pinecone/core-2020-05-10-deduplication huggingface dataset:
Translation Elongation Factor-1α, La, and PTB Interact with the 3′ Untranslated Region of Dengue 4 Virus RNA

AbstractThe 384-nt long 3′ untranslated region (3′UTR) of dengue 4 virus (DEN4) is not polyadenylated, but contains the adjacent thermodynamically stable conserved short and long stem-loop structures (L-SL) and the conserved sequences CS1 and CS2. The latter are duplicated (CS2A and CS2B) in DEN4. Dengue virus replication, like that of other RNA viruses, might involve the cis-elements located within the 3′UTR and the trans-acting factors that could interact with the viral replicase to function as a replicase complex. The identification and characterization of viral and cellular proteins involved in the interaction with the 3′UTR of dengue virus will help us to understand the cellular requirements for viral replication. To determine these requirements, mobility shift and cross-linking assays were performed with uninfected and DEN4-infected C6/36 cell extracts as well as the different segments of the 3′UTR. Our results revealed that RNA–protein complexes were formed with the RNAs which involved the domains CS2A, CS2B, CS1, and L-SL. The minimum RNA sequence that was able to form specific and stable complexes with cellular proteins was the CS1–L-SL region. Using UV-induced cross-linking we identified eight proteins with molecular weights of 34, 39, 51, 52, 56, 62, 72, and 84 kDa that bound to the complete 3′UTR. The translation elongation factor-1α (EF-1α) bound to the complete 3′UTR and to the CS1–L-SL region. In addition, the recombinant GST–human La autoantigen bound to the 3′UTR and to the CS1–L-SL region as demonstrated by mobility shift and cross-linking assays. Although different antibodies against PTB were unable to react with any of the cellular proteins from C6/36, the recombinant His-PTB protein did bind to the complete 3′UTR and to the CS1–L-SL region. The specific binding of La and PTB to the sequences considered essential for viral RNA replication may suggest that these proteins could function as RNA chaperones to maintain RNA structure in a conformation that favors viral replication, while EF-1α may function as an RNA helicase
A Variable Region on the Chlorovirus CVK2 Genome Contains Five Copies of the Gene for Vp260, a Viral-Surface Glycoprotein

AbstractA 22.2-kb variable region near the left end of the chlorovirus CVK2 genome that was previously supposed to be expanded compared to the PBCV-1 genome was characterized. This region contains a tandem array of five gene copies for the Vp260-like protein, a viral-surface glycoprotein. The authentic 104-kDa Vp260 was found to be encoded at another site on the genome and to contain 13 internal tandem repeats of 61–65 amino acids, similar to the prominent Rickettsia surface antigen. The extra copies were also found to retain 10 of the internal repeats, despite the C-terminal deletions or extensions. These extra copies are conserved among chloroviruses isolated in various areas of Japan. By Northern blot analysis, these genes were demonstrated to be expressed late in infection. The proteins are incorporated into virions, as revealed by comparing viral structural proteins between wild-type and deletion mutants. These results indicate that extra copies of Vp260-like proteins encoded in a variable region on the genome may give variations in the surface nature of the chloroviral particles
Microarray Analysis of Gene Expression during Bacteriophage T4 Infection

AbstractGenomic microarrays were used to examine the complex temporal program of gene expression exhibited by bacteriophage T4 during the course of development. The microarray data confirm the existence of distinct early, middle, and late transcriptional classes during the bacteriophage replicative cycle. This approach allows assignment of previously uncharacterized genes to specific temporal classes. The genomic expression data verify many promoter assignments and predict the existence of previously unidentified promoters
Antigenic Peptides of the Epizootic Hematopoietic Necrosis Virus

