bluuebunny/crossref_metadata_2025_split huggingface dataset:
Role of AKT Kinase in Measles Virus Replication
Authors: Carsillo Mary , Kim Dhohyung , Niewiesk Stefan 
Published: November 2009

ABSTRACT

Many RNA and DNA viruses activate serine-threonine kinase AKT to increase virus replication. In contrast, measles virus (MV) infection leads to downregulation of AKT. This is thought to be beneficial for the virus because it correlates with immune suppression. To determine whether this is a sacrifice for the virus, we used a recombinant virus and transfected cells expressing constitutively active AKT and evaluated its effect on virus replication.
In vitro
, overexpression of AKT did not influence virus replication but did affect (cell-type dependent) virus release.
In vivo
, the recombinant virus did not abrogate inhibition of proliferation of spleen cells from MV-infected cotton rats.
Immunization with Live Attenuated Influenza Viruses That Express Altered NS1 Proteins Results in Potent and Protective Memory CD8
+
T-Cell Responses
Authors: Mueller Scott N. , Langley William A. , Carnero Elena , García-Sastre Adolfo , Ahmed Rafi 
Published: November 2009

ABSTRACT

The generation of vaccines that induce long-lived protective immunity against influenza virus infections remains a challenging goal. Ideally, vaccines should elicit effective humoral and cellular immunity to protect an individual from infection or disease. Cross-reactive T- and B-cell responses that are elicited by live virus infections may provide such broad protection. Optimal induction of T-cell responses involves the action of type I interferons (IFN-I). Influenza virus expressed nonstructural protein 1 (NS1) functions as an inhibitor of IFN-I and promotes viral growth. We wanted to examine the priming of CD8
+
T-cell responses to influenza virus in the absence of this inhibition of IFN-I production. We generated recombinant mouse-adapted influenza A/PR/8/34 viruses with NS1 truncations and/or deletions that also express the gp33-41 epitope from lymphocytic choriomeningitis virus. Intranasal infection of mice with the attenuated viruses primed long-lived T- and B-cell responses despite significantly reduced viral replication in the lungs compared to wild-type virus. Antigen-specific CD8
+
T cells expanded upon rechallenge and generated increased protective memory T-cell populations after boosting. These results show that live attenuated influenza viruses expressing truncated NS1 proteins can prime protective immunity and may have implications for the design of novel modified live influenza virus vaccines.
Hemagglutinin-Pseudotyped Green Fluorescent Protein-Expressing Influenza Viruses for the Detection of Influenza Virus Neutralizing Antibodies
Authors: Martínez-Sobrido Luis , Cadagan Richard , Steel John , Basler Christopher F. , Palese Peter , Moran Thomas M. , García-Sastre Adolfo 
Published: November 2009

ABSTRACT
Influenza virus is a highly contagious virus that causes yearly epidemics and occasional pandemics of great consequence. Influenza virus neutralizing antibodies (NAbs) are promising prophylactic and therapeutic reagents. Detection of NAbs in serum samples is critical to evaluate the prevalence and spread of new virus strains. Here we describe the development of a simple, sensitive, specific, and safe screening assay for the rapid detection of NAbs against highly pathogenic influenza viruses under biosafety level 2 (BSL-2) conditions. This assay is based on the use of influenza viruses in which the hemagglutinin (HA) gene is replaced by a gene expressing green fluorescent protein (GFP). These GFP-expressing influenza viruses replicate to high titers in HA-expressing cell lines, but in non-HA-expressing cells, their replication is restricted to a single cycle.
Nonpathogenesis of Simian Immunodeficiency Virus Infection Is Associated with Reduced Inflammation and Recruitment of Plasmacytoid Dendritic Cells to Lymph Nodes, Not to Lack of an Interferon Type I Response, during the Acute Phase
Authors: Campillo-Gimenez Laure , Laforge Mireille , Fay Michèle , Brussel Audrey , Cumont Marie-Christine , Monceaux Valérie , Diop Ousmane , Lévy Yves , Hurtrel Bruno , Zaunders John , Corbeil Jacques , Elbim Carole , Estaquier Jérôme 
Published: November 2009

ABSTRACTDivergent Toll-like receptor 7 (TLR7) and TLR9 signaling has been proposed to distinguish pathogenic from nonpathogenic simian immunodeficiency virus infection in primate models. We demonstrate here that increased expression of type I interferon in pathogenic rhesus macaques compared to nonpathogenic African green monkeys was associated with the recruitment of plasmacytoid dendritic cells in the lymph nodes and the presence of an inflammatory environment early after infection, instead of a difference in the TLR7/9 response.
A Single Amino Acid Difference in Human APOBEC3H Variants Determines HIV-1 Vif Sensitivity
Authors: Zhen Anjie , Wang Tao , Zhao Ke , Xiong Yong , Yu Xiao-Fang 
Published: November 2009

