bluuebunny/crossref_metadata_2025_split huggingface dataset:
Demonstration that orf2 Encodes the Feline Immunodeficiency Virus Transactivating (Tat) Protein and Characterization of a Unique Gene Product with Partial Rev Activity
Authors: de Parseval Aymeric , Elder John H. 
Published: December 2019

ABSTRACT

The long PCR technique was used to amplify the three size classes of viral mRNAs produced in cells infected by feline immunodeficiency virus (FIV). We identified in the
env
region a new splice acceptor site that generated two transcripts, each coding for an 11-kDa protein, p11

rev

, whose function is unknown. The small-size class of mRNAs included two bicistronic orf2/
rev
mRNAs and two
rev
-like mRNAs, consisting only of the second exon of
rev
and coding for a 15-kDa protein, p15

rev

. p15

rev

contained the minimal effector domain of Rev and was sufficient to mediate partial Rev activity. The bicistronic mRNAs encoded two distinct proteins, one of 23 kDa corresponding to Rev and a 9-kDa protein encoded by the orf2 gene. The orf2 gene product is a protein of 79 amino acids with characteristics similar to those of the Tat (transactivator) proteins of the ungulate lentiviruses. Transient expression assays, using the FIV long terminal repeat (LTR) to drive transcription of the bacterial gene for chloramphenicol acetyltransferase demonstrated that the orf2 gene transactivates gene expression an average of 14- to 20-fold above the basal level. Deletion mutants of the FIV LTR were generated to locate sequences responsive to transactivation mediated by the orf2 gene. A 5′ deletion mutant that removed the AP1 site resulted in residual low-level transactivation by orf2. Further experiments using LTR mutants with internal deletions identified three regions located between positions −126 and −47 relative to the cap site that were important for orf2-directed transactivation. These regions include the AP1 site, a C/EBP tandem repeat, and an ATF site.
Limited Breadth of the Protective Immunity Elicited by Simian Immunodeficiency Virus SIVmne gp160 Vaccines in a Combination Immunization Regimen
Authors: Polacino Patricia , Stallard Virginia , Klaniecki James E. , Montefiori David C. , Langlois Alphonse J. , Richardson Barbra A. , Overbaugh Julie , Morton William R. , Benveniste Raoul E. , Hu Shiu-Lok 
Published: December 2019

ABSTRACT
We previously reported that immunization with recombinant simian immunodeficiency virus SIVmne envelope (gp160) vaccines protected macaques against an intravenous challenge by the cloned homologous virus, E11S. In this study, we confirmed this observation and found that the vaccines were effective not only against virus grown on human T-cell lines but also against virus grown on macaque peripheral blood mononuclear cells (PBMC). The breadth of protection, however, was limited. In three experiments, 3 of 10 animals challenged with the parental uncloned SIVmne were completely protected. Of the remaining animals, three were transiently virus positive and four were persistently positive after challenge, as were 10 nonimmunized control animals. Protection was not correlated with levels of serum-neutralizing antibodies against the homologous SIVmne or a related virus, SIVmac251. To gain further insight into the protective mechanism, we analyzed nucleotide sequences in the envelope region of the uncloned challenge virus and compared them with those present in the PBMC of infected animals. The majority (85%) of the uncloned challenge virus was homologous to the molecular clone from which the vaccines were made (E11S type). The remaining 15% contained conserved changes in the V1 region (variant types). Control animals infected with this uncloned virus had different proportions of the two genotypes, whereas three of four immunized but persistently infected animals had >99% of the variant types early after infection. These results indicate that the protective immunity elicited by recombinant gp160 vaccines is restricted primarily to the homologous virus and suggest the possibility that immune responses directed to the V1 region of the envelope protein play a role in protection.
Reovirus-Induced Apoptosis Is Preceded by Increased Cellular Calpain Activity and Is Blocked by Calpain Inhibitors
Authors: Debiasi Roberta L. , Squier Margaret K. T. , Pike Bobbi , Wynes Murry , Dermody Terence S. , Cohen J. John , Tyler Kenneth L. 
Published: December 2019

