pinecone/core-2020-05-10-deduplication huggingface dataset:
Molecular Cloning and Biological Characterization of Full-Length HIV-1 Subtype C from Botswana

AbstractHuman immunodeficiency virus type 1 (HIV-1) subtype C is now responsible for more than half of all HIV-1 infections in the global epidemic and for the high levels of HIV-1 prevalence in southern Africa. To facilitate studies of the biological nature and the underlying molecular determinants of this virus, we constructed eight full-length proviral clones from two asymptomatic and three AIDS patients infected with HIV-1 subtype C from Botswana. Analysis of viral lysates showed that Gag, Pol, and Env structural proteins were present in the virions. In four clones, the analysis suggested inefficient envelope glycoprotein processing. Nucleotide sequence analysis of the eight clones did not reveal frameshifts, deletions, premature truncations, or translational stop codons in any structural, regulatory, or accessory genes. None of the subtype C clones were replication competent in donor peripheral blood mononuclear cells (PBMCs), macrophages, Jurkattat cells, or U87.CD4.CCR5 cells. However, infection by two clones could be rescued by complementation with a functional subtype C envelope clone, resulting in a productive infection of PBMCs, macrophages, and U87.CD4.CCR5 cells
The Human Papillomavirus E7 Protein Is Able to Inhibit the Antiviral and Anti-growth Functions of Interferon-α

AbstractIt is now well recognized that cervical cancer is caused by infection with certain human papillomavirus (HPV) subtypes and while interferon-α (IFN-α) is used to treat HPV-infected lesions, HPV appears to have developed a means to avoid the effects of IFN-α. Clinically, resistance appears to be associated with the expression of the E7 oncoprotein. Here we investigated the effects of expression in cells of the E7 protein from high- and low-risk papillomavirus subtypes on a range of responses to IFN-α. 2fTGH, a cell line dependent on IFN-α for growth in selection medium, grew significantly less well in the presence of E7, and the antiproliferative effects of IFN-α upon epithelial cells was lost upon E7 expression. The antiviral effects of IFN-α were abrogated in E7-expressing cells. Loss of response to IFN-α was found to occur in both high- and low-risk papillomaviruses. Finally, deletion of amino acids 21–24 of HPV type 16 E7 protein partially reversed repression. We conclude that E7 inhibits the functional effects of IFN-α and that this property is shared by all HPV subtypes tested
H9N2 Subtype Influenza A Viruses in Poultry in Pakistan Are Closely Related to the H9N2 Viruses Responsible for Human Infection in Hong Kong

AbstractFollowing the outbreak of H5N1 “bird flu” in Hong Kong in 1997, the isolation of H9N2 subtype viruses from patients in southern China and Hong Kong SAR once again raised the spectre of a possible influenza pandemic. H9N2 viruses have recently been responsible for disease in poultry in various parts of the world and preliminary studies of the H9 haemagglutinin (HA) genes of viruses isolated during 1998 and 1999 in Germany, Iran, Pakistan, and Saudi Arabia showed a close relationship to the HA genes of the viruses that infected two children in Hong Kong SAR. Analysis of the complete genome of a Pakistan isolate, A/chicken/Pakistan/2/99, showed that it is closely related in all eight genes (97–99% homology) to the human H9N2 isolates and furthermore that the six genes encoding internal components of the virus are similar to the corresponding genes of the H5N1 viruses that caused 6 (out of 18) fatal cases of human infection. Thus H9N2 viruses similar to those that caused human infections in Hong Kong are circulating more widely in other parts of the world. Whether or not these H9N2 viruses also have features that facilitate avian-to-human transmission is not known. Since avian H9N2 viruses are currently perceived to represent a significant threat to human health it is important to determine whether or not viruses of this subtype circulating in poultry in various parts of the world have the potential to infect people
Autographa californica M Nucleopolyhedrovirus chiA Is Required for Processing of V-CATH

