bluuebunny/crossref_metadata_2025 huggingface dataset:
Lethality to Ferrets of H5N1 Influenza Viruses Isolated from Humans and Poultry in 2004
Authors: Govorkova Elena A. , Rehg Jerold E. , Krauss Scott , Yen Hui-Ling , Guan Yi , Peiris Malik , Nguyen Tien D. , Hanh Thi H. , Puthavathana Pilipan , Long Hoang T. , Buranathai Chantanee , Lim Wilina , Webster Robert G. , Hoffmann Erich
Published: January 2005

The 2004 outbreaks of H5N1 influenza viruses in Vietnam and Thailand were highly lethal to humans and to poultry; therefore, newly emerging avian influenza A viruses pose a continued threat, not only to avian species but also to humans. We studied the pathogenicity of four human and nine avian H5N1/04 influenza viruses in ferrets (an excellent model for influenza studies). All four human isolates were fatal to intranasally inoculated ferrets. The human isolate A/Vietnam/1203/04 (H5N1) was the most pathogenic isolate; the severity of disease was associated with a broad tissue tropism and high virus titers in multiple organs, including the brain. High fever, weight loss, anorexia, extreme lethargy, and diarrhea were observed. Two avian H5N1/04 isolates were as pathogenic as the human viruses, causing lethal systemic infections in ferrets. Seven of nine H5N1/04 viruses isolated from avian species caused mild infections, with virus replication restricted to the upper respiratory tract. All chicken isolates were nonlethal to ferrets. A sequence analysis revealed polybasic amino acids in the hemagglutinin connecting peptides of all H5N1/04 viruses, indicating that multiple molecular differences in other genes are important for a high level of virulence. Interestingly, the human A/Vietnam/1203/04 isolate had a lysine substitution at position 627 of PB2 and had one to eight amino acid changes in all gene products except that of the M1 gene, unlike the A/chicken/Vietnam/C58/04 and A/quail/Vietnam/36/04 viruses. Our results indicate that viruses that are lethal to mammals are circulating among birds in Asia and suggest that pathogenicity in ferrets, and perhaps humans, reflects a complex combination of different residues rather than a single amino acid difference.
Kinetics of Human Immunodeficiency Virus Type 1 Decay following Entry into Resting CD4
+
T Cells
Authors: Zhou Yan , Zhang Haili , Siliciano Janet D. , Siliciano Robert F.
Published: January 2005

In untreated human immunodeficiency virus type 1 (HIV-1) infection, most viral genomes in resting CD4
+
T cells are not integrated into host chromosomes. This unintegrated virus provides an inducible latent reservoir because cellular activation permits integration, virus gene expression, and virus production. It remains controversial whether HIV-1 is stable in this preintegration state. Here, we monitored the fate of HIV-1 in resting CD4
+
cells by using a green fluorescent protein (GFP) reporter virus carrying an X4 envelope. After virus entry into resting CD4
+
T cells, both rescuable virus gene expression, visualized with GFP, and rescuable virion production, assessed by p24 release, decayed with a half-life of 2 days. In these cells, reverse transcription goes to completion over 2 to 3 days, and 50% of the viruses that have entered undergo functional decay before reverse transcription is complete. We distinguished two distinct but closely related factors contributing to loss of rescuable virus. First, some host cells undergo virus-induced apoptosis upon viral entry, thereby reducing the amount of rescuable virus. Second, decay processes directly affecting the virus both before and after the completion of reverse transcription contribute to the loss of rescuable virus. The functional half-life of full-length, integration-competent reverse transcripts is only 1 day. We propose that rapid intracellular decay processes compete with early steps in viral replication in infected CD4
+
T cells. Decay processes dominate in resting CD4
+
T cells as a result of the slow kinetics of reverse transcription and blocks at subsequent steps. Therefore, the reservoir of unintegrated HIV-1 in recently infected resting CD4
+
T cells is highly labile.
RNA Recombination of Hepatitis Delta Virus in Natural Mixed-Genotype Infection and Transfected Cultured Cells
Authors: Wang Tzu-Chi , Chao Mei
Published: January 2005

