pinecone/core-2020-05-10-deduplication huggingface dataset:
Inhibition of Rb and p53 Is Insufficient for SV40 T-Antigen Transformation

AbstractThe SV40 large T-antigen (TAg) has proven useful in studying pathways involved with cell division and tissue homeostasis. TAg disrupts the normal action of tumor suppressors pRb and p53. It is unclear whether T-antigen inhibition of p53 and pRb is sufficient for oncogenic transformation or if additional T-antigen activities are required. To pursue this question, cell lines were generated that coexpress an amino-terminal fragment of T-antigen (TAgN136), which has been shown to be sufficient to block pRb function, together with a dominant-negative p53. Neither focus formation nor saturation density was enhanced by coexpression of the dominant-negative p53 molecule, p53DD, along with TAgN136. Furthermore, a full-length TAg mutant incapable of binding p53 was capable of relieving contact inhibition, a hallmark of transformation. These results suggest the presence of a novel transforming activity in addition to the binding and inactivation of p53, requiring TAg amino acids 137 to 708
Activated Notch1 Signaling Cooperates with Papillomavirus Oncogenes in Transformation and Generates Resistance to Apoptosis on Matrix Withdrawal through PKB/Akt

AbstractInvasive cervical tumors, a major subset of human epithelial neoplasms, are characterized by the consistent presence of papillomavirus oncogenes 16 or 18 E6 and E7 products. Cervical tumors also consistently exhibit cytosolic and nuclear forms of Notch1, suggesting the possible persistent activation of the Notch pathway. Here we show that activated Notch1 synergizes with papillomavirus oncogenes in transformation of immortalized epithelial cells and leads to the generation of resistance to anoikis, an apoptotic response induced on matrix withdrawal. This resistance to anoikis by activated Notch1 is mediated through the activation of PKB/Akt, a key effector of activated Ras in transformation. We suggest that activated Notch signaling may serve to substitute for the lack of activated Ras mutations in the majority of human cervical neoplasms
Detection of Viral DNA and E4 Protein in Basal Keratinocytes of Experimental Canine Oral Papillomavirus Lesions

AbstractWe studied experimental canine oral papillomavirus (COPV) infection by in situ hybridization and immunohistochemistry of weekly biopsies. After 4 weeks, viral DNA in rete ridges suggested a keratinocyte stem cell target. Abundant viral DNA was seen in E4-positive cells only. E4 was predominantly cytoplasmic but also nuclear, being concentrated in the nucleoli during wart formation. Infected cells spread laterally along the basal layer and into the parabasal layers, accompanied by E7 transcription and increased mitoses. Most of the lower epithelium was positive for viral DNA, but, in mature warts, higher levels of E4 expression and genome amplification occurred in only sporadic superficial cells. L1 expression was late and in only a subset of E4-positive cells. During regression, viral DNA was less abundant in deep epithelial layers, suggesting downregulation of replication prior to replacement of infected cells from beneath. Detection of viral DNA in post-regression tissue indicated latent infection
Domains in the Simian Immunodeficiency Virus gp41 Cytoplasmic Tail Required for Envelope Incorporation into Particles

AbstractThe mechanism by which lentivirus envelope (Env) glycoproteins are packaged into budding virions is poorly understood. Simian immunodeficiency virus (SIV) contains an Env protein with an unusually long cytoplasmic tail. To investigate the role of this domain in the incorporation of the SIV Env into virions, we generated a series of SIV Env mutants carrying small in-frame deletions within the cytoplasmic domain. The effects of these mutations on Env synthesis, processing, and association with Gag particles were analyzed by means of the vaccinia virus expression system. All of the mutant Env glycoproteins were synthesized and processed in a manner similar to that of the wild-type Env. However, deletions affecting domains C-terminal to residue 832 in the SIV Env protein significantly impaired Env incorporation into particles. Cell surface biotinylation assays showed that this phenotype could not be attributed to inefficient cell surface expression of the Env mutants. Furthermore, when the Env deletion mutants were tested for their ability to mediate virus entry in single-cycle infectivity assays, those mutations that impaired Env incorporation also caused a severe defect in virus infectivity. Our results suggest that domains in the C-terminal third of the SIV Env protein are required for Env incorporation into particles and Env-mediated virus entry
High Virus Loads in Naturally and Experimentally SIVagm-Infected African Green Monkeys