ABSTRACT
Several variants of APOBEC3H (A3H) have been identified in different human populations. Certain variants of this protein are particularly potent inhibitors of retrotransposons and retroviruses, including HIV-1. However, it is not clear whether HIV-1 Vif can recognize and suppress the antiviral activity of A3H variants, as it does with other APOBEC3 proteins. We now report that A3H_Haplotype II (HapII), a potent inhibitor of HIV-1 in the absence of Vif, can indeed be degraded by HIV-1 Vif. Vif-induced degradation of A3H_HapII was blocked by the proteasome inhibitor MG132 and a Cullin5 (Cul5) dominant negative mutant. In addition, Vif mutants that were incapable of assembly with the host E3 ligase complex factors Cul5, ElonginB, and ElonginC were also defective for A3H_HapII suppression. Although we found that Vif hijacks the same E3 ligase to degrade A3H_HapII as it does to inactivate APOBEC3G (A3G) and APOBEC3F (A3F), more Vif motifs were involved in A3H_HapII inactivation than in either A3G or A3F suppression. In contrast to A3H_HapII, A3H_Haplotype I (HapI), which differs in only three amino acids from A3H_HapII, was resistant to HIV-1 Vif-mediated degradation. We also found that residue 121 was critical for determining A3H sensitivity and binding to HIV-1 Vif.
Viral Oncolysis That Targets Raf-1 Signaling Control of Nuclear Transport
Authors: Riolobos Laura , Valle Noelia , Hernando Eva , Maroto Beatriz , Kann Michael , Almendral José M. 
Published: November 2009

ABSTRACT

The central role of Raf protein kinase isoforms in human cancer demands specific anti-Raf therapeutic inhibitors. Parvoviruses are currently used in experimental cancer therapy due to their natural oncotropism and lytic life cycle. In searching for mechanisms underlying parvovirus oncolysis, we found that trimers of the major structural protein (VP) of the parvovirus minute virus of mice (MVM), which have to be imported into the nucleus for capsid assembly, undergo phosphorylation by the Raf-1 kinase. Purified Raf-1 phosphorylated the capsid subunits
in vitro
to the two-dimensional pattern found in natural MVM infections. VP trimers isolated from mammalian cells translocated into the nucleus of digitonin-permeabilized human cells. In contrast, VP trimers isolated from insect cells, which are devoid of Raf-1, were neither phosphorylated nor imported into the mammalian nucleus. However, the coexpression of a constitutively active Raf-1 kinase in insect cells restored VP trimer phosphorylation and nuclear transport competence. In MVM-infected normal and transformed cells, Raf-1 inhibition resulted in cytoplasmic retention of capsid proteins, preventing their nuclear assembly and progeny virus maturation. The level of Raf-1 activity in cancer cells was consistent with the extent of VP specific phosphorylation and with the permissiveness to MVM infection. Thus, Raf-1 control of nuclear translocation of MVM capsid assembly intermediates provides a novel target for viral oncolysis. MVM may reinforce specific therapies against frequent human cancers with deregulated Raf signaling.
Dynamin- and Clathrin-Dependent Endocytosis in African Swine Fever Virus Entry
Authors: Hernaez Bruno , Alonso Covadonga 
Published: November 2009

ABSTRACT

African swine fever virus (ASFV) is a large DNA virus that enters host cells after receptor-mediated endocytosis and depends on acidic cellular compartments for productive infection. The exact cellular mechanism, however, is largely unknown. In order to dissect ASFV entry, we have analyzed the major endocytic routes using specific inhibitors and dominant negative mutants and analyzed the consequences for ASFV entry into host cells. Our results indicate that ASFV entry into host cells takes place by clathrin-mediated endocytosis which requires dynamin GTPase activity. Also, the clathrin-coated pit component Eps15 was identified as a relevant cellular factor during infection. The presence of cholesterol in cellular membranes, but not lipid rafts or caveolae, was found to be essential for a productive ASFV infection. In contrast, inhibitors of the Na
+
/H
+
ion channels and actin polymerization inhibition did not significantly modify ASFV infection, suggesting that macropinocytosis does not represent the main entry route for ASFV. These results suggest a dynamin-dependent and clathrin-mediated endocytic pathway of ASFV entry for the cell types and viral strains analyzed.
Viable Polioviruses That Encode 2A Proteins with Fluorescent Protein Tags
Authors: Teterina Natalya L. , Levenson Eric A. , Ehrenfeld Ellie 
Published: November 2009