AbstractInfection of permissive insect hosts by the baculovirus Autographa californica M nucleopolyhedrovirus results in liquefaction, a pathogenic effect that enhances the dispersal of progeny virions. Two viral gene products—a protease, V-CATH, and a chitinase, chiA—have been shown to be required for liquefaction to occur. It has been generally accepted that the primary functions of these proteins is to degrade the proteinaceous and chitinous components of the host cadaver, respectively. We have generated suggestive evidence, however, that chiA may also serve as a molecular chaperone for proV-CATH, the precursor of V-CATH. When cells were infected with virus lacking a functional chiA gene, proV-CATH failed to undergo processing in vivo and in vitro and formed insoluble aggregates in the endoplasmic reticulum of infected cells. Thus, expression of chiA may be required for the proper folding of the nascent V-CATH polypeptide in the endoplasmic reticulum. Identical results were obtained when tunicamycin was used to block N-linked glycosylation in cells infected with wildtype virus, suggesting that the putative chiA/V-CATH interaction is mediated by N-linked oligosaccharides
Construction of Full-Length cDNA Clones to Soil-Borne Wheat Mosaic Virus RNA1 and RNA2, from Which Infectious RNAs Are Transcribed In Vitro: Virion Formation and Systemic Infection without Expression of the N-Terminal and C-Terminal Extensions to the Capsid Protein

AbstractThe 19-kDa capsid protein (CP) of Soil-borne wheat mosaic furovirus (SBWMV) is encoded in the 5′-terminal region of RNA2. In addition to CP, two CP-related proteins are translated from SBWMV RNA2: (1) a 24-kDa protein (N-CP) with an N-terminal 40-amino-acid extension initiated at an upstream in-frame CUG codon; and (2) an 83-kDa protein (CP-RT) with an about 580-amino-acid, C-terminal extension by partial translational readthrough at the UGA termination codon at the end of the CP gene. We examined requirements for N-CP and CP-RT on virion formation and systemic infection in wheat plants using full-length cDNA clones, from which infectious RNA can be transcribed in vitro. RNA2 mutants, which could not synthesize N-CP, CP-RT, or either infected wheat plants systemically in combination with the wild-type RNA1 transcripts, produced rod-shaped virus particles in uninoculated upper leaves. Original mutations which abolished translation of N-CP and CP-RT were confirmed on RNA2 extracted from purified virus from the upper leaves by nucleotide sequence analysis. These results indicate that neither N-terminal nor C-terminal extensions to the CP are required for virion formation and systemic infection of SBWMV in wheat plants
Species-Specific TT Viruses in Humans and Nonhuman Primates and Their Phylogenetic Relatedness

AbstractBy means of polymerase chain reaction with a primer pair (NG133-NG147) deduced from the untranslated region (UTR) of TT virus (TTV), TTVs with markedly distinct genomic lengths were recovered from sera of humans and nonhuman primates, and their entire nucleotide sequences were determined. A human TTV [TGP96 of 2908 nucleotides (nt)] was obtained that was about 900 nt shorter than heretofore reported TTVs (3787–3853 nt). Likewise, TTVs of chimpanzee occurred in two distinct genomic sizes [Pt-TTV6 (3690 nt) and Pt-TTV8-II (2785 nt)]. Two TTVs of Japanese macaque [Mf-TTV3 (3798 nt) and Mf-TTV9 (3763 nt)] were comparable in genomic length, but only 55% similar in sequence. These five human and nonhuman primate TTVs, along with TTVs of tamarin [So-TTV2 (3371 nt)] and douroucouli [At-TTV3 (3718 nt)], were compared over the entire nucleotide sequence. Although the seven TTVs were only ≤55% similar, they share a common genomic organization with two open reading frames (ORFs), designated ORF1 (654–735 amino acids) and ORF2 (91–152 amino acids). The N-terminal sequences of ORF1 proteins were rich in arginine, and sequence motifs necessary for transcription and replication were conserved among them all. Like the human prototype TTV (TA278), all seven TTVs from various animals possessed in common two 15-nt sequences (CGAATGGCTGAGTTT and AGGGGCAATTCGGGC) in the UTR that were covered by NG133 and NG147, respectively. These primers would be instrumental in research on TTVs in previously unexamined species for defining their virological characteristics and evolutionary relationships
Infectious Hypodermal and Hematopoietic Necrosis Virus of Shrimp Is Related to Mosquito Brevidensoviruses