ABSTRACTThe cellular pathways of apoptosis have not been fully characterized; however, calpain, a cytosolic calcium-activated cysteine protease, has been implicated in several forms of programmed cell death. Reoviruses induce apoptosis both in vitro and in vivo and serve as a model for studying virus-induced cell death. We investigated the potential role of calpain in reovirus-induced apoptosis in vitro by measuring calpain activity as well as evaluating the effects of calpain inhibitors. L929 cells were infected with reovirus type 3 Abney (T3A), and calpain activity, measured as cleavage of the fluorogenic calpain substrate Suc-Leu-Leu-Val-Tyr-AMC, was monitored. There was a 1.6-fold increase in calpain activity in T3A-infected cells compared to mock-infected cells; this increase was completely inhibited by preincubation with calpain inhibitor I (N-acetyl-leucyl-leucyl-norleucinal [aLLN]), an active-site inhibitor. Both aLLN and PD150606, a specific calpain inhibitor that interacts with the calcium-binding site, inhibited reovirus-induced apoptosis in L929 cells by 54 to 93%. Apoptosis induced by UV-inactivated reovirus was also reduced 65 to 69% by aLLN, indicating that inhibition of apoptosis by calpain inhibitors is independent of effects on viral replication. We conclude that calpain activation is a component of the regulatory cascade in reovirus-induced apoptosis.
Cleavage of Poly(A)-Binding Protein by Coxsackievirus 2A Protease In Vitro and In Vivo: Another Mechanism for Host Protein Synthesis Shutoff?
Authors: Kerekatte Vaishali , Keiper Brett D. , Badorff Cornel , Cai Aili , Knowlton Kirk U. , Rhoads Robert E. 
Published: December 2019

ABSTRACT

Infection of cells by picornaviruses of the rhinovirus, aphthovirus, and enterovirus groups results in the shutoff of host protein synthesis but allows viral protein synthesis to proceed. Although considerable evidence suggests that this shutoff is mediated by the cleavage of eukaryotic translation initiation factor eIF4G by sequence-specific viral proteases (2A protease in the case of coxsackievirus), several experimental observations are at variance with this view. Thus, the cleavage of other cellular proteins could contribute to the shutoff of host protein synthesis and stimulation of viral protein synthesis. Recent evidence indicates that the highly conserved 70-kDa cytoplasmic poly(A)-binding protein (PABP) participates directly in translation initiation. We have now found that PABP is also proteolytically cleaved during coxsackievirus infection of HeLa cells. The cleavage of PABP correlated better over time with the host translational shutoff and onset of viral protein synthesis than did the cleavage of eIF4G. In vitro experiments with purified rabbit PABP and recombinant human PABP as well as in vivo experiments with
Xenopus
oocytes and recombinant
Xenopus
PABP demonstrate that the cleavage is catalyzed by 2A protease directly. N- and C-terminal sequencing indicates that cleavage occurs uniquely in human PABP at
482
VANTSTQTM↓GPRPAAAAAA
500
, separating the four N-terminal RNA recognition motifs (80%) from the C-terminal homodimerization domain (20%). The N-terminal cleavage product of PABP is less efficient than full-length PABP in restoring translation to a PABP-dependent rabbit reticulocyte lysate translation system. These results suggest that the cleavage of PABP may be another mechanism by which picornaviruses alter the rate and spectrum of protein synthesis.
Nuclear Magnetic Resonance Analysis of Solution Conformations in C4-V3 Hybrid Peptides Derived from Human Immunodeficiency Virus (HIV) Type 1 gp120: Relation to Specificity of Peptide-Induced Anti-HIV Neutralizing Antibodies
Authors: Vu Hai M. , Myers David , de Lorimier Robert , Matthews Thomas J. , Moody M. Anthony , Heinly Craig , Torres Jose V. , Haynes Barton F. , Spicer Leonard 
Published: December 2019

ABSTRACTImmunogenic peptides containing epitopes of the gp120 C4 and V3 regions from human immunodeficiency virus strains MN and EV91 have been studied by nuclear magnetic resonance and molecular modeling and used as immunogens in rhesus monkeys. The results, combined with those for other peptides, suggest a correlation between solution conformation and immunologic cross-reactivity.
RNA Replication for the Paramyxovirus Simian Virus 5 Requires an Internal Repeated (CGNNNN) Sequence Motif
Authors: Murphy Susan K. , Parks Griffith D. 
Published: December 2019

