bluuebunny/crossref_metadata_2025_split huggingface dataset:
Two Retroviral Entry Pathways Distinguished by Lipid Raft Association of the Viral Receptor and Differences in Viral Infectivity
Authors: Narayan Shakti , Barnard Richard J. O. , Young John A. T. 
Published: January 2003

ABSTRACT

The receptor “priming” model for entry of the retrovirus avian sarcoma and leukosis virus (ASLV) predicts that upon binding cell surface receptors, virions are endocytosed and trafficked to acidic endosomes where fusion occurs. To test this model directly, we have now followed subgroup A ASLV (ASLV-A) virions entering cells via either the transmembrane (TVA950) or glycophosphatidylinositol (GPI)-anchored (TVA800) forms of the cellular receptor. Our results suggest that viruses entering via these two forms of receptor are subjected to different intracellular fates, perhaps due to use of different endocytic trafficking pathways to access acidic fusion compartments. Kinetic analyses demonstrated that virus bound to TVA800 was taken up from the cell surface more slowly but then trafficked to the site of fusion more quickly than that entering via TVA950. Furthermore, transiently arresting virions within putative fusion compartments with NH
4
Cl led to a substantially greater decrease in the infectivity of virions using TVA950 than with those using TVA800. The increased infectivity of virions using TVA800 correlated with the localization of this receptor to lipid rafts, since this effect was abolished by pharmacological disruption of lipid rafts. Together these results suggest that, in the presence of NH
4
Cl, virus bound to the GPI-anchored receptor may utilize a lipid raft-dependent pathway to accumulate within a fusion compartment where it is more stable than if it enters via the transmembrane receptor. The TVA800/ASLV-A system should prove useful for the molecular analysis of lipid raft-dependent endocytosis and may provide a tool for the biochemical dissection of the poorly understood uncoating step of retroviral replication.
Replication of Human Immunodeficiency Viruses Engineered with Heterologous Tat-Transactivation Response Element Interactions
Authors: Xie Baode , Wainberg Mark A. , Frankel Alan D. 
Published: January 2003

ABSTRACT
Human immunodeficiency viruses (HIVs) and the related bovine lentiviruses bovine immunodeficiency virus (BIV) and Jembrana disease virus (JDV) utilize the viral Tat protein to activate viral transcription. The arginine-rich RNA-binding domains of the Tat proteins bind to their cognate transactivation response element (TAR) RNA hairpins located at the 5′ ends of the viral mRNAs, resulting in enhanced processivity of RNA polymerase II. It has previously been shown that HIV type 1 (HIV-1) Tat requires the cellular cyclin T1 protein for high-affinity RNA binding whereas BIV Tat and JDV Tat bind with high affinity on their own and adopt distinct β-hairpin conformations when complexed to RNA. Here we have engineered the BIV and JDV Tat-TAR interactions into HIV-1 and show that the heterologous interactions support viral replication, correlating well with their RNA-binding affinities. Viruses engineered with a variant TAR able to bind all three Tat proteins replicate efficiently with any of the proteins. In one virus containing a noncognate Tat-TAR pair that neither interacts nor efficiently replicates (HIV-1 TAR and BIV Tat), viral revertants were isolated in which TAR had become mutated to generate a functional BIV Tat binding site. Our results support the view that incremental changes to TAR structure can provide routes for evolving new Tat-TAR complexes while maintaining active viral replication.
Human Papillomavirus Type 16 E2 Protein Has No Effect on Transcription from Episomal Viral DNA
Authors: Bechtold Viviane , Beard Peter , Raj Kenneth 
Published: January 2003

ABSTRACT
The human papillomavirus (HPV) E2 protein plays an important role in viral DNA replication. Many studies with high-risk HPVs have demonstrated that the E2 protein can also repress transcription of the E6 and E7 oncogenes. This conclusion, based on experiments carried out with cervical cancer cells bearing integrated HPV genomes, is currently assumed to be applicable to the normal HPV life cycle, in which the viral genomes are episomal. Here, we have tested experimentally whether this assumption is correct. We made use of a pair of isogenic cell lines, W12 and S12. W12 cells contain episomal HPV16 genomes, whereas S12 cells, which are derived from the W12 line, contain HPV DNA as integrated copies. When we expressed E2 in S12 cells, we observed strong repression of E6 and E7 transcription. In contrast, no effect of E2 on the transcription of these genes was detected in W12 cells. While integration of the viral genome into the host DNA contributes to the difference between W12 and S12 cells, integration by itself is not sufficient to explain this difference. Instead, the chromatin structure in the region of the E6 and E7 promoter (p97), which we show to be very different in these two cell lines, is likely to be the cause of the different responsiveness of p97 to the E2 protein. Experiments with the histone deacetylase inhibitor trichostatin A (TSA) indicated that the episomal HPV16 DNA is in a relatively inaccessible state prior to TSA treatment. Our results, together with those of others, suggest that any effect of the E2 protein on the expression of the E6 and E7 genes during the normal viral life cycle is of secondary importance compared to the function of E2 in replication.
Depletion of Wee-1 Kinase Is Necessary for both Human Immunodeficiency Virus Type 1 Vpr- and Gamma Irradiation-Induced Apoptosis
Authors: Yuan Huidong , Xie Yi-Ming , Chen Irvin S. Y. 
Published: January 2003