Most RNA viruses encode their own RNA polymerases for genome replication, and increasing numbers of them appear to be capable of undergoing RNA recombination. Here, we provide the first report of intergenotypic recombination in hepatitis delta virus (HDV), the only animal RNA virus that requires host RNA polymerase(s) for viral replication. In vivo, we analyzed RNA species derived from the serum of a patient with mixed genotype I and genotype IIb HDV infection by using multiple restriction fragment length polymorphism (RFLP) assays and sequence analysis of cloned reverse transcription (RT)-PCR products. Six HDV recombinants were isolated from 101 tested clones, and HDV recombination in this patient was further confirmed by RT-PCR with genotype-specific primer pairs. Analysis of the recombination junctions suggested that the HDV genome rearrangement occurred through faithful homologous recombination. We then used an RNA cotransfection cell culture system to investigate HDV RNA recombination in vitro. We found that HDV recombinants could indeed be detected in the transfected cells; some of these possessed recombination junctions identical to those identified in vivo. Furthermore, using a PCR-independent RNase protection assay, we were able to readily identify the recombined HDV RNA species in cultured cells. Taken together, our results demonstrate that HDV RNA recombination occurs in both natural HDV infections and cultured cells, thereby presenting a novel mechanism for HDV evolution.
Involvement of the Interferon-Regulated Antiviral Proteins PKR and RNase L in Reovirus-Induced Shutoff of Cellular Translation
Authors: Smith Jennifer A. , Schmechel Stephen C. , Williams Bryan R. G. , Silverman Robert H. , Schiff Leslie A.
Published: January 2005

Cellular translation is inhibited following infection with most strains of reovirus, but the mechanisms responsible for this phenomenon remain to be elucidated. The extent of host shutoff varies in a strain-dependent manner; infection with the majority of strains leads to strong host shutoff, while infection with strain Dearing results in minimal inhibition of cellular translation. A genetic study with reassortant viruses and subsequent biochemical analyses led to the hypothesis that the interferon-induced, double-stranded RNA-activated protein kinase, PKR, is responsible for reovirus-induced host shutoff. To directly determine whether PKR is responsible for reovirus-induced host shutoff, we used a panel of reovirus strains and mouse embryo fibroblasts derived from knockout mice. This approach revealed that PKR contributes to but is not wholly responsible for reovirus-induced host shutoff. Studies with cells lacking RNase L, the endoribonuclease component of the interferon-regulated 2′,5′-oligoadenylate synthetase-RNase L system, demonstrated that RNase L also down-regulates cellular protein synthesis in reovirus-infected cells. In many viral systems, PKR and RNase L have well-characterized antiviral functions. An analysis of reovirus replication in cells lacking these molecules indicated that, while they contributed to host shutoff, neither PKR nor RNase L exerted an antiviral effect on reovirus growth. In fact, some strains of reovirus replicated more efficiently in the presence of PKR and RNase L than in their absence. Data presented in this report illustrate that the inhibition of cellular translation following reovirus infection is complex and involves multiple interferon-regulated gene products. In addition, our results suggest that reovirus has evolved effective mechanisms to avoid the actions of the interferon-stimulated antiviral pathways that include PKR and RNase L and may even benefit from their expression.
Expression Profiling of Human Hepatoma Cells Reveals Global Repression of Genes Involved in Cell Proliferation, Growth, and Apoptosis upon Infection with Parvovirus H-1
Authors: Li Jianhong , Werner Ekkehard , Hergenhahn Manfred , Poirey Rémy , Luo Zuyu , Rommelaere Jean , Jauniaux Jean-Claude
Published: January 2005

