bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequence-Specific Binding of Human Immunodeficiency Virus Type 1 Nucleocapsid Protein to Short Oligonucleotides
Authors: Fisher Robert J. , Rein Alan , Fivash Matthew , Urbaneja Maria A. , Casas-Finet José R. , Medaglia Maxine , Henderson Louis E. 
Published: December 2019

ABSTRACT

We have analyzed the binding of recombinant human immunodeficiency virus type 1 nucleocapsid protein (NC) to very short oligonucleotides by using surface plasmon resonance (SPR) technology. Our experiments, which were conducted at a moderate salt concentration (0.15 M NaCl), showed that NC binds more stably to runs of d(G) than to other DNA homopolymers. However, it exhibits far more stable binding with the alternating base sequence d(TG)
n
than with any homopolymeric oligodeoxyribonucleotide; thus, it shows a strong sequence preference under our experimental conditions. We found that the minimum length of an alternating d(TG) sequence required for stable binding was five nucleotides. Stable binding to the tetranucleotide d(TG)
2
was observed only under conditions where two tetranucleotide molecules were held in close spatial proximity. The stable, sequence-specific binding to d(TG)
n
required that both zinc fingers be present, each in its proper position in the NC protein, and was quite salt resistant, indicating a large hydrophobic contribution to the binding. Limited tests with RNA oligonucleotides indicated that the preferential sequence-specific binding observed with DNA also occurs with RNA. Evidence was also obtained that NC can bind to nucleic acid molecules in at least two distinct modes. The biological significance of the specific binding we have detected is not known; it may reflect the specificity with which the parent Gag polyprotein packages genomic RNA or may relate to the functions of NC after cleavage of the polyprotein, including its role as a nucleic acid chaperone.
Temporal Gene Regulation of the Channel Catfish Virus (Ictalurid Herpesvirus 1)
Authors: Huang Suming , Hanson Larry A. 
Published: December 2019

ABSTRACT

To identify promoter regions that impart differential temporal regulation of channel catfish virus (CCV) genes, the transcriptional kinetics of an immediate-early gene and prospective early and late genes were characterized. A cDNA clone, designated IE3C, representing a third immediate-early transcript was identified. The 5′ end of the IE3C transcript was mapped to nucleotides 15,368 and 131,043 in the terminal repeat regions of the CCV genome. The full length of the transcript represented by the IE3C clone is 1,412 bp, and it most likely codes for the protein specified by open reading frame (ORF) 12. The putative product of ORF12 contains a consensus RING finger metal binding motif (C
3
HC
4
structure). Temporal expression studies, in conjunction with protein synthesis and DNA replication inhibition, demonstrated that the IE3C transcript belongs to an immediate-early kinetic class, the ORF5 transcript is a member of the early kinetic class, and ORF39 and ORF46 are true late-kinetic-class genes. Additionally, we demonstrated that ORF38 transcription overlaps ORF39 and the products presumably share the same poly(A) signal. The 5′ ends of the transcripts encoding ORF38, ORF39, and ORF46 were mapped to nucleotides 44,862, 45,254, and 59,644, respectively, and potential transcriptional control elements were located.
A Peptide Mimic of a Protective Epitope of Respiratory Syncytial Virus Selected from a Combinatorial Library Induces Virus-Neutralizing Antibodies and Reduces Viral Load In Vivo
Authors: Chargelegue Daniel , Obeid Obeid E. , Hsu Shiou-Chih , Shaw Michael D. , Denbury Andrew N. , Taylor Geraldine , Steward Michael W. 
Published: December 2019