ABSTRACT

Human immunodeficiency virus (HIV) protein R (Vpr) induces G
2
arrest, and prolonged G
2
arrest leads to apoptosis. We find that in HeLa cells the cell cycle regulatory kinase, Wee-1, is depleted following prolonged G
2
arrest induced by Vpr. Of note, small interfering RNAs directed to Wee-1 triggered apoptosis, suggesting a direct role for Wee-1 in apoptosis. In support of this hypothesis, overexpression of Wee-1 suppressed Vpr-mediated apoptosis. Importantly, similar results were observed with cells induced to undergo apoptosis gamma irradiation. Thus, Wee-1 may serve as a key regulator of both HIV type 1 Vpr- and gamma irradiation-mediated apoptosis and possibly serve as a general regulator linking the cell cycle to some pathways of apoptosis.
Influence of Reverse Transcriptase Variants, Drugs, and Vpr on Human Immunodeficiency Virus Type 1 Mutant Frequencies
Authors: Mansky Louis M. , Le Rouzic Erwann , Benichou Serge , Gajary Lisa C. 
Published: January 2003

ABSTRACT

The evolution of drug resistance is a major complication of human immunodeficiency virus type 1 (HIV-1) chemotherapy. HIV-1 reverse transcriptase (RT) is a major target of antiretroviral therapy and ultimately the target of drug resistance mutations. Previous studies have indicated that drug-resistant HIV-1 RTs can alter HIV-1 mutant frequencies. In this study, we have tested a panel of HIV-1 RT variants for their ability to influence virus mutant frequencies. The RT variants tested included drug-resistant RT variants as well as other variants analyzed in enzyme fidelity studies with the
lacZ
α gene as a mutation target and/or implicated as being important for enzyme fidelity by structural studies. Combinations of mutations that alone had a statistically significant influence on virus mutant frequencies resulted in different mutant frequency phenotypes. Furthermore, when virus replication occurred in the presence of drugs [e.g., 3′-azido-3′-deoxythymidine, (−)2/,3′-dideoxy-3′-thiacytidine, hydroxyurea, thymidine, or thioguanine] with selected RT variants, virus mutant frequencies increased. Similarly, Vpr variants deficient for binding to the uracil DNA glycosylase repair enzyme were observed to influence HIV-1 virus mutant frequencies when tested alone or in combination with RT variants. In summary, these observations indicate that HIV-1 mutant frequencies can significantly change by single amino acid substitutions in RT and that these effects can be altered by additional mutations in RT, by drugs, and/or by expression of Vpr variants. Such altered virus mutant frequencies could impact HIV-1 dynamics and evolution in small population sizes.
Comprehensive Epitope Analysis of Human Immunodeficiency Virus Type 1 (HIV-1)-Specific T-Cell Responses Directed against the Entire Expressed HIV-1 Genome Demonstrate Broadly Directed Responses, but No Correlation to Viral Load
Authors: Addo M. M. , Yu X. G. , Rathod A. , Cohen D. , Eldridge R. L. , Strick D. , Johnston M. N. , Corcoran C. , Wurcel A. G. , Fitzpatrick C. A. , Feeney M. E. , Rodriguez W. R. , Basgoz N. , Draenert R. , Stone David R. , Brander C. , Goulder P. J. R. , Rosenberg E. S. , Altfeld M. , Walker B. D. 
Published: January 2003

