pinecone/core-2020-05-10-deduplication huggingface dataset:
Various Types of HIV Mixed Infections in Cameroon

AbstractIn order to assess the incidence of HIV mixed infection as well as to clarify the molecular epidemiology of HIV in central Africa, we investigated 43 HIVs obtained from 211 Cameroonian AC, ARC, and AIDS patients in 1994 and 1995. Part of thepolregion and part of theenvregion were phylogenetically analyzed. The genotypes observed were varied: of 43 specimens, 28 (65%) were subtype A, 1 (2%) was subtype B, 2 (5%) were subtype D, 3 (7%) were subtype F, and 2 (5%) were group O. Of the remaining 7 specimens, 3 were mixed infections with HIV-1 subtypes A and C, HIV-1 subtypes C and F, and HIV-2 subtype A and HIV-1 subtype A; 1 was a mixed infection with HIV-1 subtypes A and D and the highly divergent group O (triple infection); another 3 appeared to consist of mosaic genomes (A/G, A/E, and B/A recombinant). These data show that various types of mixed infection, such as between different subtypes of HIV-1 group M, between HIV-1 and HIV-2, and even between HIV-1 groups O and M, were confirmed at a rather high frequency (approximately 10%). The mixed infection is particularly significant where there is a greater variety of HIV-1 subtypes circulating, since it results in new genetic diversity generated by intersubtype recombination
Immunization with a Recombinant Envelope Protein (rgp90) of EIAV Produces a Spectrum of Vaccine Efficacy Ranging from Lack of Clinical Disease to Severe Enhancement

AbstractWe have previously reported that immunization of ponies with a baculovirus-expressed recombinant surface unit envelope protein (rgp90) for equine infectious anemia virus (EIAV) resulted in enhancement of disease symptoms and virus replication in 4 of 4 vaccine recipients subjected to a heterologous virus challenge (rgp90 I vaccine trial) (Wanget al.,1994). To extend these studies of EIAV vaccine enhancement, two additional and independent rgp90 vaccine trials (rgp90 II and rgp90 III) were performed. Combined, a total of 13 ponies were immunized with the rgp90 immunogen using our standard vaccination procedures and challenged with a heterologous strain of EIAV. In contrast to the uniform enhancement observed in the rgp90 I vaccine trial, the severity of clinical symptoms varied markedly among the rgp90 recipients: 5 ponies experienced enhanced disease symptoms, 5 ponies experienced moderate disease symptoms, and 3 ponies remained asymptomatic. Of particular interest, in the 5 ponies with enhanced clinical symptoms was a severe thrombocytopenia (≤105,000 platelets/μl) evident coincident with the first febrile episode following virus challenge. Thrombocytopenia was either absent (7/10 ponies) or substantially delayed (3/10 ponies) in naive control ponies inoculated with the standard EIAVPVchallenge. Measurements of virus replication in the challenged vaccine recipients indicated a correlation between the level of viral RNA in plasma and the severity of the disease. Interestingly, an association was not observed between serum antibody reactivity to the vaccine or native viral antigens and the frequency of enhancement. Thus, these observations demonstrate a previously unrecognized complexity of rgp90 vaccine efficacy that has important implications for AIDS vaccine development
Release of Viral Glycoproteins during Ebola Virus Infection

AbstractMaturation and release of the Ebola virus glycoprotein GP were studied in cells infected with either Ebola or recombinant vaccinia viruses. Significant amounts of GP were found in the culture medium in nonvirion forms. The major form represented the large subunit GP1that was shed after release of its disulfide linkage to the smaller transmembrane subunit GP2. The minor form were intact GP1,2complexes incorporated into virosomes. Vector-expressed GP formed spikes morphologically indistinguishable from spikes on virus particles, indicating that spike assembly is independent of other viral proteins. Analysis of a truncation mutant revealed an early and almost complete release of GP1,2molecules, showing that membrane anchoring is mediated by the carboxy-terminal hydrophobic domain of GP2. We have also compared wild-type virus which requires transcriptional editing for synthesis of full-length GP with a variant that does not depend on editing. Both viruses released comparable amounts of GP1, but the variant expressed only minute amounts of the small, soluble GP which is the expression product of nonedited mRNA species of the GP gene. The abundant shedding of soluble GP1may play an important role in the immunopathology of Ebola hemorrhagic fever in experimentally and naturally infected hosts
The Adeno-Associated Virus Rep78 Major Regulatory Protein Binds the Cellular TATA-Binding Proteinin Vitroandin Vivo

