bluuebunny/crossref_metadata_2025_split huggingface dataset:
Vectorial Entry and Release of Hepatitis A Virus in Polarized Human Hepatocytes
Authors: Snooks Michelle J. , Bhat Purnima , Mackenzie Jason , Counihan Natalie A. , Vaughan Nicola , Anderson David A. 
Published: June 2008

ABSTRACT
Hepatitis A virus (HAV) is an enterically transmitted virus that replicates predominantly in hepatocytes within the liver before excretion via bile through feces. Hepatocytes are polarized epithelial cells, and it has been assumed that the virus load in bile results from direct export of HAV via the apical domain of polarized hepatocytes. We have developed a subclone of hepatocyte-derived HepG2 cells (clone N6) that maintains functional characteristics of polarized hepatocytes but displays morphology typical of columnar epithelial cells, rather than the complex morphology that is typical of hepatocytes. N6 cells form microcolonies of polarized cells when grown on glass and confluent monolayers of polarized cells on semipermeable membranes. When N6 microcolonies were exposed to HAV, infection was restricted to peripheral cells of polarized colonies, whereas all cells could be infected in colonies of nonpolarized HepG2 cells (clone C11) or following disruption of tight junctions in N6 colonies with EGTA. This suggests that viral entry occurs predominantly via the basolateral plasma membrane, consistent with uptake of virus from the bloodstream after enteric exposure, as expected. Viral export was also found to be markedly vectorial in N6 but not C11 cells. However, rather than being exported from the apical domain as expected, more than 95% of HAV was exported via the basolateral domain of N6 cells, suggesting that virus is first excreted from infected hepatocytes into the bloodstream rather than to the biliary tree. Enteric excretion of HAV may therefore rely on reuptake and transcytosis of progeny HAV across hepatocytes into the bile. These studies provide the first example of the interactions between viruses and polarized hepatocytes.
Ligand-Independent Exhaustion of Killer Immunoglobulin-Like Receptor-Positive CD8
+
T Cells in Human Immunodeficiency Virus Type 1 Infection
Authors: Alter Galit , Rihn Suzannah , Streeck Hendrik , Teigen Nickolas , Piechocka-Trocha Alicja , Moss Kristin , Cohen Kristen , Meier Angela , Pereyra Florencia , Walker Bruce , Altfeld Marcus 
Published: June 2008

ABSTRACT

Virus-specific CD8
+
T cells play a central role in the control of viral infections, including human immunodeficiency virus type 1 (HIV-1) infection. However, despite the presence of strong and broad HIV-specific CD8
+
T-cell responses in chronic HIV-1 infection, these cells progressively lose critical effector functions and fail to clear the infection. Mounting evidence suggests that the upregulation of several inhibitory regulatory receptors on the surface of CD8
+
T cells during HIV-1 infection may contribute directly to the impairment of T-cell function. Here, we investigated the role of killer immunoglobulin receptors (KIR), which are expressed on NK cells and on CD8
+
T cells, in regulating CD8
+
T-cell function in HIV-1 infection. KIR expression was progressively upregulated on CD8
+
T cells during HIV-1 infection and correlated with the level of viral replication. Expression of KIR was associated with a profound inhibition of cytokine secretion, degranulation, proliferation, and activation by CD8
+
T cells following stimulation with T-cell receptor (TCR)-dependent stimuli. In contrast, KIR
+
CD8
+
T cells responded potently to TCR-independent stimulation, demonstrating that these cells are functionally competent. KIR-associated suppression of CD8
+
T-cell function was independent of ligand engagement, suggesting that these regulatory receptors may constitutively repress TCR activation. This ligand-independent repression of TCR activation of KIR
+
CD8
+
T cells may represent a significant barrier to therapeutic interventions aimed at improving the quality of the HIV-specific CD8
+
T-cell response in infected individuals.
Association of Respiratory Syncytial Virus M Protein with Viral Nucleocapsids Is Mediated by the M2-1 Protein
Authors: Li Dongsheng , Jans David A. , Bardin Phillip G. , Meanger Jayesh , Mills John , Ghildyal Reena 
Published: June 2008