ABSTRACT

A functional RNA replication promoter for the paramyxovirus simian virus 5 (SV5) requires two essential and discontinuous elements: 19 bases at the 3′ terminus (conserved region I) and an 18-base internal region (conserved region II [CRII]) that is contained within the coding region of the L protein gene. A reverse-genetics system was used to determine the sequence requirements for the internal CRII element to function in RNA replication. A series of copyback defective interfering (DI) RNA analogs were constructed to contain point mutations in the 18 nucleotides composing CRII, and their relative replication levels were analyzed. The results indicated that SV5 DI RNA replication was reduced by substitutions for two CG dinucleotides, which in the nucleocapsid template are in the first two positions of the first two hexamers of CRII nucleotides. Substitutions for other bases within CRII did not reduce RNA synthesis. Thus, two consecutive 5′-CGNNNN-3′ hexamers form an important sequence in the SV5 CRII promoter element. The position of the CG dinucleotide within the SV5 leader and antitrailer promoters was highly conserved among other members of the
Rubulavirus
genus, but this motif differed significantly in both sequence and position from that previously identified for Sendai virus. The possible roles of the CRII internal promoter element in paramyxovirus RNA replication are discussed.
A Hairpin Structure in the R Region of the Human Immunodeficiency Virus Type 1 RNA Genome Is Instrumental in Polyadenylation Site Selection
Authors: Das Atze T. , Klaver Bep , Berkhout Ben 
Published: December 2019

ABSTRACT
Some retroviruses with an extended repeat (R) region encode the polyadenylation signal within the R region such that this signal is present at both the 5′ and 3′ ends of the viral transcript. This necessitates differential regulation to either repress recognition of the 5′ polyadenylation signal or enhance usage of the 3′ signal. The human immunodeficiency virus type 1 (HIV-1) genome encodes an inherently efficient polyadenylation signal within the 97-nucleotide R region. Polyadenylation at the 5′ HIV-1 polyadenylation site is inhibited by downstream splicing signals, and usage of the 3′ polyadenylation site is triggered by an upstream enhancer element. In this paper, we demonstrate that this on-off switch of the HIV-1 polyadenylation signal is controlled by a secondary RNA structure that occludes part of the AAUAAA hexamer motif, which we have termed the polyA hairpin. Opening the 5′ hairpin by mutation triggered premature polyadenylation and caused reduced synthesis of viral RNA, indicating that the RNA structure plays a pivotal role in repression of the 5′ polyadenylation site. Apparently, the same hairpin structure does not interfere with efficient usage of the 3′ polyadenylation site, which may be due to the presence of the upstream enhancer element. However, when the 3′ hairpin was further stabilized by mutation, we measured a complete loss of 3′ polyadenylation. Thus, the thermodynamic stability of the polyA hairpin is delicately balanced to allow nearly complete repression of the 5′ site yet efficient activation of the 3′ site. This is the first report of regulated polyadenylation that is mediated by RNA secondary structure. A similar hairpin motif that occludes the polyadenylation signal can be proposed for other lentiviruses and members of the spumaretroviruses, suggesting that this represents a more general gene expression strategy of complex retroviruses.
Genetic Selection of Poliovirus 2A
pro
-Binding Peptides
Authors: Ventoso Iván , Barco Angel , Carrasco Luis 
Published: December 2019

ABSTRACT

The yeast two-hybrid system has been used to identify mammalian clones that interact with poliovirus 2A proteinase (2A
pro
). Eight clones which encode previously unidentified human proteins were selected from a HeLa cell cDNA expression library. In addition, five clones encoding short peptides that interact with poliovirus 2A
pro
were also identified. The lengths of these peptides range from 6 to 30 amino acids, but all of them contain the Leu-X-Thr-Z motif (X represents any amino acid; Z represents a hydrophobic residue). This sequence is invariably located just at the carboxy terminus of each peptide. This approach raises the possibility of designing substrate analogue inhibitors of 2A
pro
. Thus, two nonhydrolyzable peptides containing the Leu-X-Thr-Z motif prevented cleavage of eukaryotic initiation factor 4G by poliovirus 2A
pro
in vitro. A more general method for identifying peptides with antiproteinase activity is discussed.
Truncated Particles Produced in Fish Surviving Infectious Hematopoietic Necrosis Virus Infection: Mediators of Persistence?
Authors: Kim Carol H. , Dummer Dina M. , Chiou Pinwen P. , Leong Jo-Ann C. 
Published: December 2019