AbstractRep78 is the major regulatory protein of adenoassociated virus (AAV). Rep78 is able to transcriptionally regulate all three of AAV's promoters, as well as a variety of heterologous promoters. In an attempt to understand the mechanism of action by which Rep78 is able to regulate gene expression, we are investigating Rep78's possible protein–protein interaction with basal transcription factors. One such critical basal transcription factor is the human TATA binding protein, TBP. TBP is a core factor required for the assemblage of the transcription initiation complex, TFIID. In this report anin vitrointeraction between Rep78 and TBP was demonstrated in three different assay systems, including West(far)–Western analysis, electrophoretic mobility shift assay–supershift, and coimmunoprecipitation. Furthermore, using the yeast GAL4 two-hybrid system, anin vivointeraction between Rep78 and TBP was also demonstrated. Further still, the amino half of Rep78 is shown to be needed for Rep78–TBP interaction. Mutations within this region of Rep78 are known to be defective for transcriptional regulatory ability, suggesting a biological role for this interaction. Thus, Rep78 may regulate transcription through binding and regulating TBP's numerous interactions. Furthermore, as Rep78 is known to bind at least one other transcription factor (Sp1) and likely others, Rep78 may function as a TBP-associated factor in an altered TFIID-like complex
Internal/Structures Containing Transcriptase-Related Proteins in Top Component Particles of Mammalian Orthoreovirus

AbstractThe structure of mammalian orthoreovirus top component particles, which are profoundly deficient in the content of double-stranded RNA genome, was determined at 30 Å resolution by transmission cryoelectron microscopy and three-dimensional image reconstruction. Previously undetected, ordered densities, appearing primarily as pentameric flowers in the reconstruction, were seen to extend 65 Å inwardly from the inner capsid at the icosahedral fivefold axes. Identically positioned but lower density elements were observed in two types of partially uncoated top component particles obtained by limited proteolysis. The levels of three inner-capsid proteins—λ1, λ3, and μ2—were reduced in concert with the internal densities during proteolytic uncoating. Since λ3 contains the catalytic regions of the viral RNA polymerase and since both λ1 and μ2 appear to play roles in transcription or mRNA capping, the internal structures are concluded to be complexes of the viral transcriptase-related enzymes. The findings have implications for the mechanisms of transcription and mRNA capping by orthoreovirus particles
Variability of Persisting MHV RNA Sequences Constituting Immune and Replication-Relevant Domains

AbstractSurvivors of acute infection with the neurotropic JHM strain of mouse hepatitis virus develop a persistent infection of the central nervous system associated with chronic ongoing demyelination. Persistence is characterized by viral RNA in the absence of infectious virus. To associate persistence with possible immune evasion and/or replication defects, viral RNA from brains of acutely and persistently infected mice was examined for mutations by reverse transcriptase–PCR. Sequences analyzed included the encapsidation sequence (ECS), the transmembrane domains of the matrix (M) protein, and a cytotoxic T cell (CTL) epitope within the nucleocapsid (N) protein. The ECS, present only on genomic RNA, revealed minimal variability and was detected out to 120 days postinfection, suggesting low levels of replication. The M gene sequence also remained stable during persistence despite random mutations during the acute phase. Although the N gene sequence exhibited the greatest diversity, mutations were random and not selected for during persistence. A single exception was detected comprising a prominent Pro to Ser substitution in a region of N not associated with any known regulatory or immune function. Of the N gene mutations found within the CTL epitope in responder mice (H-2d), one resulted in reduced CTL recognition with no evidence of antagonist activity. However, this mutation was also detected in nonresponder mice (H-2b), suggesting that escape variants arising from CTL pressure play no role in establishing persistence in immunocompetent hosts infected as adults
Role of a Subdominant H-2Kd-Restricted SV40 Tumor Antigen Cytotoxic T Lymphocyte Epitope in Tumor Rejection