ABSTRACT
Cytoplasmic inclusions in respiratory syncytial virus-infected cells comprising viral nucleocapsid proteins (L, N, P, and M2-1) and the viral genome are sites of viral transcription. Although not believed to be necessary for transcription, the matrix (M) protein is also present in these inclusions, and we have previously shown that M inhibits viral transcription. In this study, we have investigated the mechanisms for the association of the M protein with cytoplasmic inclusions. Our data demonstrate for the first time that the M protein associates with cytoplasmic inclusions via an interaction with the M2-1 protein. The M protein colocalizes with M2-1 in the cytoplasm of cells expressing only the M and M2-1 proteins and directly interacts with M2-1 in a cell-free binding assay. Using a cotransfection system, we confirmed that the N and P proteins are sufficient to form cytoplasmic inclusions and that M2-1 localizes to these inclusions; additionally, we show that M associates with cytoplasmic inclusions only in the presence of the M2-1 protein. Using truncated mutants, we show that the N-terminal 110 amino acids of M mediate the interaction with M2-1 and the subsequent association with nucleocapsids. The interaction of M2-1 with M and, in particular, the N-terminal region of M may represent a target for novel antivirals that block the association of M with nucleocapsids, thereby inhibiting virus assembly.
Changes in p19
Arf
Localization Accompany Apoptotic Crisis during Pre-B-Cell Transformation by Abelson Murine Leukemia Virus
Authors: Zimmerman Rebekah Stackpole , Rosenberg Naomi 
Published: June 2008

ABSTRACT

Transformation by Abelson murine leukemia virus (Ab-MLV) is a multistep process in which growth-stimulatory signals from the v-Abl oncoprotein and growth-suppressive signals from the p19
Arf
-p53 tumor suppressor pathway oppose each other and influence the outcome of infection. The process involves a proliferative phase during which highly viable primary transformants expand, followed by a period of marked apoptosis (called “crisis”) that is dependent on the presence of p19
Arf
and p53; rare cells that survive this phase emerge as fully transformed and malignant. To understand the way in which v-Abl expression affects p19
Arf
expression, we examined changes in expression of
Arf
during all stages of Ab-MLV transformation process. As is consistent with the ability of v-Abl to stimulate Myc, a transcription factor known to induce p19
Arf
,
Myc
and
Arf
are induced soon after infection and p19
Arf
is expressed. At these early time points, the infected cells remain highly viable. The onset of crisis is marked by an increase in p19
Arf
expression and a change in localization of the protein from the nucleoplasm to the nucleolus. These data together suggest that the localization and expression levels of p19
Arf
modulate the effects of the protein during oncogenesis and reveal that the p19
Arf
-mediated response is subject to multiple layers of regulation that influence its function during Ab-MLV-mediated transformation.
Structural Domains within the 3′ Untranslated Region of Turnip Crinkle Virus
Authors: McCormack John C. , Yuan Xuefeng , Yingling Yaroslava G. , Kasprzak Wojciech , Zamora Rodolfo E. , Shapiro Bruce A. , Simon Anne E. 
Published: June 2008

ABSTRACT

The genomes of positive-strand RNA viruses undergo conformational shifts that complicate efforts to equate structures with function. We have initiated a detailed analysis of secondary and tertiary elements within the 3′ end of
Turnip crinkle virus
(TCV) that are required for viral accumulation in vivo. MPGAfold, a massively parallel genetic algorithm, suggested the presence of five hairpins (H4a, H4b, and previously identified hairpins H4, H5, and Pr) and one H-type pseudoknot (Ψ
3
) within the 3′-terminal 194 nucleotides (nt). In vivo compensatory mutagenesis analyses confirmed the existence of H4a, H4b, Ψ
3
and a second pseudoknot (Ψ
2
) previously identified in a TCV satellite RNA. In-line structure probing of the 194-nt fragment supported the coexistence of H4, H4a, H4b, Ψ
3
and a pseudoknot that connects H5 and the 3′ end (Ψ
1
). Stepwise replacements of TCV elements with the comparable elements from
Cardamine chlorotic fleck virus
indicated that the complete 142-nt 3′ end, and subsets containing Ψ
3
, H4a, and H4b or Ψ
3
, H4a, H4b, H5, and Ψ
2
, form functional domains for virus accumulation in vivo. A new 3-D molecular modeling protocol (RNA2D3D) predicted that H4a, H4b, H5, Ψ
3
, and Ψ
2
are capable of simultaneous existence and bears some resemblance to a tRNA. The related
Japanese iris necrotic ring virus
does not have comparable domains. These results provide a framework for determining how interconnected elements participate in processes that require 3′ untranslated region sequences such as translation and replication.
Loss of the N-Linked Glycan at Residue 173 of Human Parainfluenza Virus Type 1 Hemagglutinin-Neuraminidase Exposes a Second Receptor-Binding Site
Authors: Alymova Irina V. , Taylor Garry , Mishin Vasiliy P. , Watanabe Makiko , Murti K. Gopal , Boyd Kelli , Chand Pooran , Babu Y. Sudhakara , Portner Allen 
Published: June 2008