AbstractThe epizootic hematopoietic necrosis virus (EHNV) is a strain of the Iridovirus genus, which includes viruses seriously affecting native and aquacultured fish and amphibians. Despite its growing importance as a threat to fish farming, very little information is available on the biochemical and immunological nature of this virus. To identify and characterize the main antigenic determinants of EHNV, a panel of murine monoclonal antibodies was produced upon parenteral inoculation with live virus. A total of 124 primary hybridoma cultures from two fusions was found to produce antibodies reacting with EHNV by ELISA, but no neutralizing monoclonal antibody was detected. Twenty hybridoma cultures were randomly chosen for further study, and the antibodies secreted were analyzed by Western blotting, radioimmunoprecipitation, and immunostaining of infected cells. Only three MAbs immunoprecipitated the 50-kDa EHNV major capsid protein (MCP) from infected cell lysates, but they did not stain this protein in Western blotting. Eight and five further MAbs recognized peptides of approximately 15 and 18 kDa, respectively. Four antibodies could not be mapped into any viral protein, although they specifically immunostained virus-infected cells and reacted with purified EHNV virions by ELISA. These latter MAbs and the three antibodies directed at the MCP are likely to recognize conformation-dependent epitopes on the virus capsid proteins
Identification and Genomic Analysis of a Second Species of Nucleopolyhedrovirus Isolated from Mamestra configurata

AbstractMacoNPV-96B is a nucleopolyhedrovirus isolated from naturally infected Mamestra configurata (Lepidoptera: Noctuidae) larvae. It was initially identified due to its completely different restriction endonuclease profile relative to the previously sequenced Mamestra configurata virus MacoNPV-90/2 (Q. Li, C. Donly, L. Li, L. G. Willis, D. A. Theilmann, and M. Erlandson, 2002, Virology 294, 106–121). The MacoNPV-96B host range and virulence were also found to differ significantly from those of the previous isolate. To further understand the complex of viruses infecting M. configurata, the genome of MacoNPV-96B was completely sequenced and analyzed in comparison with the genome of MacoNPV-90/2 and other sequenced baculoviruses. MacoNPV-96B consists of 158,482 bp, and 168 open reading frames (ORFs) of 150 nucleotides or longer with minimal overlap have been identified. The genome of MacoNPV-96B is 3422 bp larger than MacoNPV-90/2 and although gene arrangement is virtually identical, there are 9 ORFs unique to MacoNPV-96B and 10 unique to MacoNPV-90/2. bro genes were found to be associated with nonhomologous regions, suggesting that bro genes may facilitate recombination between genomes. A major difference in the gene content between the two viruses is a 5.4-kb insert in MacoNPV-96B, which is highly homologous to a cluster of Xestia c-nigrum granulovirus (XecnGV) ORFs, suggesting recent recombination events between these two viruses. Nucleotide sequence and amino acid sequence identity between the common ORFs of MacoNPV-96B and MacoNPV-90/2 average 87 and 90%, respectively. The sequence data suggest that MacoNPV-96B and MacoNPV-90/2 are closely related but have diverged and evolved into two separate species. This is the first study to identify highly related but separately evolving viruses in the same insect host and geographic location. A new Identity-GeneParityPlot analysis was developed to perform a comparison of two viral genomes in gene content and arrangement as well as homology level of individual ORFs
The Group-Specific Murine Coronavirus Genes Are Not Essential, but Their Deletion, by Reverse Genetics, Is Attenuating in the Natural Host

AbstractIn addition to a characteristic set of essential genes coronaviruses contain several so-called group-specific genes. These genes differ distinctly among the three coronavirus groups and are specific for each group. While the essential genes encode replication and structural functions, hardly anything is known about the products and functions of the group-specific genes. As a first step to elucidate their significance, we deleted the group-specific genes from the group 2 mouse hepatitis virus (MHV) genome via a novel targeted recombination system based on host switching (L. Kuo, G. J.Godeke, M. J. Raamsman, P. S. Masters, and P. J. M. Rottier, 2000, J. Virol. 74, 1393–1406). Thus, we obtained recombinant viruses from which the two clusters of group-specific genes were deleted either separately or in combination in a controlled genetic background. As all recombinant deletion mutant viruses appeared to be viable, we conclude that the MHV group-specific genes are nonessential, accessory genes. Importantly, all deletion mutant viruses were attenuated when inoculated into their natural host, the mouse. Therefore, deletion of the coronavirus group-specific genes seems to provide an attractive approach to generate attenuated live coronavirus vaccines
Induction of Interferon-α and Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand in Human Blood Mononuclear Cells by Hemagglutinin-Neuraminidase but Not F Protein of Newcastle Disease Virus

