S and 80S Initiation Complexes Independently of Initiator AUG Location
Authors: Ochs Kerstin , Rust RenéC. , Niepmann Michael 
Published: December 2019

ABSTRACT
Most eukaryotic initiation factors (eIFs) are required for internal translation initiation at the internal ribosome entry site (IRES) of picornaviruses. eIF4B is incorporated into ribosomal 48S initiation complexes with the IRES RNA of foot-and-mouth disease virus (FMDV). In contrast to the weak interaction of eIF4B with capped cellular mRNAs and its release upon entry of the ribosomal 60S subunit, eIF4B remains tightly associated with the FMDV IRES during formation of complete 80S ribosomes. Binding of eIF4B to the IRES is energy dependent, and binding of the small ribosomal subunit to the IRES requires the previous energy-dependent association of initiation factors with the IRES. The interaction of eIF4B with the IRES in 48S and 80S complexes is independent of the location of the initiator AUG and thus independent of the mechanism by which the small ribosomal subunit is placed at the actual start codon, either by direct internal ribosomal entry or by scanning. eIF4B does not greatly rearrange its binding to the IRES upon entry of the ribosomal subunits, and the interaction of eIF4B with the IRES is independent of the polypyrimidine tract-binding protein, which enhances FMDV translation.
An Adenovirus–Epstein-Barr Virus Hybrid Vector That Stably Transforms Cultured Cells with High Efficiency
Authors: Tan Brenton T. , Wu Lily , Berk Arnold J. 
Published: December 2019

ABSTRACT
EBV episomes are nuclear plasmids that are stably maintained through multiple cell divisions in primate and canine cells (J. L. Yates, N. Warren, and B. Sugden, Nature 313:812–815, 1985). In this report, we describe the construction and characterization of an E1-deleted recombinant adenovirus vector system that delivers an EBV episome to infected cells. This adenovirus-EBV hybrid vector system utilizes Cre-mediated, site-specific recombination to excise an EBV episome from a target recombinant adenovirus genome. We demonstrate that this vector system efficiently delivers the EBV episome and stably transforms a large fraction of infected canine D-17 cells. Using a colony-forming assay, we demonstrate stable transformation of 37% of cells that survive the infection. However, maximal transformation efficiency is achieved at doses of the E1-deleted recombinant adenoviruses that are toxic to the infected cells. Consequently, E1-deleted vector toxicity imposes a limitation on our current vector system.
Localization of Mouse Hepatitis Virus Nonstructural Proteins and RNA Synthesis Indicates a Role for Late Endosomes in Viral Replication
Authors: van der Meer Yvonne , Snijder Eric J. , Dobbe Jessika C. , Schleich Sibylle , Denison Mark R. , Spaan Willy J. M. , Locker Jacomine Krijnse 
Published: December 2019

ABSTRACT
The aim of the present study was to define the site of replication of the coronavirus mouse hepatitis virus (MHV). Antibodies directed against several proteins derived from the gene 1 polyprotein, including the 3C-like protease (3CLpro), the putative polymerase (POL), helicase, and a recently described protein (p22) derived from the C terminus of the open reading frame 1a protein (CT1a), were used to probe MHV-infected cells by indirect immunofluorescence (IF) and electron microscopy (EM). At early times of infection, all of these proteins showed a distinct punctate labeling by IF. Antibodies to the nucleocapsid protein also displayed a punctate labeling that largely colocalized with the replicase proteins. When infected cells were metabolically labeled with 5-bromouridine 5′-triphosphate (BrUTP), the site of viral RNA synthesis was shown by IF to colocalize with CT1a and the 3CLpro. As shown by EM, CT1a localized to LAMP-1 positive late endosomes/lysosomes while POL accumulated predominantly in multilayered structures with the appearance of endocytic carrier vesicles. These latter structures were also labeled to some extent with both anti-CT1a and LAMP-1 antibodies and could be filled with fluid phase endocytic tracers. When EM was used to determine sites of BrUTP incorporation into viral RNA at early times of infection, the viral RNA localized to late endosomal membranes as well. These results demonstrate that MHV replication occurs on late endosomal membranes and that several nonstructural proteins derived from the gene 1 polyprotein may participate in the formation and function of the viral replication complexes.