ABSTRACT

BCX 2798 (4-azido-5-isobutyrylamino-2,3-didehydro-2,3,4,5-tetradeoxy-
d
-glycero-
d
-galacto-2-nonulopyranosic acid) effectively inhibited the activities of the hemagglutinin-neuraminidase (HN) of human parainfluenza viruses (hPIV) in vitro and protected mice from lethal infection with a recombinant Sendai virus whose HN was replaced with that of hPIV-1 (rSeV[hPIV-1HN]) (I. V. Alymova, G. Taylor, T. Takimoto, T. H. Lin., P. Chand, Y. S. Babu, C. Li, X. Xiong, and A. Portner, Antimicrob. Agents Chemother. 48:1495-1502, 2004). The ability of BCX 2798 to select drug-resistant variants in vivo was examined. A variant with an Asn-to-Ser mutation at residue 173 (N173S) in HN was recovered from mice after a second passage of rSeV(hPIV-1HN) in the presence of BCX 2798 (10 mg/kg of body weight daily). The N173S mutant remained sensitive to BCX 2798 in neuraminidase inhibition assays but was more than 10,000-fold less sensitive to the compound in hemagglutination inhibition tests than rSeV(hPIV-1HN). Its susceptibility to BCX 2798 in plaque reduction assays was reduced fivefold and did not differ from that of rSeV(hPIV-1HN) in mice. The N173S mutant failed to be efficiently eluted from erythrocytes and released from cells. It demonstrated reduced growth in cell culture and superior growth in mice. The results for gel electrophoresis analysis were consistent with the loss of the N-linked glycan at residue 173 in the mutant. Sequence and structural comparisons revealed that residue 173 on hPIV-1 HN is located close to the region of the second receptor-binding site identified in Newcastle disease virus HN. Our study suggests that the N-linked glycan at residue 173 masks a second receptor-binding site on hPIV-1 HN.
Preservation of FoxP3+Regulatory T Cells in the Peripheral Blood of Human Immunodeficiency Virus Type 1-Infected Elite Suppressors Correlates with Low CD4+T-Cell Activation
Authors: Chase Amanda J. , Yang Hung-Chih , Zhang Hao , Blankson Joel N. , Siliciano Robert F. 
Published: June 2008

ABSTRACTElite suppressors (ES) are untreated human immunodeficiency virus type 1 (HIV-1)-infected individuals who maintain normal CD4+T-cell counts and control viremia to levels that are below the limit of detection of current assays. The mechanisms involved in long-term control of viremia have not been fully elucidated. CD4+CD25+regulatory T cells (Tregs) downmodulate chronic inflammation by suppressing the activation and proliferation of effector lymphocytes. We found that while Tregs were functional in ES and patients on highly active antiretroviral therapy (HAART), ES maintained high levels of Tregs in peripheral blood mononuclear cells whereas patients on HAART had evidence of Treg depletion. We also demonstrated that Tregs can serve as reservoirs for HIV-1 in vivo. These data suggest that both direct infection by HIV-1 and tissue redistribution are possible explanations for declining FoxP3+Tregs in progressive HIV-1 infection. Furthermore, the maintenance of Tregs may be one mechanism associated with the nonprogressive nature of HIV-1 infection in ES.
Evidence for a Structural Relationship between BRCT Domains and the Helicase Domains of the Replication Initiators Encoded by thePolyomaviridaeandPapillomaviridaeFamilies of DNA Tumor Viruses
Authors: Kumar Anuradha , Joo Woo S. , Meinke Gretchen , Moine Stephanie , Naumova Elena N. , Bullock Peter A. 
Published: June 2008

ABSTRACTStudies of DNA tumor viruses have provided important insights into fundamental cellular processes and oncogenic transformation. They have revealed, for example, that upon expression of virally encoded proteins, cellular pathways involved in DNA repair and cell cycle control are disrupted. Herein, evidence is presented that BRCT-related regions are present in the helicase domains of the viral initiators encoded by thePolyomaviridaeandPapillomaviridaeviral families. Of interest, BRCT domains in cellular proteins recruit factors involved in diverse pathways, including DNA repair and the regulation of cell cycle progression. Therefore, the viral BRCT-related regions may compete with host BRCT domains for particular cellular ligands, a process that would help to explain the pleiotropic effects associated with infections with many DNA tumor viruses.
Human Papillomavirus Type 16 E7 Oncoprotein Associates with E2F6
Authors: McLaughlin-Drubin Margaret E. , Huh Kyung-Won , Münger Karl 
Published: June 2008