AbstractTo determine molecular viral components which can induce innate immune responses in human peripheral blood mononuclear cells (PBMC), we investigated the anti-neoplastic agent Newcastle disease virus (NDV) and its two spike proteins hemagglutinin-neuraminidase (HN) and fusion protein (F). NDV was an excellent inducer in PBMC of IFN-α production and capable of inducing upregulation of plasma membrane expression of tumor necrosis factor related apoptosis inducing ligand (TRAIL). Viral replication was not required for these responses because NDV inactivated for 5 min by UV was as good as live NDV. NDV-modified and paraformaldehyde-fixed BHK cells could also trigger IFN-α and TRAIL induction, indicating that contacts of responder cells with NDV-modified cell surfaces are sufficient to induce these activities in PBMC. Antibodies against HN but not F were able to block these responses. Finally we could show that HN but not F induced IFN-α and TRAIL in PBMC. This was possible through the use of respective gene transfectants generated with the help of Semliki Forest virus (SFV) replicase-based DNA recombinant expression systems. Upon contact with BHK cells expressing HN but not F at their cell surface, human PBMC produced IFN-α and some cells, including monocytes and T lymphocytes, upregulated cell surface TRAIL expression
Golgi Localization of Hantaan Virus Glycoproteins Requires Coexpression of G1 and G2

AbstractThe membrane glycoproteins G1 and G2 of Hantaan virus (HTNV; family Bunyaviridae) are encoded on the M RNA genome segment as a precursor polypeptide that is cotranslationally cleaved by host proteases. G1 and G2 accumulate in the Golgi complex in cells following either virus infection or transfection with M segment cDNA. However, there are conflicting reports in the literature concerning the Golgi targeting of separately expressed G1. To resolve these differences, a series of M segment and G1 coding region cDNA mutants was constructed containing either C-terminal or internal deletions. The intracellular localization of proteins expressed from these constructs was investigated by using confocal microscopy and double-staining immunofluorescence with G1 and G2 specific monoclonal antibodies and antisera specific for markers of the Golgi complex (GM130 and mannosidase II) and of the ER (calnexin). When expressed individually, G1 and G2 were retained in the ER, whereas when coexpressed from separate plasmids, both proteins localized to the Golgi. A construct expressing the whole G1 coding region and the complete signal sequence of G2 (amino acids 1–648 of the precursor) was found to be the minimal G1 protein competent to rescue G2 to the Golgi. This suggests that the G1 cytoplasmic tail including the downstream G2 signal peptide plays an important role in Golgi localization of HTNV glycoproteins. None of the constructs with internal deletions in the cDNA expressed proteins that localized to the Golgi. Our results indicate that the Golgi retention signal of HTNV glycoproteins may depend on the conformation of oligomerized G1 and G2 complex rather than a precise primary amino acid sequence
Evaluation of the Replication and Immunogenicity of Recombinant Human Parainfluenza Virus Type 3 Vectors Expressing up to Three Foreign Glycoproteins

AbstractThe level of replication and immunogenicity of recombinant parainfluenza virus type 3 (rHPIV3) bearing one, two, or three gene insertions expressing foreign protective antigens was examined. cDNA-derived recombinant HPIV3s bearing genes encoding the open reading frames (ORFs) of the hemagglutinin-neuraminidase (HN) of HPIV1, the HN of HPIV2, or the hemagglutinin (HA) of measles virus replicated efficiently in vitro, including the largest recombinant, which had three gene unit insertions and which was almost 23 kb in length, 50% longer than unmodified HPIV3. Several viruses were recovered from cDNAs whose genome length was not a multiple of six nucleotides and these contained nucleotide insertions that corrected the length to be a multiple of 6, confirming that the “rule of six” applies to HPIV3. Using a hemagglutination inhibition assay, we determined that the HPIV1 HN expressed by recombinant HPIV3 was incorporated into HPIV3 virions, whereas using this assay incorporation of the HPIV2 HN could not be detected. HPIV3 virions bearing HPIV1 HN were not neutralized by HPIV1 antiserum but were readily neutralized by antibodies to the HPIV3 HN or fusion protein (F). Viruses with inserts were restricted for replication in the respiratory tract of hamsters, and the level of restriction was a function of the total number of genes inserted, the nature of the insert, and the position of the inserted gene in the gene order. A single insert of HPIV2 HN or measles virus HA reduced the in vivo replication of rHPIV3 up to 25-fold, whereas the HPIV1 HN insert decreased replication almost 1000-fold. This indicates that the HPIV1 HN insert has an attenuating effect in addition to that of the extra gene insert itself, presumably because it is incorporated into the virus particle. Viruses containing two inserts were generally more attenuated than those with a single insert, and viruses with three inserts were over-attenuated for replication in hamsters. Inserts between the N and P genes were slightly more attenuating than those between the P and the M genes. A recombinant HPIV3 bearing both the HPIV1 and the HPIV2 HN genes (r1HN 2HN) was attenuated, immunogenic, and protected immunized hamsters from challenge with HPIV1, HPIV2, and HPIV3. Thus, it is possible to use a single HPIV vector expressing two foreign gene inserts to protect infants and young children from the severe lower respiratory tract disease caused by the three major human PIV pathogens
Sendai Virus, a Murine Parainfluenza Virus Type 1, Replicates to a Level Similar to Human PIV1 in the Upper and Lower Respiratory Tract of African Green Monkeys and Chimpanzees