AbstractA quantitative RT–PCR assay was developed for SIVagm and was used to measure the levels of viral RNA in the plasma of experimentally and naturally infected African green monkeys. The number of productively infected PBMCs and the number of cells carrying integrated provirus were also measured. Plasma virus loads in experimentally infected animals peaked at 2 weeks postinfection, ranging from 2.9 × 105 to 4.2 × 107 RNA copies/ml plasma. Set points of 2.1 × 103 to 2.8 × 106 RNA copies/ml plasma were maintained for one year. Similarly, the number of cells carrying integrated SIVagm provirus remained relatively stable in individual animals for one year with set points ranging from 73 to 810 proviral copies per 106 PBMC. However, the number of productively infected cells fluctuated considerably during this period. Virus loads in the 26 naturally infected AGMs ranged from 8.3 × 103 to 1.1 × 107 (mean 1.7 × 106) RNA copies/ml plasma. These levels of viremia are similar to those seen in pathogenic systems (HIV-1, SIVmac), indicating that control of SIVagm replication is not the reason for the natural host's resistance to disease progression
Analysis of the Complete DNA Sequence of the Temperate Bacteriophage TP901-1: Evolution, Structure, and Genome Organization of Lactococcal Bacteriophages

AbstractA complete analysis of the entire genome of the temperate lactococcal bacteriophage TP901-1 has been performed and the function of 21 of 56 TP901-1-encoded ORFs has been assigned. This knowledge has been used to propose 10 functional modules each responsible for specific functions during bacteriophage TP901-1 proliferation. Short regions of microhomology in intergenic regions present in several lactococcal bacteriophages and chromosomal fragments of Lactococcus lactis are suggested to be points of exchange of genetic material through homologous recombination. Our results indicate that TP901-1 may have evolved by homologous recombination between the host chromosome and a mother phage and support the observation that phage remnants as well as prophages located in the Lactococcus chromosome contribute significantly to bacteriophage evolution. Some proteins encoded in the early transcribed region of the TP901-1 genome were more homologous to proteins encoded by phages infecting gram-positive hosts other than L. lactis. This protein homology argues for the occurrence of horizontal genetic exchange among these bacteriophages and indicates that they have access to a common gene pool
Down-Modulation of the Costimulatory Molecule, CD28, Is a Conserved Activity of Multiple SIV Nefs and Is Dependent on Histidine 196 of Nef

AbstractIn this study Nef proteins derived from simian immunodeficiency virus (SIV) and human immunodeficiency virus (HIV) were compared to assess their abilities to down-modulate the cell surface levels of the T-cell costimulatory molecule CD28. We demonstrate that in addition to Nef derived from the prototypic SIVmac239, Nef proteins encoded by the pathogenic SIVsmmPBj molecular clone and the SIVsmmB670 isolate also down-modulate cell surface CD28. In contrast, Nef proteins derived from HIV failed to down-modulate CD28. We have also identified H196 as a critical residue which influences the capacity of SIVmac Nef to down-modulate CD28. Nef derived from SIVmacJ5 failed to down-modulate cell surface CD28, whereas a Q196H substitution mutant of SIVmacJ5 Nef was able to down-modulate cell surface CD28. Conversely, substitution of H196 to Q196 in SIVmac239 Nef resulted in a mutant that had minimal effect on cell surface CD28 expression, despite retaining the capacity to down-modulate cell surface CD3ϵ. H196 lies immediately adjacent to a documented di-leucine endocytic motif and mutation of this motif also abrogated the ability of SIVmac239 Nef to down-modulate CD28. These findings demonstrate that down-modulation of the costimulatory molecule, CD28, and clonotypic TCR/CD3 complex are conserved attributes of SIV Nef
The Hepatitis C Virus Nonstructural Protein 4B Is an Integral Endoplasmic Reticulum Membrane Protein

AbstractThe hepatitis C virus (HCV) nonstructural protein 4B (NS4B) is a relatively hydrophobic 27-kDa protein of unknown function. A tetracycline-regulated gene expression system, a novel monoclonal antibody, and in vitro transcription–translation were employed to investigate the subcellular localization and to characterize the membrane association of this viral protein. When expressed individually or in the context of the entire HCV polyprotein, NS4B was localized in the endoplasmic reticulum (ER), as shown by subcellular fractionation, immunofluorescence analyses, and double-label confocal laser scanning microscopy. In this compartment NS4B colocalized with the other HCV nonstructural proteins. Association of NS4B with the ER membrane occurred cotranslationally, presumably via engagement of the signal recognition particle by an internal signal sequence. In membrane extraction and proteinase protection assays NS4B displayed properties of a cytoplasmically oriented integral membrane protein. Taken together, our findings suggest that NS4B is a component of a membrane-associated cytoplasmic HCV replication complex. An efficient replication system will be essential to further define the role of NS4B in the viral life cycle
The Whitewater Arroyo Virus: Natural Evidence for Genetic Recombination among Tacaribe Serocomplex Viruses (Family Arenaviridae)