AbstractWe purified and sequenced infectious hypodermal and hematopoietic necrosis virus (IHHNV), a small DNA virus of shrimp, from wild Penaeus stylirostris. The virion has a buoyant density of 1.45 as determined by cesium chloride gradient. Analysis of 3873 nucleotides of the viral genome revealed three large open reading frames (ORFs) and parts of the noncoding termini of the viral genome. The left, mid, and right ORFs on the complementary (plus) strand have potential coding capacities of 666 amino acids (aa) (75.77 kDa), 363 aa (42.11 kDa), and 329 aa (37.48 kDa), respectively. The overall genomic organization is similar to that of the mosquito brevidensoviruses. The left ORF most likely encodes the major nonstructural (NS) protein (NS-1) since it contains conserved replication initiator motifs and NTP-binding and helicase domains similar to those in NS-1 from all other parvoviruses. The IHHNV putative NS-1 shares the highest aa sequence homology with the NS-1 of mosquito brevidensoviruses, Aedes densovirus and Aedes albopictus parvovirus. A search for putative splicing sites revealed that the N-terminal region of NS-1 is very likely located in a small ORF upstream of the left ORF. The right ORF is presumed to encode structural polypeptides (VPs), as in other parvoviruses. Two putative promoters, located upstream of the left and right ORFs, are presumed to regulate expression of NS and VP genes, respectively. Thus, IHHNV is closely related to densoviruses of the genus Brevidensovirus in the family Parvoviridae, and we therefore propose to rename this virus Penaeus stylirostris densovirus (PstDNV)
Translation Initiation of a Bicistronic mRNA of Borna Disease Virus: A 16-kDa Phosphoprotein Is Initiated at an Internal Start Codon

AbstractWe examined translational initiation of a bicistronic 0.8-kb mRNA of Borna disease virus (BDV) using a cDNA clone of the mRNA. Upon transfection with the clone, COS-7 cells produced a 16-kDa protein (P′), in addition to the previously identified products of BDV, 24- (P) and 14.5-kDa proteins. The 16-kDa product was detected by anti-P monoclonal antibody and was shown to exist in BDV-infected cell lines as well as in infected animal brain cells. Transient expression analysis of mutated cDNA clones encoding the BDV 0.8-kb mRNA revealed that the 16-kDa protein was initiated at the second AUG codon on the same open reading frame of the P protein. The mutational analysis also demonstrated that the first AUG within the 0.8-kb mRNA is not optimal, although the signal contains a better Kozak's motif. These results demonstrated the presence of three functional AUG codons in the smallest mRNA of BDV and also suggested that a leaky scanning mechanism is involved in translational initiation at AUG codons downstream of the bicistronic mRNA of BDV. Furthermore, the 16-kDa protein was located in the BDV-specific nuclear foci and was found to associate with the other viral proteins in BDV-infected cells, demonstrating an important role of the novel identified BDV protein in viral replication
Acetylation of HIV-1 Tat by CBP/P300 Increases Transcription of Integrated HIV-1 Genome and Enhances Binding to Core Histones

AbstractThe HIV-1 Tat protein is required for viral replication and is a potent stimulator of viral transcription. Although Tat has been extensively studied in various reductive paradigms, to date there is little information as to how this activator mediates transcription from natural nucleosomally packaged long terminal repeats. Here we show that CREB-binding protein (CBP)/p300 interacts with the HIV-1 Tat protein and serves as a coactivator of Tat-dependent HIV-1 gene expression on an integrated HIV-1 provirus. The site of acetylation of Tat was mapped to the double-lysine motif in a highly conserved region, 49RKKRRQ54, of the basic RNA-binding motif of Tat. Using HLM1 cells (HIV-1+/Tat−), which contain a single copy of full-length HIV-1 provirus with a triple termination codon at the first AUG of the Tat gene, we find that only wild type, and not K50A, K51A, or K50A/K51A alone or in combination of ectopic CBP/p300, is able to produce full-length infectious virions, as measured by p24 gag ELISAs. Tat binds CBP/p300 in the minimal histone acetyltransferase domain (1253–1710) and the binding is stable up to 0.85 M salt wash conditions. Interestingly, wild-type peptide 41–54, and not other Tat peptides, changes the conformation of the CBP/p300 such that it can acquire and bind better to basal factors such as TBP and TFIIB, indicating that Tat may influence the transcription machinery by helping CBP/p300 to recruit new partners into the transcription machinery. Finally, using biotinylated wild-type or acetylated peptides, we find that acetylation decreases Tat's ability to bind the TAR RNA element, as well as to bind basal factors such as TBP, CBP, Core-Pol II, or cyclin T. However, the acetylated Tat peptide is able to bind to core histones on a nucleosome assembled HIV-1 proviral DNA
Cloning of Human Picobirnavirus Genomic Segments and Development of an RT-PCR Detection Assay