ABSTRACT

Infectious hematopoietic necrosis virus (IHNV) is a rhabdovirus that produces an acute, lethal infection in rainbow trout (
Oncorhynchus mykiss
). Fish that survive infection cease to produce detectable infectious virus at approximately 46 days after infection, yet there is evidence that survivor fish continue to harbor virus particles (B. S. Drolet, P. P. Chiou, J. Heidel, and J. C. Leong, J. Virol. 69:2140–2147, 1995). In an effort to determine the biological function of these particles, the kidneys and livers from IHNV survivors were harvested and divided into samples for nested reverse transcriptase PCR analysis and explant culture. Sequences for the IHNV nucleoprotein and polymerase genes were detected in 50 and 89%, respectively, of the organs from survivor fish. When explant tissue cultures were infected with purified standard IHNV, the liver tissues from survivor fish produced up to 10-fold less virus than naive control fish liver tissues. In addition, immunosorbent electron microscopy analysis of the supernatant media from the cultured explants of survivor fish revealed truncated particles, whereas the control tissue supernatants contained only standard viral particles. These results suggest that the truncated IHNV particles observed in persistently infected fish are defective interfering particles that may mediate virus persistence.
Viral Load and a Locus on Chromosome 11 Affect the Late Clinical Disease Caused by Theiler’s Virus
Authors: Aubagnac Stéphanie , Brahic Michel , Bureau Jean-François 
Published: December 2019

ABSTRACTTheiler’s virus causes a persistent infection and a demyelinating disease of mice which is a model for multiple sclerosis. Susceptibility to viral persistence maps to several loci, including the interferon gamma locus. Inactivating the gene coding for the interferon gamma receptor makes 129/Sv mice susceptible to persistent infection and clinical disease, whereas inactivating the interferon gamma gene makes C57BL/6 mice susceptible to persistent infection but not to clinical disease. This difference in phenotype is due to the difference in genetic background. Clinical disease depends on high viral load andTmevd5, a locus on chromosome 11. These results have consequences for the identification of viruses which might be implicated in multiple sclerosis.
CD40-Mediated Induction of CD4 and CXCR4 on B Lymphocytes Correlates with Restricted Susceptibility to Human Immunodeficiency Virus Type 1 Infection: Potential Role of B Lymphocytes as a Viral Reservoir
Authors: Moir Susan , Lapointe Réjean , Malaspina Angela , Ostrowski Mario , Cole Charsey E. , Chun Tae-Wook , Adelsberger Joseph , Baseler Michael , Hwu Patrick , Fauci Anthony S. 
Published: December 2019

ABSTRACTHuman immunodeficiency virus type 1 (HIV-1) replicates primarily in lymphoid tissues where it has ready access to activated immune competent cells. We used one of the major pathways of immune activation, namely, CD40-CD40L interactions, to study the infectability of B lymphocytes isolated from peripheral blood mononuclear cells. Highly enriched populations of B lymphocytes generated in the presence of interleukin-4 and oligomeric soluble CD40L upregulated costimulatory and activation markers, as well as HIV-1 receptors CD4 and CXCR4, but not CCR5. By using single-round competent luciferase viruses complemented with either amphotropic or HIV-derived envelopes, we found a direct correlation between upregulation of HIV-1 receptors and the susceptibility of the B lymphocytes to infection with dual-tropic and T-tropic strains of HIV-1; in contrast, cells were resistant to M-tropic strains of HIV-1. HIV-1 envelope-mediated infection was completely abolished with either an anti-CD4 monoclonal antibody or a peptide known to directly block CXCR4 usage and partially blocked with stromal cell-derived factor 1, all of which had no effect on the entry of virus pseudotyped with amphotropic envelope. Full virus replication kinetics confirmed that infection depends on CXCR4 usage. Furthermore, productive cycles of virus replication occurred rapidly yet under most conditions, without the appearance of syncytia. Thus, an activated immunological environment may induce the expression of HIV-1 receptors on B lymphocytes, priming them for infection with selective strains of HIV-1 and allowing them to serve as a potential viral reservoir.
Discovery of a New Endogenous Type C Retrovirus (FcEV) in Cats: Evidence for RD-114 Being an FcEV
Gag-Pol
/Baboon Endogenous Virus BaEV
Env
Recombinant
Authors: van der Kuyl Antoinette C. , Dekker John T. , Goudsmit Jaap 
Published: December 2019