ABSTRACT

Respiratory syncytial virus (RSV) is the most important cause of bronchiolitis and pneumonia in infants and young children worldwide. As yet, there is no effective vaccine against RSV infection, and previous attempts to develop a formalin-inactivated vaccine resulted in exacerbated disease in recipients subsequently exposed to the virus. In the work described here, a combinatorial solid-phase peptide library was screened with a protective monoclonal antibody (MAb 19) to identify peptide mimics (mimotopes) of a conserved and conformationally-determined epitope of RSV fusion (F) protein. Two sequences identified (S1 [HWYISKPQ] and S2 [HWYDAEVL]) reacted specifically with MAb 19 when they were presented as solid-phase peptides. Furthermore, after amino acid substitution analyses, three sequences derived from S1 (S1S [HWSISKPQ], S1K [KWYISKPQ], and S1P [HPYISKPQ]), presented as multiple antigen peptides (MAPs), also showed strong reactivity with MAb 19. The affinity constants of the binding of MAb 19, determined by surface plasmon resonance analyses, were 1.19 × 10
9
and 4.93 × 10
9
M
−1
for S1 and S1S, respectively. Immunization of BALB/c mice with these mimotopes, presented as MAPs, resulted in the induction of anti-peptide antibodies that inhibited the binding of MAb 19 to RSV and neutralized viral infection in vitro, with titers equivalent to those in sera from RSV-infected animals. Following RSV challenge of S1S mimotope-immunized mice, a 98.7% reduction in the titer of virus in the lungs was observed. Furthermore, there was a greatly reduced cell infiltration in the lungs of immunized mice compared to that in controls. These results indicate the potential of peptide mimotopes to protect against RSV infection without exacerbating pulmonary pathology.
Activation of Heterologous Gene Expression by the Large Isoform of Hepatitis Delta Antigen
Authors: Wei Yu , Ganem Don 
Published: December 2019

ABSTRACT
Hepatitis delta virus (HDV) encodes two isoforms of its principal gene product, hepatitis delta antigen (HDAg). These two forms play distinctive and complementary roles in viral replication. Here we report that the large (LHDAg), but not the small (SHDAg), isoform of HDAg has the capacity to activate the expression of cotransfected genes driven by a variety of promoters, including the pre-S, S, and C promoters of hepatitis B virus. Mutational analysis of the C-terminal 19 amino acids unique to LHDAg shows that changing prolines to alanines in the two PXXP motifs in this region specifically ablates the activation function without abolishing another activity of LHDAg, namely, its ability to inhibit HDV RNA synthesis. However, C-terminal truncations that also disrupt these PXXP motifs only slightly diminished the activation function, indicating that the proline mutations were not acting by inactivating potential SH3 interactions that could be mediated by these motifs. Mutation of the isoprenylated cysteine to serine decreases but does not abolish the activation activity, and overexpression of SHDAg does not interfere with the transactivation function of LHDAg. Although the mechanism and biological significance of this activity of LHDAg remain unknown, the presence of this activity serves as yet another marker that functionally distinguishes this protein from the closely related isoform SHDAg.
Modulation of Feline Immunodeficiency Virus Infection by Stromal Cell-Derived Factor
Authors: Hosie Margaret J. , Broere Nelleke , Hesselgesser Joseph , Turner Julie D. , Hoxie James A. , Neil James C. , Willett Brian J. 
Published: December 2019

ABSTRACTThe α-chemokine receptor CXCR4 has recently been shown to support syncytium formation mediated by strains of feline immunodeficiency virus (FIV) that have been selected for growth in the Crandell feline kidney cell line (CrFK-tropic virus). Given that both human and feline CXCR4 support syncytium formation mediated by FIV, we investigated whether human stromal cell-derived factor (SDF-1) would inhibit infection with FIV. Human SDF-1α and SDF-1β bound with a high affinity (KDs of 12.0 and 10.4 nM, respectively) to human cells stably expressing feline CXCR4, and treatment of CrFK cells with human SDF-1α resulted in a dose-dependent inhibition of infection by FIVPET. No inhibitory activity was detected when the interleukin-2 (IL-2)-dependent feline T-cell line Mya-1 was used in place of CrFK cells, suggesting the existence of a CXCR4-independent mechanism of infection. Furthermore, neither the human β-chemokines RANTES, MIP-1α, MIP-1β, and MCP-1 nor the α-chemokine IL-8 had an effect on infection of either CrFK or Mya-1 cells with CrFK-tropic virus. Envelope glycoprotein purified from CrFK-tropic virus competed specifically for binding of SDF-1α to feline CXCR4 and CXCR4 expression was reduced in FIV-infected cells, suggesting that the inhibitory activity of SDF-1α in CrFK cells may be the result of steric hindrance of the virus-receptor interaction following the interaction between SDF and CXCR4. Prolonged incubation of CrFK cells with SDF-1α led to an enhancement rather than an inhibition of infection. Flow cytometric analysis revealed that this effect may be due largely to up-regulation of CXCR4 expression by SDF-1α on CrFK cells, an effect mimicked by treatment of the cells with phorbol myristate acetate. The data suggest that infection of feline cells with FIV can be mediated by CXCR4 and that, depending on the assay conditions, infection can be either inhibited or enhanced by SDF-1α. Infection with FIV may therefore prove a valuable model in which to study the development of novel therapeutic interventions for the treatment of AIDS.
T Cells Expressing Activated LFA-1 Are More Susceptible to Infection with Human Immunodeficiency Virus Type 1 Particles Bearing Host-Encoded ICAM-1
Authors: Fortin Jean-François , Cantin Réjean , Tremblay Michel J. 
Published: December 2019