Autonomous parvoviruses are characterized by their stringent dependency on host cell S phase and their cytopathic effects on neoplastic cells. To better understand the interactions between the virus and its host cell, we used oligonucleotide arrays that carry more than 19,000 unique human gene sequences to profile the gene expression of the human hepatocellular carcinoma cell line QGY-7703 at two time points after parvovirus H-1 infection. At the 6-h time point, a single gene was differentially expressed with a >2.5-fold change. At 12 h, 105 distinct genes were differentially expressed in virus-infected cells compared to mock-treated cells, with 93% of these genes being down-regulated. These repressed genes clustered mainly into classes involved in transcriptional regulation, signal transduction, immune and stress response, and apoptosis, as exemplified by genes encoding the transcription factors Myc, Jun, Fos, Ids, and CEBPs. Quantitative real-time reverse transcription-PCR analysis on selected genes validated the array data and allowed the changes in cellular gene expression to be correlated with the accumulation of viral transcripts and NS1 protein. Western blot analysis of several cellular proteins supported the array results and substantiated the evidence given by these and other data to suggest that the H-1 virus kills QGY-7703 cells by a nonapoptotic process. The promoter regions of most of the differentially expressed genes analyzed fail to harbor any motif for sequence-specific binding of NS1, suggesting that direct binding of NS1 to cellular promoters may not participate in the modulation of cellular gene expression in H-1 virus-infected cells.
The Topology of Bulges in the Long Stem of the Flavivirus 3′ Stem-Loop Is a Major Determinant of RNA Replication Competence
Authors: Yu Li , Markoff Lewis
Published: January 2005

All flavivirus genomes contain a 3′terminal stem-loop secondary structure (3′SL) formed by the most downstream ∼100 nucleotides (nt) of the viral RNA. The 3′SL is required for virus replication and has been shown to bind both virus-coded and cellular proteins. Results of the present study using an infectious DNA for WN virus strain 956 initially demonstrated that the dengue virus serotype 2 (DEN2) 3′SL nucleotide sequence could not substitute for that of the WN 3′SL to support WN genome replication. To determine what WN virus-specific 3′SL nucleotide sequences were required for WN virus replication, WN virus 3′SL nucleotide sequences were selectively deleted and replaced by analogous segments of the DEN2 3′SL nucleotide sequence such that the overall 3′SL secondary structure was not disrupted. Top and bottom portions of the WN virus 3′SL were defined according to previous studies (J. L. Blackwell and M. A. Brinton, J. Virol.
71:
6433-6444, 1997; L. Zeng, L., B. Falgout, and L. Markoff, J. Virol.
72:
7510-7522, 1998). A bulge in the top portion of the long stem of the WN 3′SL was essential for replication of mutant WN RNAs, and replication-defective RNAs failed to produce negative strands in transfected cells. Introduction of a second bulge into the bottom portion of the long stem of the wild-type WN 3′SL markedly enhanced the replication competence of WN virus in mosquito cells but had no effect on replication in mammalian cells. This second bulge was identified as a host cell-specific enhancer of flavivirus replication. Results suggested that bulges and their topological location within the long stem of the 3′SL are primary determinants of replication competence for flavivirus genomes.
Disruption of the Shc/Grb2 Complex during Abelson Virus Transformation Affects Proliferation, but Not Apoptosis
Authors: Baughn Linda B. , Rosenberg Naomi
Published: January 2005