ABSTRACT
The papillomavirus life cycle is intimately coupled to the differentiation state of the infected epithelium. Since papillomaviruses lack most of the rate-limiting enzymes required for genome synthesis, they need to uncouple keratinocyte differentiation from cell cycle arrest and maintain or reestablish a replication-competent state within terminally differentiated keratinocytes. The human papillomavirus (HPV) E7 protein appears to be a major determinant for this activity and induces aberrant S-phase entry through the inactivation of the retinoblastoma tumor suppressor and related pocket proteins. In addition, E7 can abrogate p21 and p27. Together, this leads to the activation of E2F1 to E2F5, enhanced expression of E2F-responsive genes, and increased cdk2 activity. E2F6 is a pRB-independent, noncanonical member of the E2F transcription factor family that acts as a transcriptional repressor. E2F6 expression is activated in S phase through an E2F-dependent mechanism and thus may provide a negative-feedback mechanism that slows down S-phase progression and/or exit in response to the activation of the other E2F transcription factors. Here, we show that low- and high-risk HPV E7 proteins, as well as simian virus 40 T antigen and adenovirus E1A, can associate with and inactivate the transcriptional repression activity of E2F6, thereby subverting a critical cellular defense mechanism. This may result in the extended S-phase competence of HPV-infected cells. E2F6 is a component of polycomb group complexes, which bind to silenced chromatin and are critical for the maintenance of cell fate. We show that E7-expressing cells show decreased staining for E2F6/polycomb complexes and that this is at least in part dependent on the association with E2F6.
Protein Kinase A-Dependent Step(s) in Hepatitis C Virus Entry and Infectivity
Authors: Farquhar Michelle J. , Harris Helen J. , Diskar Mandy , Jones Sarah , Mee Christopher J. , Nielsen Søren U. , Brimacombe Claire L. , Molina Sonia , Toms Geoffrey L. , Maurel Patrick , Howl John , Herberg Friedrich W. , van IJzendoorn Sven C. D. , Balfe Peter , McKeating Jane A. 
Published: June 2008

ABSTRACTViruses exploit signaling pathways to their advantage during multiple stages of their life cycle. We demonstrate a role for protein kinase A (PKA) in the hepatitis C virus (HCV) life cycle. The inhibition of PKA with H89, cyclic AMP (cAMP) antagonists, or the protein kinase inhibitor peptide reduced HCV entry into Huh-7.5 hepatoma cells. Bioluminescence resonance energy transfer methodology allowed us to investigate the PKA isoform specificity of the cAMP antagonists in Huh-7.5 cells, suggesting a role for PKA type II in HCV internalization. Since viral entry is dependent on the host cell expression of CD81, scavenger receptor BI, and claudin-1 (CLDN1), we studied the role of PKA in regulating viral receptor localization by confocal imaging and fluorescence resonance energy transfer (FRET) analysis. Inhibiting PKA activity in Huh-7.5 cells induced a reorganization of CLDN1 from the plasma membrane to an intracellular vesicular location(s) and disrupted FRET between CLDN1 and CD81, demonstrating the importance of CLDN1 expression at the plasma membrane for viral receptor activity. Inhibiting PKA activity in Huh-7.5 cells reduced the infectivity of extracellular virus without modulating the level of cell-free HCV RNA, suggesting that particle secretion was not affected but that specific infectivity was reduced. Viral particles released from H89-treated cells displayed the same range of buoyant densities as did those from control cells, suggesting that viral protein association with lipoproteins is not regulated by PKA. HCV infection of Huh-7.5 cells increased cAMP levels and phosphorylated PKA substrates, supporting a model where infection activates PKA in a cAMP-dependent manner to promote virus release and transmission.
Influenza Virus Protecting RNA: an Effective Prophylactic and Therapeutic Antiviral
Authors: Dimmock Nigel J. , Rainsford Edward W. , Scott Paul D. , Marriott Anthony C. 
Published: June 2008