ABSTRACT

Analysis of a cat genomic DNA library showed that cats harbor a previously unrecognized endogenous type C retrovirus, whose
env
gene has homology to the murine
Fv-4
resistance gene. This unique retrovirus, designated FcEV (
Felis catus
endogenous retrovirus), has a type C
pol
gene, closely related to the primate
Papio cynocephalus
endogenous virus (PcEV)
pol
, not overlapping the
env
gene, unlike in other type C retroviruses, and is presumably present in a higher copy number than RD-114. Phylogenetic analysis of FcEV and RD-114 fragments amplified from cat species and comparison with baboon endogenous virus (BaEV) fragments from monkeys suggested that RD-114 does not represent the cat strain of BaEV but is actually a new recombinant between FcEV type C genes and the
env
gene of BaEV. Although BaEV did appear to have infected an ancestor of the domestic cat lineage, it was a de novo recombinant that made its way into the cat germ line.
Translation from the 5′ Untranslated Region (UTR) of mRNA 1 Is Repressed, but That from the 5′ UTR of mRNA 7 Is Stimulated in Coronavirus-Infected Cells
Authors: Senanayake Savithra D. , Brian David A. 
Published: December 2019

ABSTRACT

Viral gene products are generally required in widely differing amounts for successful virus growth and assembly. For coronaviruses, regulation of transcription is a major contributor to these differences, but regulation of translation may also be important. Here, we examine the possibility that the 5′ untranslated regions (UTRs), unique for each of the nine species of mRNA in the bovine coronavirus and ranging in length from 70 nucleotides (nt) to 210 nt (inclusive of the common 5′-terminal 65-nt leader), can differentially affect the rate of protein accumulation. When the natural 77-nt 5′ UTR on synthetic transcripts of mRNA 7 (mRNA for N and I proteins) was replaced with the 210-nt 5′ UTR from mRNA 1 (genomic RNA, mRNA for viral polymerase), approximately twofold-less N, or (N) CAT fusion reporter protein, was made in vitro. Twofold less was also made in vivo in uninfected cells when a T7 RNA polymerase-driven transient-transfection system was used. In coronavirus-infected cells, this difference surprisingly became 12-fold as the result of both a stimulated translation from the 77-nt 5′ UTR and a repression of translation from the 210-nt 5′ UTR. These results reveal that a differential 5′ UTR-directed regulation of translation can occur in coronavirus-infected cells and lead us to postulate that the direction and degree of regulation is carried out by viral or virally induced cellular factors acting in
trans
on
cis
-acting elements within the 5′ UTR.
Genetic Reassortment of Rift Valley Fever Virus in Nature
Authors: Sall A. A. , Zanotto P. M. de A. , Sene O. K. , Zeller H. G. , Digoutte J. P. , Thiongane Y. , Bouloy M. 
Published: December 2019

ABSTRACT

Rift Valley fever virus (RVFV), a phlebovirus of the
Bunyaviridae
family, is an arthropod-borne virus which emerges periodically throughout Africa, emphasizing that it poses a major threat for animal and human populations. To assess the genetic variability of RVFV, several isolates from diverse localities of Africa were investigated by means of reverse transcription-PCR followed by direct sequencing of a region of the small (S), medium (M), and large (L) genomic segments. Phylogenetic analysis showed the existence of three major lineages corresponding to geographic variants from West Africa, Egypt, and Central-East Africa. However, incongruences detected between the L, M, and S phylogenies suggested that genetic exchange via reassortment occurred between strains from different lineages. This hypothesis, depicted by parallel phylogenies, was further confirmed by statistical tests. Our findings, which strongly suggest exchanges between strains from areas of endemicity in West and East Africa, strengthen the potential existence of a sylvatic cycle in the tropical rain forest. This also emphasizes the risk of generating uncontrolled chimeric viruses by using live attenuated vaccines in areas of endemicity.