ABSTRACTThe incorporation of host-derived proteins in nascent human immunodeficiency virus type 1 (HIV-1) particles is a well-established phenomenon. We recently demonstrated that the physical presence of host-encoded ICAM-1 glycoproteins on HIV-1 leads to a significant increase in virus infectivity in an ICAM-1/LFA-1-dependent fashion (J.-F. Fortin, R. Cantin, G. Lamontagne, and M. Tremblay, J. Virol. 71:3588–3596, 1997). We show here that conversion of LFA-1 to high affinity for ICAM-1 with the use of anti-LFA-1 antibodies (clones NKI-L16 and MEM83) markedly enhances the susceptibility of different target T-lymphoid cell lines, as well as of primary peripheral blood mononuclear cells, to infection by ICAM-1-bearing HIV-1 particles (6- to 95-fold). It is known that T-cell receptor (TCR) cross-linking induces a transient increase in LFA-1 affinity for ICAM-1. Treatment of peripheral blood mononuclear cells with anti-TCR antibodies (clone OKT3) resulted in a transient increase in susceptibility to infection by ICAM-1-positive virions that parallels the previously reported kinetics of the LFA-1/ICAM-1 adhesion mechanism. Our results led us to postulate that the strong interaction taking place between virally incorporated ICAM-1 and cell surface-activated LFA-1 markedly enhances the efficiency of virus binding and entry, thus favoring greater infection by ICAM-1-bearing HIV-1 particles. In view of the knowledge that primary HIV-1 isolates harbor host-derived ICAM-1 on their surfaces, these results provide new information about the role of host-derived ICAM-1 in the life cycle of HIV-1 and how it could positively modulate the dynamics of the viral infection, mainly in cellular compartments, such as the lymphoid tissues, where the level of cellular activation is high and where the probability of encountering a T cell expressing the activated LFA-1 form is also elevated.
Chimeric Measles Viruses with a Foreign Envelope
Authors: Spielhofer Pius , Bächi Thomas , Fehr Thomas , Christiansen Gudrun , Cattaneo Roberto , Kaelin Karin , Billeter Martin A. , Naim Hussein Y. 
Published: December 2019

ABSTRACT

Measles virus (MV) and vesicular stomatitis virus (VSV) are both members of the
Mononegavirales
but are only distantly related. We generated two genetically stable chimeric viruses. In MGV, the reading frames of the MV envelope glycoproteins H and F were substituted by a single reading frame encoding the VSV G glycoprotein; MG/FV is similar but encodes a G/F hybrid in which the VSV G cytoplasmic tail was replaced by that of MV F. In contrast to MG/FV, MGV virions do not contain the MV matrix (M) protein. This demonstrates that virus assembly is possible in the absence of M; conversely, the cytoplasmic domain of F allows incorporation of M and enhances assembly. The formation of chimeric viruses was substantially delayed and the titers obtained were reduced about 50-fold in comparison to standard MV. In the novel chimeras, transcription and replication are mediated by the MV ribonucleoproteins but the envelope glycoproteins dictate the host range. Mice immunized with the chimeric viruses were protected against lethal doses of wild-type VSV. These findings suggest that it is feasible to construct MV variants bearing a variety of different envelopes for use as vaccines or for gene therapeutic purposes.
Autologous and Heterologous Neutralization Analyses of Primary Feline Immunodeficiency Virus Isolates
Authors: del Mauro Daniela , Matteucci Donatella , Giannecchini Simone , Maggi Fabrizio , Pistello Mauro , Bendinelli Mauro 
Published: December 2019