The v-Abl protein tyrosine kinase encoded by Abelson murine leukemia virus (Ab-MLV) induces pre-B-cell transformation. Signals emanating from the SH2 domain of the protein are required for transformation, and several proteins bind this region of v-Abl. One such protein is the adaptor molecule Shc, a protein that complexes with Grb2/Sos and facilitates Ras activation, an event associated with Ab-MLV transformation. To test the role this interaction plays in growth and survival of infected pre-B cells, dominant-negative (DN) Shc proteins were coexpressed with v-Abl and transformation was examined. Expression of DN Shc reduced Ab-MLV pre-B-cell transformation and decreased the ability of v-Abl to stimulate Ras activation and Erk phosphorylation in a Raf-dependent but Rac-independent fashion. Further analysis revealed that Shc is required for v-Abl-mediated Raf tyrosine 340 and 341 phosphorylation, an event associated with Erk phosphorylation. In contrast to effects on proliferation, survival of the cells and activation of Akt were not affected by expression of DN Shc. Together, these data reveal that v-Abl-Shc interactions are a critical part of the growth stimulatory signals delivered during transformation but that they do not affect antiapoptotic pathways. Furthermore, these data highlight a novel role for Shc in signaling from v-Abl to Raf.
Functional Analysis of the Inhibitor of Apoptosis (iap) Gene Carried by the Entomopoxvirus ofAmsacta moorei
Authors: Li Qianjun , Liston Peter , Moyer Richard W.
Published: January 2005

The entomopoxvirus fromAmsacta moorei(AmEPV) contains none of the commonly recognized vertebrate poxvirus apoptotic suppressor genes. However, AmEPV carries a single inhibitor of apoptosis (iap) gene (AMViap) not present in vertebrate poxviruses. The AMViapgene was active when coexpressed with theDrosophilaproapoptotic genehidin Ld652 cells and can rescue cells from apoptosis as shown by increased number of surviving cells and reduced levels of caspase-3-like activity. We also showed that expression of the AMViapgene rescued polyhedron production inAutographa californica Mnucleopolyhedrovirus (AcMNPV)Δp35-infected Sf9 cells during an otherwise abortive infection induced by apoptosis. Surprisingly, deletion of the AMViapgene from the AmEPV genome led to only a modest (10-fold) loss of virion production in infected Ld652 cells, indicating that the AMViapgene is nonessential for virus replication under these conditions. However, infection of Ld652 cells by AmEPV lacking a functionaliapgene led to a more rapid induction of cytotoxicity and increased levels of caspase-3-like activity. Similar results were observed and were more pronounced in infected Sf9 and S2 cells. The purified AMVIAP protein also inhibits the enzymatic activities of human caspase-9 and caspase-3 in vitro. Our results indicate that while the AMViapgene was active in controlling apoptosis through the intrinsic pathway, the virus likely encodes additional proteins that also regulate apoptosis.
Analysis of the Interaction of the Adenovirus L1 52/55-Kilodalton and IVa2 Proteins with the Packaging Sequence In Vivo and In Vitro
Authors: Perez-Romero Pilar , Tyler Ryan E. , Abend Johanna R. , Dus Monica , Imperiale Michael J.
Published: January 2005

We previously showed that the adenovirus IVa2 and L1 52/55-kDa proteins interact in infected cells and the IVa2 protein is part of two virus-specific complexes (x and y) formed in vitro with repeated elements of the packaging sequence called the A1-A2 repeats. Here we demonstrate that both the IVa2 and L1 52/55-kDa proteins bind in vivo to the packaging sequence and that each protein-DNA interaction is independent of the other. There is a strong and direct interaction of the IVa2 protein with DNA in vitro. This interaction is observed when probes containing the A1-A2 or A4-A5 repeats are used, but it is not found by using an A5-A6 probe. Furthermore, we show that complex x is likely a heterodimer of IVa2 and an unknown viral protein, while complex y is a monomer or multimer of IVa2. No in vitro interaction of purified L1 52/55-kDa protein with the packaging sequence was found, suggesting that the L1 52/55-kDa protein-DNA interaction may be mediated by an intermediate protein. Results support roles for both the L1 52/55-kDa and IVa2 proteins in DNA encapsidation.
Influence of the Human Parainfluenza Virus 3 Attachment Protein's Neuraminidase Activity on Its Capacity To Activate the Fusion Protein
Authors: Porotto Matteo , Murrell Matthew , Greengard Olga , Doctor Lynne , Moscona Anne
Published: January 2005