ABSTRACT

Cellular immune responses play a critical role in the control of human immunodeficiency virus type 1 (HIV-1); however, the breadth of these responses at the single-epitope level has not been comprehensively assessed. We therefore screened peripheral blood mononuclear cells (PBMC) from 57 individuals at different stages of HIV-1 infection for virus-specific T-cell responses using a matrix of 504 overlapping peptides spanning all expressed HIV-1 proteins in a gamma interferon-enzyme-linked immunospot (Elispot) assay. HIV-1-specific T-cell responses were detectable in all study subjects, with a median of 14 individual epitopic regions targeted per person (range, 2 to 42), and all 14 HIV-1 protein subunits were recognized. HIV-1 p24-Gag and Nef contained the highest epitope density and were also the most frequently recognized HIV-1 proteins. The total magnitude of the HIV-1-specific response ranged from 280 to 25,860 spot-forming cells (SFC)/10
6
PBMC (median, 4,245) among all study participants. However, the number of epitopic regions targeted, the protein subunits recognized, and the total magnitude of HIV-1-specific responses varied significantly among the tested individuals, with the strongest and broadest responses detectable in individuals with untreated chronic HIV-1 infection. Neither the breadth nor the magnitude of the total HIV-1-specific CD8
+
-T-cell responses correlated with plasma viral load. We conclude that a peptide matrix-based Elispot assay allows for rapid, sensitive, specific, and efficient assessment of cellular immune responses directed against the entire expressed HIV-1 genome. These data also suggest that the impact of T-cell responses on control of viral replication cannot be explained by the mere quantification of the magnitude and breadth of the CD8
+
-T-cell response, even if a comprehensive pan-genome screening approach is applied.
Replication of
Carnation Italian Ringspot Virus
Defective Interfering RNA in
Saccharomyces cerevisiae
Authors: Pantaleo Vitantonio , Rubino Luisa , Russo Marcello 
Published: January 2003

ABSTRACT

Two plasmids from which the sequences coding for the 36- and 95-kDa proteins of
Carnation Italian ringspot virus
(CIRV) could be transcribed in vivo in the yeast
Saccharomyces cerevisiae
under the control of the ADH1 promoter and terminator were constructed. The two proteins, which constitute the viral replicase, were correctly translated and integrated into membranes of the yeast cells. An additional plasmid was introduced in yeasts expressing the CIRV replicase, from which a defective interfering (DI) RNA (DI-7 RNA) could be transcribed under the control of the GAL1 promoter and terminated by the
Tobacco ringspot virus
satellite ribozyme, which cleaved 19 nucleotides downstream of the 3′ end of DI RNA. The DI-7 RNA transcripts were amplified by the viral replicase as demonstrated by the restoration of the authentic 3′ end, the requirement of a specific
cis
-acting signal at this terminus, the preferential accumulation of molecules with the authentic 5′ terminus (AGAAA), the synthesis of head-to-tail dimers, the presence of negative strands, and the incorporation of 5-bromo-UTP. Additionally, transformation with a dimeric construct of DI-7 RNA led to the synthesis of monomers, mimicking the activity of the viral replicase in plant cells.
Human Immunodeficiency Virus Type 1 Nef-Mediated Downregulation of CD4 Correlates with Nef Enhancement of Viral Pathogenesis
Authors: Stoddart Cheryl A. , Geleziunas Romas , Ferrell Sharon , Linquist-Stepps Valerie , Moreno Mary E. , Bare Christopher , Xu Weiduan , Yonemoto Wes , Bresnahan Patricia A. , McCune Joseph M. , Greene Warner C. 
Published: January 2003

ABSTRACT

The
nef
gene products encoded by human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus type 1 (SIV-1) increase viral loads in infected hosts and accelerate clinical progression to AIDS. Nef exhibits a spectrum of biological activities, including the ability to downregulate surface expression of CD4 and major histocompatibility complex (MHC) class I antigens, to alter the state of T-cell activation, and to enhance the infectivity of viral particles. To determine which of these in vitro functions most closely correlates with the pathogenic effects of Nef in vivo, we constructed recombinant HIV-1 NL4-3 viruses carrying mutations within the
nef
gene that selectively impair these functions. These mutant viruses were evaluated for pathogenic potential in severe combined immunodeficiency (SCID) mice implanted with human fetal thymus and liver (SCID-hu Thy/Liv mice), in which virus-mediated depletion of thymocytes is known to be Nef dependent. Disruption of the polyproline type II helix (Pxx)
4
within Nef (required for binding of Hck and p21-activated kinase-like kinases, downregulation of MHC class I, and enhancement of HIV-1 infectivity in vitro but dispensable for CD4 downregulation) did not impair thymocyte depletion in virus-infected Thy/Liv human thymus implants. Conversely, three separate point mutations in Nef that compromised its ability to downregulate CD4 attenuated thymocyte depletion while not diminishing viral replication. These findings indicate that the functional ability of Nef to downregulate CD4 and not MHC class I downregulation, Hck or PAK binding, or (Pxx)
4
-associated enhancement of infectivity most closely correlates with Nef-mediated enhancement of HIV-1 pathogenicity in vivo. Nef-mediated CD4 downregulation merits consideration as a new target for the development of small-molecule inhibitors.
Virus Replication in Engineered Human Cells That Do Not Respond to Interferons
Authors: Young D. F. , Andrejeva L. , Livingstone A. , Goodbourn S. , Lamb R. A. , Collins P. L. , Elliott R. M. , Randall R. E. 
Published: January 2003