ABSTRACT
Feline immunodeficiency virus (FIV) provides a model system with which the significance of neutralizing antibody (NA) in immunosuppressive lentivirus infections may be studied. To date, no detailed analysis of the neutralization properties of primary FIV isolates has been reported. In this study, we have conducted the first comprehensive study of the sensitivity to autologous and heterologous neutralization in a lymphoid cell-based assay of 15 primary FIV isolates and, for comparison, of one tissue culture-adapted strain. Primary isolates in general proved highly NA resistant, although there was considerable individual variation. Variation was also observed in the capacity of immune sera to neutralize heterologous FIV isolates. The ability of sera to neutralize isolates or for isolates to be neutralized by sera did not correlate with epidemiological and genetic relatedness or with the quasispecies complexity of the isolates. From the study of specific-pathogen-free cats experimentally infected with viral isolates associated with NA of different breadths, it appears that the development of FIV vaccines cannot rely on the existence of viral strains inherently capable of inducing especially broad NA responses.
Requirement for an Aromatic Amino Acid or Histidine at the N Terminus of Sindbis Virus RNA Polymerase
Authors: Shirako Yukio , Strauss James H. 
Published: December 2019

ABSTRACT
The N terminal amino acid of nonstructural protein nsP4, the viral RNA polymerase, is a tyrosine in all sequenced alphaviruses; this is a destabilizing amino acid for the N-end rule pathway and results in rapid degradation of nsP4 produced in infected cells or in reticulocyte lysates. We have constructed 11 mutants of Sindbis virus bearing Phe, Ala, Thr, Cys, Leu, Met, Asn, Gln, Glu, Arg, or Pro at the N terminus of nsP4. Translation of RNAs in reticulocyte lysates showed that cleavage at the nsP3/nsP4 site occurred efficiently for all mutants except for Glu-nsP4, which was cleaved inefficiently, and Pro-nsP4, which was not detectably cleaved, and that Tyr, Cys, Leu, Arg, and Phe destabilized nsP4 but Ala, Met, Thr, Asn, Gln, and Glu stabilized nsP4 to various extents. The viability of the mutants was examined by transfection of chicken cells at 30 or 40°C. The Phe-nsP4 mutant formed large plaques at both temperatures. The Met-nsP4 mutant was also viable but formed small plaques at 30°C and minute plaques at 40°C. The remaining mutants did not form plaques at either temperature. However, after prolonged incubation at 30°C, all the mutants except Glu-nsP4 and Pro-nsP4 produced viable viruses. In the case of Cys-, Leu-, Asn-, Gln-, or Arg-nsP4, revertants that were indistinguishable in plaque phenotype from the wild-type virus arose by same-site reversion to Tyr, Trp, Phe, or His by a single nucleotide substitution in the original mutant codon. Viable viruses also arose from the Ala-, Leu-, Cys-, Thr-, Asn-, Gln-, and Arg-nsP4 mutants that retained the original mutations at the N terminus of nsP4, but these viruses formed smaller plaques than the wild-type virus and many were temperature sensitive. Our results indicate that only nsP4s bearing N-terminal Tyr, Phe, Trp, or His have wild-type or near-wild-type activity for RNA replication and that rapid degradation of nsP4 is not a prerequisite for its function. nsP4s bearing other N-terminal residues, with the exception of Met-nsP4, have only very low or negligible activity, so that no detectable infectious virus can be produced. However, suppressor mutations can arise that enable most such nsP4s to regain significant but still suboptimal activity.
Analysis of Host Range Restriction Determinants in the Rabbit Model: Comparison of Homologous and Heterologous Rotavirus Infections
Authors: Ciarlet Max , Estes Mary K. , Barone Christopher , Ramig Robert F. , Conner Margaret E. 
Published: December 2019

ABSTRACT

The main limitation of both the rabbit and mouse models of rotavirus infection is that human rotavirus (HRV) strains do not replicate efficiently in either animal. The identification of individual genes necessary for conferring replication competence in a heterologous host is important to an understanding of the host range restriction of rotavirus infections. We recently reported the identification of the P type of the spike protein VP4 of four lapine rotavirus strains as being P[14]. To determine whether VP4 is involved in host range restriction in rabbits, we evaluated infection in rotavirus antibody-free rabbits inoculated orally with two P[14] HRVs, PA169 (G6) and HAL1166 (G8), and with several other HRV strains and animal rotavirus strains of different P and G types. We also evaluated whether the parental rhesus rotavirus (RRV) (P5B[3], G3) and the derived RRV-HRV reassortant candidate vaccine strains RRV × D (G1), RRV × DS-1 (G2), and RRV × ST3 (G4) would productively infect rabbits. Based on virus shedding, limited replication was observed with the P[14] HRV strains and with the SA11 Cl3 (P[2], G3) and SA11 4F (P6[1], G3) animal rotavirus strains, compared to the homologous ALA strain (P[14], G3). However, even limited infection provided complete protection from rotavirus infection when rabbits were challenged orally 28 days postinoculation (DPI) with 10
3
50% infective doses of ALA rabbit rotavirus. Other HRVs did not productively infect rabbits and provided no significant protection from challenge, in spite of occasional seroconversion. Simian RRV replicated as efficiently as lapine ALA rotavirus in rabbits and provided complete protection from ALA challenge. Live attenuated RRV reassortant vaccine strains resulted in no, limited, or productive infection of rabbits, but all rabbits were completely protected from heterotypic ALA challenge. The altered replication efficiency of the reassortants in rabbits suggests a role for VP7 in host range restriction. Also, our results suggest that VP4 may be involved in, but is not exclusively responsible for, host range restriction in the rabbit model. The replication efficiency of rotavirus in rabbits also is not controlled by the product of gene 5 (NSP1) alone, since a reassortant rotavirus with ALA gene 5 and all other genes from SA11 was more severely replication restricted than either parental rotavirus strain.
Human Immunodeficiency Virus Replication and Genotypic Resistance in Blood and Lymph Nodes after a Year of Potent Antiretroviral Therapy
Authors: Günthard Huldrych F. , Wong Joseph K. , Ignacio Caroline C. , Guatelli John C. , Riggs Nanette L. , Havlir Diane V. , Richman Douglas D. 
Published: December 2019