AbstractNearly full-length genomic segments 2 and a partial-length genomic segment 1 of human picobirnavirus were cloned and sequenced. The clones were derived from viruses obtained from human immunodeficiency virus (HIV)-infected patients in Atlanta, Georgia (strains 3-GA-91 and 4-GA-91) and a nonHIV-infected person from China (strain 1-CHN-97). The picobirnavirus genomic segments lacked sequence similarities with other viral sequences in GenBank and EMBL. Comparison of genomic segment 1 from a human and a rabbit picobirnavirus identified a region of 127 nucleotides with 54.7% identity. The genomic segments 2 of the 4-GA-91 and 1-CHN-97 strains had 41.4% nucleic acid identity and 30.0% amino acid similarity and contained amino acid motifs typical of RNA-dependent RNA polymerase genes. Reverse transcription-PCR detection assays were developed with primers targeted to the genomic segments 2 of strains 4-GA-91 or 1-CHN-97. Picobirnaviruses related to the China strain were the predominant viruses detected in stool samples from people in four countries on three continents. Picobirnaviruses were detected in samples from two outbreaks of gastroenteritis in long-term elder care facilities but were not determined to be the primary pathogen. Our findings support the view that picobirnaviruses constitute a distinct family of viruses
VP5, the Nonstructural Polypeptide of Infectious Bursal Disease Virus, Accumulates within the Host Plasma Membrane and Induces Cell Lysis

AbstractInfectious bursal disease virus (IBDV) encodes a 17-kDa nonstructural polypeptide known as VP5. This polypeptide is not essential for virus replication in vitro but it plays an important role in in vivo dissemination and pathogenesis. We have characterized the expression of VP5 in three eukaryotic systems: (i) IBDV-infected chicken embryo fibroblasts; (ii) BSC-1 cells infected with a recombinant vaccinia virus vector; and (iii) Cos-1 cells transiently transfected with a plasmid vector. Immunofluorescence analyses showed that upon expression VP5 accumulates within the plasma membrane. This finding was consistent with sequence-based topology predictions, indicating that VP5 is a class II membrane protein with a cytoplasmic N-terminus and an extracellular C-terminal domain. Brefeldin A treatment of VP5-expressing cells prevented the accumulation of this polypeptide in the plasma membrane, thus showing the requirement of an active exocytic pathway to reach that compartment. Expression of VP5 was shown to be highly cytotoxic. Induction of VP5 expression resulted in the alteration of cell morphology, the disruption of the plasma membrane, and a drastic reduction of cell viability. VP5-induced cytotoxicity was prevented by blocking its transport to the membrane with Brefeldin A. Our findings suggest that VP5 plays an important role in the release of the IBDV progeny
Transcriptional Analysis of the Ribonucleotide Reductase Genes of Shrimp White Spot Syndrome Virus

AbstractThe causative agent of white spot syndrome (WSS) is a large double-stranded DNA virus, WSSV, which is probably a representative of a new genus, provisionally called Whispovirus. From previously constructed WSSV genomic libraries of a Taiwan WSSV isolate, clones with open reading frames (ORFs) that encode proteins with significant homology to the class I ribonucleotide reductase large (RR1) and small (RR2) subunits were identified. WSSV rr1 and rr2 potentially encode 848 and 413 amino acids, respectively. RNA was isolated from WSSV-infected shrimp at different times after infection and Northern blot analysis with rr1- and rr2-specific riboprobes found major transcripts of 2.8 and 1.4 kb, respectively. 5′ RACE showed that the major rr1 transcript started at a position of −84 (C) relative to the ATG translational start, while transcription of the rr2 gene started at nucleotide residue −68 (T). A consensus motif containing the transcriptional start sites for rr1 and rr2 was observed (TCAc/tTC). Northern blotting and RT-PCR showed that the transcription of rr1 and rr2 started 4–6 h after infection and continued for at least 60 h. The rr1 and rr2 genes thus appear to be WSSV “early genes.
Identification of a Cellular Repressor of Transcription of the Adenoviral Late IVa2 Gene That Is Unaltered in Activity in Infected Cells

AbstractThe gene encoding the adenovirus type 2 IVa2 protein, a sequence-specific activator of transcription from the viral major late promoter, is itself transcribed only during the late phase of infection. We previously identified a cellular protein (IVa2-RF) that binds specifically to an intragenic sequence of the IVa2 transcription unit. We now report that precise substitutions within the IVa2-RF-binding site that decreased binding affinity increased the efficiency of IVa2 transcription in in vitro reactions containing IVa2-RF. Consistent with the conclusion that this cellular protein represses IVa2 transcription, mutations that led to more efficient transcription in the presence of IVa2-RF were without effect in reactions lacking this cellular protein. No change in the concentration or activity of IVa2-RF could be detected in adenovirus-infected cells during the period in which the IVa2 gene is transcribed. We therefore propose that restriction of IVa2 transcription to the late phase is the result of titration of this cellular repressor as the number of copies of the IVa2 promoter increases upon replication of the viral genome
Association of the Human Papillomavirus Type 11 E1∧E4 Protein with Cornified Cell Envelopes Derived from Infected Genital Epithelium