In order to examine functions of the hemagglutinin-neuraminidase (HN) protein that quantitatively influence fusion promotion, human parainfluenza virus 3 (HPIV3) variants with alterations in HN were studied. The variant HNs have mutations that affect either receptor binding avidity, neuraminidase activity, or fusion protein (F) activation. Neuraminidase activity was regulated by manipulation of temperature and pH. F activation was assessed by quantitating the irreversible binding of target erythrocytes (RBC) to HN/F-coexpressing cells in the presence of 4-GU-DANA (zanamivir) to release target cells bound only by HN-receptor interactions; the remaining, irreversibly bound target cells are retained via the fusion protein. In cells coexpressing wild-type (wt) or variant HNs with wt F, the fusion promotion capacity of HN was distinguished from target cell binding by measuring changes with time in the amounts of target RBC that were (i) reversibly bound by HN-receptor interaction (released only upon the addition of 4-GU-DANA), (ii) released by HN′s neuraminidase, and (iii) irreversibly bound by F-insertion or fusion (F triggered). For wt HN, lowering the pH (to approach the optimum for HPIV3 neuraminidase) decreased F triggering via release of HN from its receptor. An HN variant with increased receptor binding avidity had F-triggering efficiency like that of wt HN at pH 8.0, but this efficiency was not decreased by lowering the pH to 5.7, which suggested that the variant HN′s higher receptor binding activity counterbalanced the receptor dissociation promoted by increased neuraminidase activity. To dissect the specific contribution of neuraminidase to triggering, two variant HNs that are triggering-defective due to a mutation in the HN stalk were evaluated. One of these variants has, in addition, a mutation in the globular head that renders it neuraminidase dead, while the HN with the stalk mutation alone has 30% of wt neuraminidase. While the variant without neuraminidase activity triggered F effectively at 37°C irrespective of pH, the variant possessing effective neuraminidase activity completely failed to activate F at pH 5.7 and was capable of only minimal triggering activity even at pH 8.0. These results demonstrate that neuraminidase activity impacts the extent of HPIV3-mediated fusion by releasing HN from contact with receptor. Any particular HN′s competence to promote F-mediated fusion depends on the balance between its inherent F-triggering efficacy and its receptor-attachment regulatory functions (binding and receptor cleavage).
Modulation of Human Herpesvirus 8/Kaposi's Sarcoma-Associated Herpesvirus Replication and Transcription Activator Transactivation by Interferon Regulatory Factor 7
Authors: Wang Jinzhong , Zhang Jun , Zhang Luwen , Harrington William , West John T. , Wood Charles
Published: January 2005

Human herpesvirus 8 (HHV-8)/Kaposi's sarcoma-associated herpesvirus infection goes through lytic and latent phases that are regulated by viral gene products, but very little is known about the involvement of host proteins. The replication and transcription activator (RTA) is a viral protein sufficient to initiate lytic replication by activating downstream genes, including the viral early gene open reading frame 57 (ORF 57), which codes for a posttranscriptional activator. In this study, we demonstrate that cellular interferon regulatory factor 7 (IRF-7) negatively regulates this process by competing with RTA for binding to the RTA response element in the ORF 57 promoter to down-regulate RTA-induced gene expression. We also show that alpha interferon represses RTA-mediated transactivation and that repression involves IRF-7. Our study indicates that upon HHV-8 infection, the host responds by suppression of lytic gene expression through binding of IRF-7 to the lytic viral gene promoter. These findings suggest that HHV-8 has developed a novel mechanism to induce but then subvert the innate antiviral response, specifically the interferon-signaling pathway, to regulate RTA activity and ultimately the viral latent/lytic replicative cycle.
Differential Effects of Dengue Virus on Infected and Bystander Dendritic Cells
Authors: Palmer Dupeh R. , Sun Peifang , Celluzzi Christina , Bisbing John , Pang Somnang , Sun Wellington , Marovich Mary A. , Burgess Timothy
Published: January 2005