ABSTRACT

Potent antiretroviral therapy can reduce human immunodeficiency virus (HIV) in plasma to levels below the limit of detection for up to 2 years, but the extent to which viral replication is suppressed is unknown. To search for ongoing viral replication in 10 patients on combination antiretroviral therapy for up to 1 year, the emergence of genotypic drug resistance across different compartments was studied and correlated with plasma viral RNA levels. In addition, lymph node (LN) mononuclear cells were assayed for the presence of multiply spliced RNA. Population sequencing of HIV-1
pol
was done on plasma RNA, peripheral blood mononuclear cell (PBMC) RNA, PBMC DNA, LN RNA, LN DNA, and RNA from virus isolated from PBMCs or LNs. A special effort was made to obtain sequences from patients with undetectable plasma RNA, emphasizing the rapidly emerging lamivudine-associated M184V mutation. Furthermore, concordance of drug resistance mutations across compartments was investigated. No evidence for viral replication was found in patients with plasma HIV RNA levels of <20 copies/ml. In contrast, evolving genotypic drug resistance or the presence of multiply spliced RNA provided evidence for low-level replication in subjects with plasma HIV RNA levels between 20 and 400 copies/ml. All patients failing therapy showed multiple drug resistance mutations in different compartments, and multiply spliced RNA was present upon examination. Concordance of nucleotide sequences from different tissue compartments obtained concurrently from individual patients was high: 98% in the protease and 94% in the reverse transcriptase regions. These findings argue that HIV replication differs significantly between patients on potent antiretroviral therapy with low but detectable viral loads and those with undetectable viral loads.
The Extracellular Domain of Vaccinia Virus Protein B5R Affects Plaque Phenotype, Extracellular Enveloped Virus Release, and Intracellular Actin Tail Formation
Authors: Mathew Elizabeth , Sanderson Christopher M. , Hollinshead Michael , Smith Geoffrey L. 
Published: December 2019

ABSTRACTVaccinia virus produces two morphologically distinct forms of infectious virus, termed intracellular mature virus (IMV) and extracellular enveloped virus (EEV). EEV is important for virus dissemination within a host and has different surface proteins which bind to cell receptors different from those used by IMV. Six genes are known to encode EEV-specific proteins. One of these, B5R, encodes a 42-kDa glycoprotein with amino acid similarity to members of the complement control protein superfamily and contains four copies of a 50- to 70-amino-acid repeat called the short consensus repeat (SCR). Deletion of B5R causes a small-plaque phenotype, a 10-fold reduction in EEV formation, and virus attenuation in vivo. In this study, we inserted mutated versions of the B5R gene lacking different combinations of the SCRs into a virus deletion mutant lacking the B5R gene. The resultant viruses each formed small plaques only slightly larger than those of the deletion mutant; however, the virus containing only SCR 1 formed plaques slightly larger than those of viruses with SCRs 1 and 2 or SCRs 1, 2, and 3. All of these viruses produced approximately 50-fold more infectious EEV than wild-type virus and formed comet-shaped plaques under liquid overlay. Despite producing more EEV, the mutant viruses were unable to induce the polymerization of actin on intracellular virus particles. The implications of these results for our understanding of EEV formation, release, and infectivity are discussed.