ABSTRACT

The 2A proteins of the
Picornaviridae
enterovirus genus are small cysteine proteinases that catalyze essential cleavages in the viral polyprotein in
cis
and in several cellular proteins in
trans
. In addition, 2A has been implicated in the process of viral RNA replication, independent of its protease functions. We have generated viable polioviruses that encode 2A proteins containing fluorescent protein tag insertions at either of two sites in the 2A protein structure. Viruses containing an insertion of
Discosoma
sp. red fluorescent protein (DsRed) after residue 144 of 2A, near the C terminus, produced plaques only slightly smaller than wild-type (wt) virus. The polyprotein harboring the 2A-DsRed fusion protein was efficiently and accurately cleaved; fluorescent 2A proteinase retained protease activity in
trans
and supported translation and replication of viral RNA, both
in vitro
and in infected cells. Intracellular membrane reorganization to support viral RNA synthesis was indistinguishable from that induced by wt virus. Infected cells exhibited strong red fluorescence from expression of the 2A-DsRed fusion protein, and the progeny virus was stable for three to four passages, after which deletions within the DsRed coding sequence began to accumulate. Confocal microscopic imaging and analysis revealed a portion of 2A-DsRed in punctate foci concentrated in the perinuclear region that colocalized with replication protein 2C. The majority of 2A, however, was associated with an extensive structural matrix throughout the cytoplasm and was not released from infected cells permeabilized with digitonin.
The Alpha/Beta Interferon Receptor Provides Protection against Influenza Virus Replication but Is Dispensable for Inflammatory Response Signaling
Authors: Goodman Alan G. , Zeng Hui , Proll Sean C. , Peng Xinxia , Cillóniz Cristian , Carter Victoria S. , Korth Marcus J. , Tumpey Terrence M. , Katze Michael G. 
Published: November 2009

ABSTRACT

The innate immune response provides the first line of defense against foreign pathogens by responding to molecules that are a signature of a pathogenic infection. Certain RNA viruses, such as influenza virus, produce double-stranded RNA as an intermediate during the replication life cycle, which activates pathogen recognition receptors capable of inducing interferon production. By engaging interferon receptors, interferon activates the JAK-STAT pathway and results in the positive feedback of interferon production, amplifying the response to viral infection. To examine how deficiencies in interferon signaling affect the cellular response to infection, we performed influenza virus infections of mouse embryonic fibroblasts lacking the alpha/beta interferon receptor, the gamma interferon receptor, or both. In the absence of the alpha/beta interferon receptor, we observed increased viral replication but decreased activation of PKR, Stat1, and NF-κB; the presence or absence of the gamma interferon receptor did not exhibit discernible differences in these readouts. Analysis of gene expression profiles showed that while cells lacking the alpha/beta interferon receptor exhibited decreased levels of transcription of antiviral genes, genes related to inflammatory and apoptotic responses were transcribed to levels similar to those of cells containing the receptor. These results indicate that while the alpha/beta interferon receptor is needed to curb viral replication, it is dispensable for the induction of certain inflammatory and apoptotic genes. We have identified potential pathways, via interferon regulatory factor 3 (IRF3) activation or
Hoxa13
,
Polr2a
,
Nr4a1
, or
Ing1
induction, that contribute to this redundancy. This study illustrates another way in which the host has evolved to establish several overlapping mechanisms to respond to viral infections.
Pandemic H1N1 2009 Influenza A Virus Induces Weak Cytokine Responses in Human Macrophages and Dendritic Cells and Is Highly Sensitive to the Antiviral Actions of Interferons
Authors: Österlund Pamela , Pirhonen Jaana , Ikonen Niina , Rönkkö Esa , Strengell Mari , Mäkelä Sanna M. , Broman Mia , Hamming Ole J. , Hartmann Rune , Ziegler Thedi , Julkunen Ilkka 
Published: November 2009