AbstractThe Tacaribe serocomplex (family Arenaviridae) comprises three phylogenetic lineages, designated A, B, and C. The sequence of a 3278-nt fragment of the small genomic segment of the Whitewater Arroyo (WWA) virus was determined to extend our knowledge on the phylogenetic relationship of this newly discovered North American Tacaribe complex virus to other arenaviruses. Independent analyses of full-length nucleoprotein (N) and glycoprotein precursor (GPC) amino acid sequences indicated that the WWA virus N and GPC genes are descended from a lineage A virus and lineage B virus, respectively. The different phylogenetic histories of the N and GPC genes indicate that the WWA virus genome is a product of recombination between two Tacaribe complex viruses
Human Immunodeficiency Virus Type 1 Clade A and D Neurotropism: Molecular Evolution, Recombination, and Coreceptor Use

AbstractHuman immunodeficiency virus type 1 (HIV-1) non-B clade viral infections of the brain have not been studied to date. Among nine AIDS patients from Nairobi, Kenya, infected with HIV-1 A (N = 5) or D (N = 4) clade strains, brain-derived HIV-1 env sequences displayed greater evolutionary distance than B clade brain-derived viruses (P < 0.001). Similarly, molecular diversity between matched brain and spleen env clones was clade-dependent and concentrated in the hypervariable V4 region (P < 0.001), with phylogenetic clustering of sequences derived from the same organ. Brain-derived A and D clade sequences displayed significantly lower ratios of nonsynonymous/synonymous substitution rates (dN/dS) compared to matched spleen-derived clones and brain-derived B clade viruses. Interclade recombination events were infrequently observed among the present env sequences. A chimeric virus containing the C2V3 region from an A clade brain-derived sequence preferentially used CD4 and CCR5 for infection. These findings demonstrate that differences in molecular diversity in brain-derived sequences were dependent on the individual clade and domain within the env gene, but both B and non-B clade brain-derived viruses exhibit a preference for CCR5 as a coreceptor
Inhibition of Coxsackievirus B4 Replication in Stably Transfected Cells Expressing Human MxA Protein

AbstractCoxsackieviruses B (CVB) (B1–B6), positive-strand RNA viruses, cause a variety of diseases. CVB4 may have a causal role in insulin-dependent diabetes mellitus. IFN-α inhibits CVB replication; however, the mechanism is not well known. The interferon-α-inducible human MxA protein exerts an antiviral activity against negative-strand RNA viruses and against Semliki Forest virus, a positive-strand RNA virus. To test the antiviral spectrum of MxA against CVB4, we took advantage of stably transfected Vero cells expressing MxA (Vero/MxA) in 98% of cells. Compared with control cells, in Vero/MxA cells, CVB4 yields were dramatically reduced and expression of the VP1 CVB protein analyzed by immunofluorescence was highly restricted. Furthermore, the accumulation of positive- and negative-strand CVB4 RNA was prevented as shown by in situ hybridization and RT-PCR. These results indicate that the antiviral activity of MxA extends to CVB4 and that its replication cycle is inhibited at an early step in Vero/MxA cells
Immunopathogenesis and Immune Modulation of Venezuelan Equine Encephalitis Virus-Induced Disease in the Mouse