AbstractThe cornified cell envelope (CCE) is an insoluble matrix of covalently linked proteins assembled in differentiating keratinocytes, providing a barrier against external insults. CCEs derived from HPV 11-infected tissue are fragile compared to those derived from healthy epithelium. To study a possible role for the E1∧E4 protein, HPV 11-infected epithelium was examined for the distribution of this protein and three CCE proteins. CCEs were then purified from genital epithelium, fragmented, washed to remove nonassociated proteins, and analyzed for E1∧E4 protein. In HPV 11-infected tissue, the E1∧E4 protein was detected in the region of the CCE in differentiated keratinocytes. Loricrin and cytokeratin 10 (K10) were absent in E1∧E4-positive cells, and E1∧E4 protein was not detected in cells containing these proteins. E1∧E4 protein was detected in immunoblots as a 10- to 11-kDa doublet in extracts of intact CCEs from infected tissue and in extracts of CCE fragments prepared without using reducing agents. Extraction with reducing agents eliminated E1∧E4 detection, suggesting that disulfide bonding was involved in the association with CCE fragments. In addition, cyanogen bromide degradation experiments, immunofluorescence, and immunoelectron microscopy provided evidence that E1∧E4 protein was associated with CCE fragments by covalent bonds other than disulfide bonds. We conclude that E1∧E4 protein expression is associated with profound alterations in detection of loricrin and K10 in HPV 11-infected genital epithelium. The E1∧E4 protein copurified with CCEs derived from infected epithelium and could be identified in CCE fragments, suggesting a possible role for E1∧E4 in the development of CCE abnormalities
Regulation of MVM NS1 by Protein Kinase C: Impact of Mutagenesis at Consensus Phosphorylation Sites on Replicative Functions and Cytopathic Effects

AbstractMinute virus of mice NS1, an 83-kDa mainly nuclear phosphoprotein, is the only viral nonstructural protein required in all cell types and it is involved in multiple processes necessary for virus propagation. The diversity of functions assigned to NS1, together with the variation of its complex phosphorylation pattern during infection, suggested that the various activities of NS1 could be regulated by distinct phosphorylation events. So far, it has been demonstrated that NS1 replicative functions, in particular, DNA-unwinding activities, are regulated by protein kinase C (PKC), as exemplified by the modulation of NS1 helicase activity by PKCλ phosphorylation. In order to determine further impact of phosphorylation on NS1 functions, including the induction of cytopathic effects, a mutational approach was pursued in order to produce NS1 variants harboring amino acid substitutions at candidate PKC target residues. Besides the determination of two additional in vivo phosphorylation sites in NS1, this mutagenesis allowed the segregation of distinct NS1 functions from one another, generating NS1 variants with a distinct activity profile. Thus, we obtained NS1 mutants that were fully proficient for trans activation of the viral P38 promoter, while being impaired in their replicative functions. Moreover, the alterations of specific PKC phosphorylation sites gave rise to NS1 polypeptides that exerted reduced cytotoxicity, leading to sustained gene expression, while keeping functions necessary for progeny virus production, i.e., viral DNA replication and activation of the capsid gene promoter. These data suggested that in the course of a viral infection, NS1 may undergo a shift from productive to cytotoxic functions as a result of a phosphorylation-dependent regulation
Downregulation of β1 Integrins by Ebola Virus Glycoprotein: Implication for Virus Entry

AbstractFiloviruses, including Ebola virus, are cytotoxic. To investigate the role of the Ebola virus glycoprotein (GP) in this cytopathic effect, we transiently expressed the GP in human kidney 293T cells. Expression of wild-type GP, but not the secretory form of the molecule lacking a membrane anchor, induced rounding and detachment of the cells, as did a chimeric GP containing its ectodomain and influenza virus hemagglutinin transmembrane–cytoplasmic domain. These results indicate that the GP ectodomain and its anchorage to the membrane are required for GP-induced morphologic changes in host cells. Since cell rounding and detachment could be associated with reduced levels of cell adhesion molecules, we also studied the expression of integrins, which are major molecules for adhesion to extracellular matrices, and found that the β1 integrin group is downregulated by the GP. This result was further extended by experiments in which anti-β1 monoclonal antibodies or purified integrins inhibited the infectivity of vesicular stomatitis virus pseudotyped with the GP. We suggest that integrins, especially the β1 group, might interact with the GP and perhaps be involved in Ebola virus entry into cells