ABSTRACTIn less than 3 months after the first cases of swine origin 2009 influenza A (H1N1) virus infections were reported from Mexico, WHO declared a pandemic. The pandemic virus is antigenically distinct from seasonal influenza viruses, and the majority of human population lacks immunity against this virus. We have studied the activation of innate immune responses in pandemic virus-infected human monocyte-derived dendritic cells (DC) and macrophages. Pandemic A/Finland/553/2009 virus, representing a typical North American/European lineage virus, replicated very well in these cells. The pandemic virus, as well as the seasonal A/Brisbane/59/07 (H1N1) and A/New Caledonia/20/99 (H1N1) viruses, induced type I (alpha/beta interferon [IFN-α/β]) and type III (IFN-λ1 to -λ3) IFN, CXCL10, and tumor necrosis factor alpha (TNF-α) gene expression weakly in DCs. Mouse-adapted A/WSN/33 (H1N1) and human A/Udorn/72 (H3N2) viruses, instead, induced efficiently the expression of antiviral and proinflammatory genes. Both IFN-α and IFN-β inhibited the replication of the pandemic (H1N1) virus. The potential of IFN-λ3 to inhibit viral replication was lower than that of type I IFNs. However, the pandemic virus was more sensitive to the antiviral IFN-λ3 than the seasonal A/Brisbane/59/07 (H1N1) virus. The present study demonstrates that the novel pandemic (H1N1) influenza A virus can readily replicate in human primary DCs and macrophages and efficiently avoid the activation of innate antiviral responses. It is, however, highly sensitive to the antiviral actions of IFNs, which may provide us an additional means to treat severe cases of infection especially if significant drug resistance emerges.
Herpes Simplex Virus VP16, but Not ICP0, Is Required To Reduce Histone Occupancy and Enhance Histone Acetylation on Viral Genomes in U2OS Osteosarcoma Cells
Authors: Hancock Meaghan H. , Cliffe Anna R. , Knipe David M. , Smiley James R. 
Published: November 2009

ABSTRACT
The herpes simplex virus (HSV) genome rapidly becomes associated with histones after injection into the host cell nucleus. The viral proteins ICP0 and VP16 are required for efficient viral gene expression and have been implicated in reducing the levels of underacetylated histones on the viral genome, raising the possibility that high levels of underacetylated histones inhibit viral gene expression. The U2OS osteosarcoma cell line is permissive for replication of ICP0 and VP16 mutants and appears to lack an innate antiviral repression mechanism present in other cell types. We therefore used chromatin immunoprecipitation to determine whether U2OS cells are competent to load histones onto HSV DNA and, if so, whether ICP0 and/or VP16 are required to reduce histone occupancy and enhance acetylation in this cell type. High levels of underacetylated histone H3 accumulated at several locations on the viral genome in the absence of VP16 activation function; in contrast, an ICP0 mutant displayed markedly reduced histone levels and enhanced acetylation, similar to wild-type HSV. These results demonstrate that U2OS cells are competent to load underacetylated histones onto HSV DNA and uncover an unexpected role for VP16 in modulating chromatin structure at viral early and late loci. One interpretation of these findings is that ICP0 and VP16 affect viral chromatin structure through separate pathways, and the pathway targeted by ICP0 is defective in U2OS cells. We also show that HSV infection results in decreased histone levels on some actively transcribed genes within the cellular genome, demonstrating that viral infection alters cellular chromatin structure.
Mutations in Sensor 1 and Walker B in the Bovine Papillomavirus E1 Initiator Protein Mimic the Nucleotide-Bound State
Authors: Liu Xiaofei , Stenlund Arne 
Published: November 2009

ABSTRACT
Viral replication initiator proteins are multifunctional proteins that utilize ATP binding and hydrolysis by their AAA+ modules for multiple functions in the replication of their viral genomes. These proteins are therefore of particular interest for understanding how AAA+ proteins carry out multiple ATP driven functions. We have performed a comprehensive mutational analysis of the residues involved in ATP binding and hydrolysis in the papillomavirus E1 initiator protein based on the recent structural data. Ten of the eleven residues that were targeted were defective for ATP hydrolysis, and seven of these were also defective for ATP binding. The three mutants that could still bind nucleotide represent the Walker B motif (D478 and D479) and Sensor 1 (N523), three residues that are in close proximity to each other and generally are considered to be involved in ATP hydrolysis. Surprisingly, however, two of these mutants, D478A and N523A, mimicked the nucleotide bound state and were capable of binding DNA in the absence of nucleotide. However, these mutants could not form the E1 double trimer in the absence of nucleotide, demonstrating that there are two qualitatively different consequences of ATP binding by E1, one that can be mimicked by D478A and N523A and one which cannot.
Protease Cleavage Sites in HIV-1 gp120 Recognized by Antigen Processing Enzymes Are Conserved and Located at Receptor Binding Sites
Authors: Yu Bin , Fonseca Dora P. A. J. , O'Rourke Sara M. , Berman Phillip W. 
Published: November 2009