AbstractHuman parainfluenza virus type 1 (HPIV1), a major cause of croup in infants and young children, accounts for 6% of hospitalizations for pediatric respiratory tract disease. The antigenically related Sendai virus, referred to here as murine PIV1 (MPIV1), is being considered for use as a live-attenuated vaccine to protect against HPIV1 (J. L. Hurwitz, K. F. Soike, M. Y., Sangster, A. Portner, R. E. Sealy, D. H. Dawson, and C. Coleclough, 1997, Vaccine 15(5), 533–540) and also as a recombinant vaccine vector expressing antigens to protect against viral disease in humans. However, in the 1950s MPIV1 was reported to have been isolated from humans, suggesting that zoonotic transmission might have occurred. It is therefore important to examine the ability of MPIV1 to replicate in nonhuman primates, i.e., surrogate hosts for humans. In the present study the level of replication of MPIV1 and HPIV1 was compared in African green monkeys and chimpanzees. Surprisingly, MPIV1 replicated as efficiently as HPIV1 in the upper and lower respiratory tract of African green monkeys at doses of 104 and 106 and replicated only slightly less efficiently at both sites in chimpanzees. African green monkeys immunized with MPIV1 were highly resistant to subsequent challenge with HPIV1 even though MPIV1 did not induce a detectable HPIV1-neutralizing antibody response. The high level of replication of MPIV1 observed in the upper and lower respiratory tract of these primates suggests that MPIV1 likely would require significant attenuation before it could be given to humans as a vaccine against HPIV1 or as a vaccine vector. Its ability to efficiently replicate in nonhuman primates suggests that MPIV1 lacks a significant host range restriction in primates and could theoretically cause zoonotic disease in humans
Sequence Analysis of an Isolate from a Fatal Human Infection of Australian Bat Lyssavirus

AbstractAustralian bat lyssavirus (ABLV), which occurs in pteropid and insectivorous bat populations, causes a rabies-like encephalitis in infected humans. We report the first complete sequence of an ABLV isolate obtained from a human who developed symptoms 27 months after being bitten by an infected flying fox. This isolate is the smallest lyssavirus to be sequenced, with a size of 11,918 nucleotides. Analyses of previously unsequenced regions and the complete genome confirm its close relationship with classical rabies viruses. In addition, a leucine zipper-like motif, not present in the other lyssaviruses, was found in the conserved domain I of the polymerase protein. This is the first report of a lyssavirus to vary in an 11-nucleotide, strictly conserved, complementary terminal sequence. This region is thought to encode important cis-acting regulatory signals; ABLV variation indicates a greater degree of flexibility than was thought for lyssaviruses in this region. A comparison of the pteropid and insectivorous isolates of ABLV indicates considerable differences between the two viruses. If the divergence of the two occurred on the Australian mainland, ABLV may have been endemic to Australia well before European colonisation
The Herpesvirus Saimiri Tip484 and Tip488 Proteins Both Stimulate Lck Tyrosine Protein Kinase Activity in Vivo and in Vitro