AbstractSV40-transformed mKSA cells (H-2d) readily induce progressively growing tumors in adult syngeneic BALB/c mice while expressing the full complement ofH-2dMHC class I antigens. BALB/c mice previously immunized with SV40, soluble SV40 T antigen, or irradiated SV40-transformed syngeneic, allogeneic, or xenogeneic cells reject an mKSA tumor challenge even though these mice have been considered low- or nonresponders to T antigen due to difficulty in demonstrating SV40 T antigen-specific CTL. We have investigated the role of H-2d-restricted CTL in the rejection of SV40 tumors in BALB/c mice. Immunization of BALB/c mice with SV40 induced T antigen-specific CTL which were largely H-2Ld-restricted. However, following repeatedin vitrorestimulation with mKSA cells, CTL emerged which recognized a subdominant H-2Kd-restricted epitope corresponding to T antigen residues 499–507. Immunization of BALB/c mice with a recombinant vaccinia virus expressing the T499–507epitope provided partial protection against a challenge of syngeneic mKSA tumor cells and induced the generation of T499–507-specific CTL. These results indicate that a subdominant H-2Kd-restricted CTL epitope can participate in the rejection of SV40 tumors in BALB/c mice
Lamivudine Therapy of WHV-Infected Woodchucks

AbstractHepatitis B viruses establish a chronic, productive, and noncytopathic infection of hepatocytes. Viral products are produced by transcription from multiple copies (5–50) of covalently closed circular (ccc) viral DNA. This cccDNA does not replicate, but can be replaced by DNA precursors that are synthesized in the cytoplasm. The present study was carried out to determine if long-term treatment with an inhibitor of viral DNA synthesis would lead to loss of virus products, including cccDNA, from the liver of woodchucks chronically infected with woodchuck hepatitis virus. Viral DNA synthesis was inhibited with the nucleoside analog, lamivudine (2′-deoxy-3′-thiacytidine). Lamivudine treatment produced a slow but progressive decline in viral titers in serum, to about 0.3% or less of the initial level. However, even after maintenance of drug therapy for 3–12 months, >95% of the hepatocytes in most animals were still infected. Significant declines in the percentage of infected hepatocytes and of intrahepatic cccDNA levels were observed in only three woodchucks, two in the group receiving lamivudine and one in the placebo control group. Moreover, virus titers eventually rose in woodchucks receiving lamivudine, suggesting that drug-resistant viruses began to spread through the liver starting at least as early as 9–12 months of treatment. Three types of mutation that may be associated with drug resistance were found at this time, in a region upstream of the YMDD motif in the active site of the viral reverse transcriptase. The YMDD motif itself remained unchanged. Not unexpectedly, the lamivudine therapy did not have a impact on development of liver cancer
High SDF-1 Expression in HIV-1 Carriers Does Not Correlate with CD8+T-Cell-Mediated Suppression of Viral Replication

AbstractStromal cell-derived factor 1 (SDF-1) inhibits T-cell tropic (T-tropic) HIV-1 infectionin vitro. In this study, we examined the regulatory role of SDF-1 on HIV-1 replication in peripheral blood mononuclear cells (PBMC) of HIV-infected individuals. We found that the amount of SDF-1 mRNA in freshly isolated PBMC of HIV-1 carriers was higher than in healthy donors. Moreover, PBMC from some asymptomatic carriers (ACs) exhibited high levels of SDF-1 mRNA expression. The level of SDF-1 expression in PBMC did not correlate with the magnitude of CD8+T cell-mediated suppression of HIV-1 among ACs. SDF-1 inhibited HIV-1 replication at the viral entry step, whereas a single-cycle HIV-1 infection system showed that the major part of the CD8+T-cell-mediated suppression occurs after intracellular penetration of the virus. Our results suggest that SDF-1 acts as a suppressor of virus replication in a CD8+T-cell-independent mechanism in HIV-infected individuals
Expression and Mutational Analysis of the Baculovirus Very Late Factor 1 (vlf-1) Gene

AbstractWe have examined the expression and function of a gene,vlf-1,ofAutographa californicanuclear polyhedrosis virus that is known to encode a regulator of very late gene transcription. Western blot analysis revealed thatvlf-1is expressed during the late phase of infection, primarily from 15 to 24 h postinfection. VLF-1 localized in the cell nucleus and was also present in the nucleocapsids of virus particles. Mapping ofvlf-1mRNA by primer extension showed that transcription initiates at a TAAG motif 71 bp upstream of thevlf-1open reading frame. Disruption of this TAAG motif abolished the ability ofvlf-1to stimulate transcription from the very late polyhedrin gene (polh) promoter in transient expression assays, suggesting thatvlf-1expression is controlled by the TAAG motif. Using a highly efficient system to construct recombinant viruses with modifications invlf-1,we confirmed that the TAAG motif was essential. Furthermore, efforts to construct null mutants ofvlf-1failed, suggesting thatvlf-1is an essential gene for virus replication. Computer-assisted sequence homology searches placevlf-1in the λ phage integrase family (McLachlin and Miller, 1994). None of the strictly conserved residues of this family which are found invlf-1could be changed in the viral genome, implying that the putative integrase activity of VLF-1 is associated with the essential function ofvlf-1.However, mutation of a crucial active-site tyrosine did not affect the ability ofvlf-1to transactivate thepolhpromoter in transient expression assays, indicating that the very late transcriptional activity of VLF-1 does not require the integrase activity
Bacteriophage P2 and P4 Morphogenesis: Structure and Function of the Connector