ABSTRACTThe identification of vaccine immunogens able to elicit broadly neutralizing antibodies (bNAbs) is a major goal in HIV vaccine research. Although it has been possible to produce recombinant envelope glycoproteins able to adsorb bNAbs from HIV-positive sera, immunization with these proteins has failed to elicit antibody responses effective against clinical isolates of HIV-1. Thus, the epitopes recognized by bNAbs are present on recombinant proteins, but they are not immunogenic. These results led us to consider the possibility that changes in the pattern of antigen processing might alter the immune response to the envelope glycoprotein to better elicit protective immunity. In these studies, we have defined protease cleavage sites on HIV gp120 recognized by three major human proteases (cathepsins L, S, and D) important for antigen processing and presentation. Remarkably, six of the eight sites identified in gp120 were highly conserved and clustered in regions of the molecule associated with receptor binding and/or the binding of neutralizing antibodies. These results suggested that HIV may have evolved to take advantage of major histocompatibility complex (MHC) class II antigen processing enzymes in order to evade or direct the antiviral immune response.
The Minor Envelope Glycoproteins GP2a and GP4 of Porcine Reproductive and Respiratory Syndrome Virus Interact with the Receptor CD163
Authors: Das Phani B. , Dinh Phat X. , Ansari Israrul H. , de Lima Marcelo , Osorio Fernando A. , Pattnaik Asit K. 
Published: November 2009

ABSTRACT
Porcine reproductive and respiratory syndrome virus (PRRSV) contains the major glycoprotein, GP5, as well as three other minor glycoproteins, namely, GP2a, GP3, and GP4, on the virion envelope, all of which are required for generation of infectious virions. To study their interactions with each other and with the cellular receptor for PRRSV, we have cloned each of the viral glycoproteins and CD163 receptor in expression vectors and examined their expression and interaction with each other in transfected cells by coimmunoprecipitation (co-IP) assay using monospecific antibodies. Our results show that a strong interaction exists between the GP4 and GP5 proteins, although weak interactions among the other minor envelope glycoproteins and GP5 have been detected. Both GP2a and GP4 proteins were found to interact with all the other GPs, resulting in the formation of multiprotein complex. Our results further show that the GP2a and GP4 proteins also specifically interact with the CD163 molecule. The carboxy-terminal 223 residues of the CD163 molecule are not required for interactions with either the GP2a or the GP4 protein, although these residues are required for conferring susceptibility to PRRSV infection in BHK-21 cells. Overall, we conclude that the GP4 protein is critical for mediating interglycoprotein interactions and, along with GP2a, serves as the viral attachment protein that is responsible for mediating interactions with CD163 for virus entry into susceptible host cell.
Molecular Determinants of Adaptation of Highly Pathogenic Avian Influenza H7N7 Viruses to Efficient Replication in the Human Host
Authors: de Wit Emmie , Munster Vincent J. , van Riel Debby , Beyer Walter E. P. , Rimmelzwaan Guus F. , Kuiken Thijs , Osterhaus Albert D. M. E. , Fouchier Ron A. M. 
Published: November 2009

ABSTRACT

Two viruses isolated during the highly pathogenic avian influenza (HPAI) H7N7 virus outbreak in The Netherlands in 2003, one isolated from a person with conjunctivitis and one from a person who died as the result of infection, were used for an
in vitro
study of influenza A virus pathogenicity factors. The two HPAI H7N7 viruses differed in 15 amino acid positions in five gene segments. Assays were designed to investigate the role of each of these substitutions in attachment and entry, transcription and genome replication, and virus production and release as determined by hemagglutinin (HA), polymerase proteins, nonstructural protein 1 (NS1), and neuraminidase (NA). These
in vitro
studies confirmed the roles of the E627K substitution in basic polymerase 2 (PB2) and the A143T substitution in HA in pathogenicity observed in a mouse model previously. However, the
in vitro
studies identified a contribution of acidic polymerase (PA) and NA to the efficient replication in human cells of the fatal case virus, despite the fact that these are rarely marked as determinants of pathogenicity in
in vivo
studies. With the exception of PB2 E627K, all substitutions contributing to enhanced replication of the fatal case virus
in vitro
were present in poultry viruses prior to transmission to the human fatal case, indicating that viruses with enhanced replication efficiency in the mammalian host can be generated in poultry. Thus, detailed
in vitro
analyses of mutations facilitating replication of avian influenza viruses in mammalian cells are important to assess the zoonotic risk posed by these viruses and, in addition, highlight the value of
in vitro
studies to complement animal models.