AbstractHerpesvirus saimiri (HVS) of subgroup C efficiently induces leukemia in New World primates and transforms human lymphocytes. The viral tyrosine kinase interacting protein (Tip) binds to the tyrosine protein kinase Lck and is essential for transformation. Understanding how Tip modulates Lck activity is important for elucidating the mechanism of herpesvirus saimiri leukemogenesis. However, there are reports suggesting that whereas the Tip protein of HVS strain 484 stimulates the activity of Lck, the Tip protein of HVS strain 488 inhibits Lck. To determine whether these two divergent Tip proteins have opposite effects on Lck activity, we compared them in parallel. We found that both Tip proteins stimulated Lck kinase activity in vivo and in vitro and that both stimulated NF-AT- and STAT3-dependent transcription in T cells. Our data support the model that HVS infection increases the activity of Lck through the action of Tip
The Nucleotide Sequence of RNA1 of Lettuce big-vein virus, Genus Varicosavirus, Reveals Its Relation to Nonsegmented Negative-Strand RNA Viruses

AbstractThe complete nucleotide sequence of RNA1 from Lettuce big-vein virus (LBVV), the type member of the genus Varicosavirus, was determined. LBVV RNA1 consists of 6797 nucleotides and contains one large ORF that encodes a large (L) protein of 2040 amino acids with a predicted Mr of 232,092. Northern blot hybridization analysis indicated that the LBVV RNA1 is a negative-sense RNA. Database searches showed that the amino acid sequence of L protein is homologous to those of L polymerases of nonsegmented negative-strand RNA viruses. A cluster dendrogram derived from alignments of the LBVV L protein and the L polymerases indicated that the L protein is most closely related to the L polymerases of plant rhabdoviruses. Transcription termination/polyadenylation signal-like poly(U) tracts that resemble those in rhabdovirus and paramyxovirus RNAs were present upstream and downstream of the coding region. Although LBVV is related to rhabdoviruses, a key distinguishing feature is that the genome of LBVV is segmented. The results reemphasize the need to reconsider the taxonomic position of varicosaviruses
Functional Replacement of the Tobacco rattle virus Cysteine-rich Protein by Pathogenicity Proteins from Unrelated Plant Viruses

AbstractMutation of the 16K gene encoded by RNA1 of Tobacco rattle virus (TRV) greatly reduced the levels of viral RNA that accumulated in both infected protoplasts and plants, showing that the 16K cysteine-rich protein (CRP) is required for efficient multiplication of TRV. Overexpression of the 16K protein, either from an additional copy of the gene carried on TRV RNA2 or from a PVX vector, led to an increase in the severity of disease symptoms, suggesting that the protein has a role in the pathogenicity of the virus. Mutation of the 16K gene could be overcome by expression from RNA2 of the Cucumber mosaic virus 2b gene, the Soil-borne wheat mosaic virus 19K gene, or the Barley stripe mosaic virus γb gene, indicating that the proteins encoded by these diverse genes may have similar functions. One known function of the CMV 2b gene is as a suppressor of posttranscriptional gene silencing, suggesting that the TRV 16K protein may also possess this activity
Internal Initiation by the Cucumber Necrosis Virus RNA-Dependent RNA Polymerase Is Facilitated by Promoter-like Sequences

AbstractTombusviruses, small positive sense RNA viruses of plants, are replicated by the viral-coded RNA-dependent RNA polymerase (RdRp) in infected cells. An unusual feature of the tombusvirus RdRp that is partially purified from cucumber necrosis virus (CNV)-infected plants is the ability to initiate complementary RNA synthesis from several internal positions on minus-strand templates derived from DI RNAs (19). In this study, we used template deletion, mutagenesis, and oligo-based inhibition of RNA synthesis to map the internal initiation sites observed with the in vitro CNV RdRp system. Comparing sequences around the internal initiation sites reveals that they have either (i) similar sequences to the core minus-strand initiation promoter; or (ii) similar structures to the core plus-strand initiation promoter. In addition, we find similarities among the internal initiation sites and the subgenomic RNA initiation sites. These similarities suggest that the mechanism of internal initiation is similar to initiation from the terminal core promoters or the putative subgenomic promoter sequences. We propose that internal initiation on full-length RNA templates may be important in defective interfering (DI) RNA formation/evolution by producing intermediate templates for RNA recombination in tombusviruses. This may explain why tombusviruses are frequently associated with DI RNAs