AbstractThe connector, the structure located between the bacteriophage capsid and tail, is interesting from several points of view. The connector is in many cases involved in the initiation of the capsid assembly process, functions as a gate for DNA transport in and out of the capsid, and is, as implied by the name, the structure connecting a tail to the capsid. Occupying a position on a 5-fold axis in the capsid and connected to a coaxial 6-fold tail, it mediates a symmetry mismatch between the two. To understand how the connector is capable of all these interactions its structure needs to be worked out. We have focused on the bacteriophage P2/P4 connector, and here we report an image reconstruction based on 2D crystalline layers of connector protein expressed from a plasmid in the absence of other phage proteins. The overall design of the connector complies well with that of other phage connectors, being a toroid structure having a conspicuous central channel. Our data suggests a 12-fold symmetry, i.e., 12 protrusions emerge from the more compact central part of the structure. However, rotational analysis of single particles suggests that there are both 12- and 13-mers present in the crude sample. The connectors used in this image reconstruction work differ from connectors in virions by having retained the amino-terminal 26 amino acids normally cleaved off during the morphogenetic process. We have used different late gene mutants to demonstrate that this processing occurs during DNA packaging, since only mutants in geneP, coding for the large terminase subunit, accumulate uncleaved connector protein. The suggestion that the cleavage might be intimately involved in the DNA packaging process is substantiated by the fact that the fragment cleaved off is highly basic and is homologous to known DNA binding sequences
Mutations in Viral Movement Protein Alter Systemic Infection and Identify an Intercellular Barrier to Entry into the Phloem Long-Distance Transport System

AbstractViral systemic infection of a plant host involves two processes, cell-to-cell movement and long-distance transport. Molecular determinants associated with these two processes were probed by investigating the effects that alanine scanning mutations in the movement protein (MP) of red clover necrotic mosaic virus (RCNMV) had on viral infection in the plant hostsNicotiana edwardsonii, Vigna unguiculata(cowpea), and the experimental plantNicotiana benthamiana.Plants were inoculated with RCNMV expressing wild-type and mutant forms of the MP. Immunocytochemical studies at the light and electron microscope levels were performed on these plants, using a polyclonal antibody raised against the RCNMV capsid protein to identify the cells/tissues that RCNMV could infect. These experiments demonstrated that one cellular boundary at which the RCNMV MP functions to facilitate entry into the phloem long-distance transport system is located at the interfaces between the bundle sheath and phloem parenchyma cells and the companion cell–sieve element complex. Interestingly, inNicotiana tabacum,a host that only allows a local infection, RCNMV cell-to-cell movement was found to be blocked at this same intercellular boundary. Four mutants that were able to systemically infectN. benthamianawere partially or completely defective for systemic infection ofN. edwardsoniiand cowpea, which indicated that these MP mutants exhibited host-specific defects. Thus, the roles of the RCNMV MP in cell-to-cell movement and in long-distance transport appear to be genetically distinct. These results are discussed in terms of the mechanism by which RCNMV enters the phloem to establish a systemic infection
Nef Proteins of Distinct HIV-1 or -2 Isolates Differ in Their Binding Properties for HCK: Isolation of a Novel Nef Binding Factor with Characteristics of an Adaptor Protein