AbstractThe course of Venezuelan equine encephalitis (VEE) disease in immunodeficient and immunologically normal mice was compared to define the role of the immune system in this disease process. Immunocompetent mice infected with VEE exhibited a biphasic illness characterized by an early self-limiting lymphoid phase and a fatal CNS phase. The lymphoid phase of the illness was characterized by extensive viral replication within spleen, thymus, Peyer's patches, and lymph nodes, was accompanied by a high-titered serum viremia, and resolved with the production of VEE-specific IgM class antibody at 72 h postinfection (p.i.). Immunocompetent animals survived an average of 6.8 ± 1.2 days before succumbing to fulminant encephalitis. In contrast, SCID mice infected with VEE showed a persistent replication of virus throughout all organs tested beginning at 24 h p.i. VEE-infected SCID mice exhibited a severe spongiform encephalopathy with 100% mortality and an average survival time of 8.9 ± 0.9 days. These studies indicated that the characteristic organ tropism of VEE in the mouse is due in large part to an early anti-viral state, the establishment of which is dependent upon the presence of an intact immune system. Finally, the CNS pathology in a VEE-infected mouse had a significant immunologic component. However, in contrast to other neurovirulent alphaviruses, VEE was directly cytopathic for the cells of the CNS, even in the absence of an immune response
Role of the LTR Region between the Enhancer and Promoter in Mink Cell Focus-Forming Murine Leukemia Virus Pathogenesis

AbstractLong terminal repeat (LTR) sequences are important determinants of mink cell focus-forming (MCF) murine leukemia virus pathogenesis. These sequences include the enhancer and sequences between the enhancer and promoter (DEN). In a previous study we showed that a virus missing the DEN region in its LTR was severely attenuated in its ability to induce thymic lymphoma. In this study we observed that a virus with an LTR consisting of DEN but no enhancer sequences was pathogenic. We compared the pathogenicity of this DEN virus with other LTR mutant MCF13 viruses that contained a single enhancer (1R) or a single enhancer plus DEN (1R + DEN). All LTR mutant viruses generated thymic lymphoma, however, at a much lower incidence and with a longer latency compared with wild-type (WT) MCF13 virus. DEN virus replication in the thymus was the lowest compared with the 1R and 1R + DEN viruses. Viral replication in a different thymic subpopulation could not explain the decreased pathogenicity of the LTR mutant viruses compared with WT virus. However, lower levels of mutant virus replication in the thymus compared with WT during the preleukemic period may contribute to the attenuation of pathogenicity. The phenotype of tumors induced by the mutant viruses was similar and differed from tumors induced by WT virus by the presence of CD3−CD4−CD8− cells. Analysis of LTR sequences of infectious virus rescued from tumors induced by the 1R and 1R + DEN viruses showed that amplification of enhancer sequences had occurred during tumor development. The lack of DEN virus expression by tumor cells led us to propose that DEN sequences may play a role at an early step in tumorigenesis
5′-Coterminal Subgenomic RNAs in Citrus Tristeza Virus-Infected Cells

AbstractThree unusual 5′ coterminal positive-stranded subgenomic (sg) RNAs, two of about 0.8 kb and one of 10 kb (designated LMT1, LMT2, and LaMT, respectively), from Citrus spp. plants and Nicotiana benthamiana protoplasts infected with Citrus tristeza virus (CTV) were characterized. The 5′ termini of the LMT RNAs were mapped by runoff reverse transcription and found to correspond with the 5′ terminus of the genomic RNA. The LMT 5′-coterminal sgRNAs consisted of two modal lengths of 744–746 and 842–854 nts. The 3′ of the LaMT RNAs terminated near the junction of ORF 1b and ORF 2 (p33). None of the 5′ sgRNAs had detectable amounts of corresponding negative-sense RNAs, as occurs with the genomic and 3′ coterminal subgenomic RNAs of CTV. The abundance of the short and long 5′ sgRNAs differed considerably in infected cells. The LMT RNAs were considerably more abundant than the genomic RNAs, while the larger LaMT RNA accumulated to much lower levels. The kinetics of accumulation of LMT1 and LMT2 in synchronously infected protoplasts differed. The larger RNA, LMT1, accumulated earlier with a strong hybridization signal at 2 days postinfection, a time when only traces of genomic and 3′ sgRNAs were detected. The lack of corresponding RNAs, that could be 3′ cleavage products corresponding to the 5′ coterminal sgRNAs and the lack of complementary negative strands, suggest that these sgRNAs were produced by termination during the synthesis of the genomic positive strands
Two Regions of the P Protein Are Required to be Active with the L Protein for Human Parainfluenza Virus Type 1 RNA Polymerase Activity