ABSTRACT
The V protein of the paramyxovirus simian virus 5 blocks interferon (IFN) signaling by targeting STAT1 for proteasome-mediated degradation. Here we report on the isolation of human cell lines that express the V protein and can no longer respond to IFN. A variety of viruses, particularly slow-growing wild-type viruses and vaccine candidate viruses (which are attenuated due to mutations that affect virus replication, virus spread, or ability to circumvent the IFN response), form bigger plaques and grow to titers that are increased as much as 10- to 4,000-fold in these IFN-nonresponsive cells. We discuss the practical applications of using such cells in vaccine development and manufacture, virus diagnostics and isolation of newly emerging viruses, and studies on host cell tropism and pathogenesis.
Cytomegalovirus-Mediated Upregulation of Chemokine Expression Correlates with the Acceleration of Chronic Rejection in Rat Heart Transplants
Authors: Streblow Daniel N. , Kreklywich Craig , Yin Qiang , De La Melena V. T. , Corless Christopher L. , Smith Patricia A. , Brakebill Christina , Cook Judith W. , Vink Cornelis , Bruggeman Cathrien A. , Nelson Jay A. , Orloff Susan L. 
Published: January 2003

ABSTRACT
Cytomegalovirus (CMV) infections have been shown to dramatically affect solid organ transplant graft survival in both human and animal models. Recently, it was demonstrated that rat CMV (RCMV) infection accelerates the development of transplant vascular sclerosis (TVS) in both rat heart and small bowel graft transplants. However, the mechanisms involved in this process are still unclear. In the present study, we determined the kinetics of RCMV-accelerated TVS in a rat heart transplant model. Acute RCMV infection enhances the development of TVS in rat heart allografts, and this process is initiated between 21 and 24 days posttransplantation. The virus is consistently detected in the heart grafts from day 7 until day 35 posttransplantation but is rarely found at the time of graft rejection (day 45 posttransplantation). Grafts from RCMV-infected recipients had upregulation of chemokine expression compared to uninfected controls, and the timing of this increased expression paralleled that of RCMV-accelerated neointimal formation. In addition, graft vessels from RCMV-infected grafts demonstrate the increased infiltration of T cells and macrophages during periods of highest chemokine expression. These results suggest that CMV-induced acceleration of TVS involves the increased graft vascular infiltration of inflammatory cells through enhanced chemokine expression.
The E6 and E7 Proteins of the Cutaneous Human Papillomavirus Type 38 Display Transforming Properties
Authors: Caldeira Sandra , Zehbe Ingeborg , Accardi Rosita , Malanchi Ilaria , Dong Wen , Giarrè Marianna , de Villiers Ethel-Michele , Filotico Raffaele , Boukamp Petra , Tommasino Massimo 
Published: January 2003

ABSTRACT

Several studies have suggested the involvement of cutaneous human papillomaviruses (HPVs) in the development of nonmelanoma skin cancers. Here we have characterized the in vitro properties of E7 proteins of three cutaneous HPV types, 10, 20, and 38, which are frequently detected in skin specimens. We show that HPV38 E7 is able to inactivate the tumor suppressor pRb and induces loss of G
1
/S transition control, a key event in carcinogenesis. In contrast, HPV10 and HPV20 E7 proteins do not display these in vitro transforming activities. We also show that the two early proteins E6 and E7 of HPV38 are sufficient to corrupt the cell cycle and senescence programs in primary cells, inducing active and long-lasting proliferation of primary human keratinocytes, the natural host cells. Our study shows that E6 and E7 of this cutaneous HPV type have transforming activity in primary human cells, suggesting a role for HPV38 infection in skin carcinogenesis. In further support of such a role, we detected HPV38 DNA in approximately 50% of nonmelanoma skin cancers, but only in 10% of healthy skin specimens (
P
< 0.001).
The Promyelocytic Leukemia Protein Does Not Mediate Foamy Virus Latency In Vitro
Authors: Meiering Christopher D. , Linial Maxine L. 
Published: January 2003