ABSTRACT
Another influenza pandemic is inevitable, and new measures to combat this and seasonal influenza are urgently needed. Here we describe a new concept in antivirals based on a defined, naturally occurring defective influenza virus RNA that has the potential to protect against any influenza A virus in any animal host. This “protecting RNA” (244 RNA) is incorporated into virions which, although noninfectious, deliver the RNA to those cells of the respiratory tract that are naturally targeted by infectious influenza virus. A 120-ng intranasal dose of this 244 protecting virus completely protected mice against a simultaneous challenge of 10 50% lethal doses with influenza A/WSN (H1N1) virus. The 244 virus also protected mice against strong challenge doses of all other subtypes tested (i.e., H2N2, H3N2, and H3N8). This prophylactic activity was maintained in the animal for at least 1 week prior to challenge. The 244 virus was 10- to 100-fold more active than previously characterized defective influenza A viruses, and the protecting activity was confirmed to reside in the 244 RNA molecule by recovering a protecting virus entirely from cloned cDNA. There was a clear therapeutic benefit when the 244 virus was administered 24 to 48 h after a lethal challenge, an effect which has not been previously observed with any defective virus. Protecting virus reduced, but did not abolish, replication of challenge virus in mouse lungs during both prophylactic and therapeutic treatments. Protecting virus is a novel antiviral, having the potential to combat human influenza virus infections, particularly when the infecting strain is not known or is resistant to antiviral drugs.
STAT2 Is a Primary Target for Measles Virus V Protein-Mediated Alpha/Beta Interferon Signaling Inhibition
Authors: Ramachandran Aparna , Parisien Jean-Patrick , Horvath Curt M. 
Published: June 2008

ABSTRACT

Measles virus, a member of the
Morbillivirus
family, infects millions of people each year despite the availability of effective vaccines. The V protein of measles virus is an important virulence factor that can interfere with host innate immunity by inactivating alpha/beta interferon (IFN-α/β) and IFN-γ signaling through protein interactions with signal transducer and activator of transcription proteins STAT1 and STAT2. Here we demonstrate that although STAT1 interference results from protein interactions within a V protein N-terminal region encompassed by amino acids 110 to 130, detection of STAT1 interaction and IFN-γ signaling inhibition requires the presence of cellular STAT2. Cell-specific variability in STAT1 interference was observed to correlate with V protein expression level. A more direct target for measles virus V protein-mediated IFN-α/β evasion is STAT2. Results indicate that the widely conserved C-terminal zinc finger domain of measles virus V protein is both necessary and sufficient to bind STAT2 and disrupt IFN-α/β signal transduction. Mutagenesis and molecular modeling define a contact surface for STAT2 association that includes aspartic acid residue 248 as critical for STAT2 interference and IFN antiviral immune suppression. These findings clearly define the molecular determinants for measles virus IFN evasion and validate specific targets as candidates for therapeutic intervention.
Epstein-Barr Virus Latent Membrane Protein 2A Preferentially Signals through the Src Family Kinase Lyn
Authors: Rovedo Mark , Longnecker Richard 
Published: June 2008

ABSTRACTLatent membrane protein 2A (LMP2A) is a viral protein expressed during Epstein-Barr virus (EBV) latency in EBV-infected B cells both in cell culture and in vivo. LMP2A has important roles in modulating B-cell receptor signal transduction and provides survival and developmental signals to B cells in vivo. Although Lyn has been shown to be important in mediating LMP2A signaling, it is still unclear if Lyn is used preferentially or if LMP2A associates promiscuously with other Src family kinase (SFK) members. To investigate the role of various SFKs in LMP2A signaling, we crossed LMP2A transgenic mice (TgE) with Lyn−/−, Fyn−/−, or Blk−/−mice. TgE Lyn−/−mice had a larger immunoglobulin M (IgM)-positive B-cell population than TgE mice, suggesting that the absence of Lyn prevents LMP2A from delivering survival and developmental signals to the B cells. Both TgE Fyn−/−and TgE Blk−/−mice have an IgM-negative population of splenic B cells, similar to the TgE mice. LMP2A was also transiently transfected into the human EBV-negative B-cell line BJAB to determine which SFK members associate with LMP2A. Lyn was detected in LMP2A immunoprecipitates, whereas Fyn was not. Both Lyn and Fyn were able to bind to an LMP2A mutant which contained a sequence shown previously to bind tightly to the SH2 domain of multiple SFK members. From these results, we conclude that LMP2A preferentially associates with and signals through Lyn compared to its association with other SFKs. This preferential association is due in part to the SH2 domain of Lyn associating with LMP2A.