AbstractThe paramyxovirus P protein is an essential component of the viral RNA polymerase composed of P and L proteins. In this study, we characterized the physical and functional interactions between P and L proteins using human parainfluenza virus type 1 (hPIV1) and its counterpart Sendai virus (SV). The hPIV1 P and SV L proteins or the SV P and hPIV1 L proteins formed complexes detected by anti-P antibodies. Functional analysis using the minigenome SV RNA containing CAT gene indicated that the hPIV1 P–SV L complex, but not the SV P–hPIV1 L complex, was biologically active. Mutant SV P or hPIV1 P cDNAs, which do not express C proteins, showed the same phenotype with wild-type P cDNAs, indicating that C proteins are not responsible for the dysfunction of SV P–hPIV1 L polymerase complex. Using the chimeric hPIV1/SV P cDNAs, we identified two regions (residues 387–423 and 511–568) on P protein, which are required for the functional interaction with hPIV1 L. These regions overlap with a previously identified domain for oligomer formation and binding to nucleocapsids. Our results indicate that in addition to a P–L binding domain, hPIV1 L requires a specific region on P protein to be biologically functional as a polymerase
How Hepatitis C Virus Counteracts the Interferon Response: The Jury Is Still out on NS5A

AbstractInterferons (IFNs) induce an antiviral state in the cell through complex and indirect mechanisms, which culminate in a direct inhibition of viral replication and stimulation of the host adaptive responses. Viruses often counteract with elaborate strategies to interfere with the induction as well as action of IFN effector molecules. This evolutionary battle between viruses and IFN components is a subject of intense research aimed at understanding the immunopathogenesis of viruses and the molecular basis of IFN signaling and action. In the case with hepatitis C virus (HCV), this may have profound implications for the therapeutic use of recombinant IFN in treating chronic hepatitis C. Depending on the subtype of HCV, current IFN-based treatment regimens are effective for only a small subset of chronic hepatitis C patients. Thus, one of the Holy Grails in HCV research is to understand the mechanisms by which the virus may evade IFN antiviral surveillance and establish persistent infection, which may eventually provide insights into new avenues for better antiviral therapy. Despite the lack of an efficient tissue culture system and an appropriate animal model for HCV infection, several mechanisms have been proposed based on clinical studies and in vitro experiments. This minireview focuses on the HCV NS5A nonstructural protein, which is implicated in playing a role in HCV tolerance to IFN treatment, possibly in part through its ability to inhibit the cellular IFN-induced PKR protein kinase
Membrane Relocation but Not Tight Binding of Human Immunodeficiency Virus Type 1 Gag Particles Myristoylated in Escherichia coli

AbstractExpression of human immunodeficiency virus Gag protein and the N-terminal matrix (MA) domain in Escherichia coli yielded spherical structures in the cytoplasm. When human N-myristoyltransferase was coexpressed, both Gag and MA were fully myristoylated and spherical structures were relocated in close proximity to the cytoplasmic membrane. However, neither myristoylated Gag nor MA exhibited tight binding to E. coli membrane, suggesting that myristoylation in E. coli did not confer membrane affinity on Gag despite the relocation. Our data also suggest that the morphogenetic pathway of Gag particles in prokaryotic cells differs from that in eukaryotic cells despite biochemical similarities of in the form of Gag expressed
Evolution of AZT Resistance in HIV-1: The 41-70 Intermediate That Is Not Observed in Vivo Has a Replication Defect

AbstractThe human immunodeficiency virus type 1 (HIV-1) is notorious for its ability to evolve drug-resistance in patients treated with potent antivirals. Resistance to inhibitors of the viral reverse transcriptase (RT) enzyme is frequently mediated by a single amino acid substitution within RT. Resistance against the nucleoside analogue AZT is remarkable in that multiple amino acid changes accumulate over time to yield virus variants with high-level drug resistance. We now report that in addition to drug-resistance properties, the relative replication capacity of the virus variants affects the evolution of AZT resistance. Some of the typical AZT-resistance mutations have a negative impact on virus replication, and the 41-70 double mutant was found to represent a particularly poor virus. Furthermore, introduction of additional AZT-resistance mutations (41-70-215) leads to nearly complete restoration of virus replication. These results may explain the absence of the 41-70 double mutant in clinical samples and indicate that the evolution of AZT resistance is also influenced by virus replication parameters. Prolonged passage of the replication-impaired 41-70 virus in the absence of AZT yielded several fast-replicating variants. These revertants have compensatory changes in the RT polymerase, some of which have been observed previously in AZT-treated patients. Because we could select for these changes without drug pressure, these changes are likely to improve the RT enzyme function and the HIV-1 replication capacity