ABSTRACT
Spumaviruses, commonly called foamy viruses, are complex retroviruses that establish life-long persistent infections in the absence of accompanying pathology. Depending upon cell type, infection of cells in tissue culture cells can result in either lytic replication, persistence, or latency. The cellular factors that mediate foamy virus (FV) latency are poorly understood. In this study we show that the only known inhibitor of FV replication, the promyelocytic leukemia protein (PML), which binds the FV transactivator (Tas), does not play an important role in FV latency in vitro. We found no significant differences in PML levels in cells that supported lytic replication compared to those that were latently infected. Furthermore, endogenous PML levels did not change following exposure to phorbol myristate acetate (PMA), which induces FV replication. We demonstrated that FV replication proceeded in the presence of substantial levels of PML, both in fully permissive cells and during reactivation of latent FV. Endogenous PML did not efficiently colocalize with Tas, even after upregulation by alpha interferon (IFN-α) treatment. IFN-α did, however, partially suppress the reactivation of latent FV by PMA. Finally, depletion of endogenous PML by small interfering RNA did not promote activation of FV in cells that responded to PMA treatment. Taken together, these data indicate that endogenous PML does not play an important role in mediating FV latency.
Frequency and Phenotype of Circulating Vα24/Vβ11 Double-Positive Natural Killer T Cells during Hepatitis C Virus Infection
Authors: Lucas Michaela , Gadola Stephan , Meier Ute , Young Neil T. , Harcourt Gillian , Karadimitris Anastasios , Coumi Nikki , Brown Dave , Dusheiko Geoff , Cerundolo Vincenzo , Klenerman Paul 
Published: January 2003

ABSTRACTNatural killer T (NKT) cells are thought to be involved in innate responses against infection. We investigated one specific type of NKT cell, Vα24/Vβ11 double positive, in hepatitis C virus (HCV) infection. Lower frequencies of this population were detected in the blood of HCV PCR-positive patients than in controls. Unlike Vα24/Vβ11 NKT cells found in blood, those in the liver appeared to be recently activated.
Recognition of a Novel Stage of Betaherpesvirus Latency in Human Herpesvirus 6
Authors: Kondo Kazuhiro , Sashihara Junji , Shimada Kazuya , Takemoto Masaya , Amo Kiyoko , Miyagawa Hiromi , Yamanishi Koichi 
Published: January 2003

ABSTRACT

Latency-associated transcripts of human herpesvirus 6 (H6LTs) (K. Kondo et al. J. Virol.
76:
4145-4151, 2002) were maximally expressed at a fairly stable intermediate stage between latency and reactivation both in vivo and in vitro. H6LTs functioned as sources of immediate-early protein 1 at this stage, which up-regulated the viral reactivation.
Fine Mapping of a
cis
-Acting Sequence Element in Yellow Fever Virus RNA That Is Required for RNA Replication and Cyclization
Authors: Corver Jeroen , Lenches Edith , Smith Kayla , Robison R. Aaron , Sando Trisha , Strauss Ellen G. , Strauss James H. 
Published: January 2003

ABSTRACT

We present fine mapping of a
cis
-acting nucleotide sequence found in the 5′ region of yellow fever virus genomic RNA that is required for RNA replication. There is evidence that this sequence interacts with a complementary sequence in the 3′ region of the genome to cyclize the RNA. Replicons were constructed that had various deletions in the 5′ region encoding the capsid protein and were tested for their ability to replicate. We found that a sequence of 18 nucleotides (residues 146 to 163 of the yellow fever virus genome, which encode amino acids 9 to 14 of the capsid protein) is essential for replication of the yellow fever virus replicon and that a slightly longer sequence of 21 nucleotides (residues 146 to 166, encoding amino acids 9 to 15) is required for full replication. This region is larger than the core sequence of 8 nucleotides conserved among all mosquito-borne flaviviruses and contains instead the entire sequence previously proposed to be involved in cyclization of yellow fever virus RNA.
In a Subset of Subjects on Highly Active Antiretroviral Therapy, Human Immunodeficiency Virus Type 1 RNA in Plasma Decays from 50 to <5 Copies per Milliliter, with a Half-Life of 6 Months
Authors: Di Mascio Michele , Dornadula Geethanjali , Zhang Hui , Sullivan Julie , Xu Yan , Kulkosky Joseph , Pomerantz Roger J. , Perelson Alan S. 
Published: January 2003

ABSTRACT
Three of five virally suppressed human immunodeficiency virus type I (HIV-1)-infected patients treated with highly active antiretroviral therapy and followed intensively with a supersensitive reverse transcriptase PCR assay with a lower limit of quantitation of 5 copies/ml showed statistically significant viral load decays below 50 copies/ml, with half-lives of 5 to 8 months and a mean of 6 months. This range of half-lives is consistent with the estimated half-life of the latent HIV-1 reservoir in the peripheral blood. Those patients without decay of viral load in plasma may have significant cryptic HIV-1 residual replication.