AbstractThe Nef gene of the human and simian immunodeficiency viruses HIV and SIV has been implicated in pathogenicity; however, the mechanism by which Nef induces disease is still unknown. An impact on signal transduction in cells has been suggested by the interaction of Nef from an HIV-1 strain and tyrosine kinases like HCK and LCK as well as serine/threonine kinases. We have confirmed the binding of HCK to HIV-1 subtype B Nef and demonstrated an equally strong interaction with a subtype E Nef protein but weaker binding to Nef of HIV-2 subtype A (HIV-2D194). No binding, however, was observed to HIV-2 subtype B Nef (HIV-2D205). Instead, this protein bound to a novel cellular protein,Nefin1,with characteristics of an adaptor protein and strong expression in all human hematopoietic tissues.Nefin1binds through an amino-terminal domain, which is related to SH3 domains. For interaction of Nef withNefin1,the PxxP motif and the three-dimensional conformation of the molecule appear necessary. In conclusion, this study demonstrates that Nef proteins of divergent strains of HIV-1 and HIV-2 may use different elements of signal transduction pathways for the induction of pathogenicityin vivo
The cAMP-Dependent Protein Kinase A and Protein Kinase C-β Pathways Synergistically Interact to Activate HIV-1 Transcription in Latently Infected Cells of Monocyte/Macrophage Lineage

AbstractThe HIV-1 long terminal repeat (LTR) responds to a variety of cellular signal transduction pathways. We demonstrate that the cAMP-dependent protein kinase A (PKA) and protein kinase C (PKC) signaling pathways synergize to increase HIV-1 LTR-mediated transcription and viral replication in a latently infected promonocytic cell line (U1). The LTR-mediated synergy induced by cholera toxin (Ctx), a potent activator of the cAMP-dependent PKA pathway, and the PKC activator phorbol 12-myristate 13-acetate (PMA) was abrogated by a PKC-β-specific inhibitor (LY333531). In contrast, the LTR-mediated synergy induced by Ctx and TNFα was not affected by LY333531. The synergy induced by Ctx and TNFα was also abrogated by mutation of the cAMP-responsive downstream sequence elements (DSE) in the 5′ untranslated leader region, whereas the DSE mutations did not affect the synergy induced by Ctx and PMA. These distinctions indicate that Ctx cooperates differently with TNFα and PMA to activate the HIV-1 LTR. Ctx and PMA synergistically activated AP-1- and NF-κB-dependent transcription, even though no cooperative binding of AP-1 or NF-κB was observed in gel shift assays. An extensive mutational analysis of the HIV-1 LTR that included the NF-κB and AP-1 binding sites revealed no distinctcis-acting element or region within the HIV-1 LTR that was required for the transcriptional synergy. Ctx and PMA also synergistically interact to activate the HTLV-1 LTR. These results indicate that the transcriptional synergy elicited by Ctx and PMA targets multiple functional elements and promoters, requires a cooperative interaction between the PKA and PKC-β pathways, and differs mechanistically from the transcriptional synergy induced by Ctx and TNFα
DNA Sequences outside the Simian Virus 40 Early Region Cause Downregulation of T-Antigen Production in Permissive Simian Cells

AbstractUsing a series of modified wtSV40 and early region SV40 DNAs, we assayed the effect of viral late region sequences on T-antigen production by the SV40 early region. We found that SV40 late region (L-SV40) DNA sequences reduced T-antigen (T-Ag) production by the SV40 early region (E-SV40) when both viral regions were linked as they are in wtSV40 DNA. This was demonstrated by Western analysis which showed that E-SV40 DNA produced 10 times more T-Ag than wtSV40 DNA. L-SV40, with its own promoter but unlinked to E-SV40 DNA, also greatly inhibited T-Ag production when it was cotransfected with E-SV40. Therefore, L-SV40 DNA inhibited T-Ag production by E-SV40 DNA when presentin cisorin trans.We have shown that expression of the SV40 late transcription unit dominated that of the early (T-Ag gene) transcription unit because late region RNA accumulated to much higher levels than early viral RNA. However, in cotransfected cells L-SV40 DNA did not replicate to higher levels than E-SV40 DNA. We offer a model for control of T-Ag expression in which a relatively small amount of T-Ag activates late transcription at the expense of T-Ag gene transcription and that this represents a switch from early to late viral gene expression. We suggest that when activation of the late transcription unit occurs at the late promoter, expression of the T-Ag gene is greatly reduced. The L-SV40 promoter may inhibit T-Ag gene transcription by sequestering cellular factors required for early transcription, factors which may be present in limited amounts. We suggest further that activation of late transcription allows for the necessary production of large amounts of capsomeres and virions and downregulation of early transcription prevents the early region from interfering with capsid synthesis. We tested the model using a construct with a wild-type T-Ag gene but with mutations in the SV40 major late promoter which prevent the promoter from being bound by cellular repressors of late transcription. We found that this construct, which overproduces late SV40 RNA, was defective for T-Ag production. This indicates that activation of the late promoter results in repression of T-Ag gene expression