Dendritic cells (DCs) play a central role as major targets of dengue virus (DV) infections and initiators of antiviral immune responses. Previous observations showed that DCs are activated by infection, presumably acquiring the capacity to promote cell-mediated immunity. However, separate evaluations of the maturation profiles of infected and uninfected bystander cells show that infection impairs the ability of DCs to upregulate cell surface expression of costimulatory, maturation, and major histocompatibility complex molecules, resulting in reduced T-cell stimulatory capacity. Infected DCs failed to respond to tumor necrosis factor alpha as an additional maturation stimulus and were apoptotic. Interleukin 10 (IL-10) was detected in supernatants from cultures of DV-infected DCs and cocultures of DCs and T cells. Taken together, these results constitute an immune evasion strategy used by DV that directly impairs antigen-presenting cell function by maturation blockade and induction of apoptosis.
Characterization of Helper Virus-Independent Cytopathogenic Classical Swine Fever Virus Generated by an In Vivo RNA Recombination System
Authors: Gallei Andreas , Rümenapf Till , Thiel Heinz-Jürgen , Becher Paul
Published: January 2005

Molecular analyses revealed that most cytopathogenic (cp) pestivirus strains evolve from noncytopathogenic (noncp) viruses by nonhomologous RNA recombination. In contrast to bovine viral diarrhea virus (BVDV), cp classical swine fever virus (CSFV) field isolates were rarely detected and always represented helper virus-dependent subgenomes. To investigate RNA recombination in more detail, we recently established an in vivo system allowing the efficient generation of recombinant cp BVDV strains in cell culture after transfecting a synthetic subgenomic and nonreplicatable transcript into cells being infected with noncp BVDV (A. Gallei, A. Pankraz, H.-J. Thiel, and P. Becher, J. Virol.
78:
6271-6281, 2004). Using an analogous approach, the first helper virus-independent cp CSFV strain (CP G1) has now been generated by RNA recombination. Accordingly, this study demonstrates the applicability of RNA recombination for designing new viral RNA genomes. The genomic RNA of CP G1 has a calculated size of 18.139 kb, almost 6 kb larger than all previously described CSFV genomes. It contains cellular sequences encoding a polyubiquitin fragment directly upstream of the nonstructural protein NS3 coding gene together with a duplication of viral sequences. CP G1 induces a cytopathic effect on different tissue culture cell lines from pigs and cattle. Subsequent analyses addressed growth kinetics, expression of NS3, and genetic stability of CP G1.
Identification of Internal Sequences in the 3′ Leader Region of Human Respiratory Syncytial Virus That Enhance Transcription and Confer Replication Processivity
Authors: McGivern David R. , Collins Peter L. , Fearns Rachel
Published: January 2005

Previous studies of respiratory syncytial virus have shown that the 44-nucleotide (nt) leader (Le) region is sufficient to initiate RNA replication, producing antigenome RNA, and that the Le and adjoining gene start (GS) signal of the first gene are sufficient to initiate transcription, producing mRNA. A
cis
-acting element necessary for both transcription and replication was mapped within the first 11 nt at the 3′ end of Le. In the present study the remainder of the Le region was mapped to identify sequences important for transcription and replication. A series of minigenomes with mutant Le sequences was generated, and their ability to direct transcription and replication was determined by Northern blot analysis, which examined full-length antigenome and mRNA, and by primer extension analysis, which examined antigenome and mRNA initiation. With regard to transcription, nt 36 to 43, located immediately upstream of the GS signal, were found to be necessary for optimal levels of mRNA synthesis, although the GS signal in conjunction with the 3′-terminal region of Le was sufficient to direct accurate mRNA synthesis initiation. With regard to replication, the first 15 nt of Le were found to be sufficient to direct initiation of antigenome synthesis, but nt 16 to 34 were required in addition for efficient encapsidation and production of full-length antigenome. Analysis of transcripts produced from di- and tricistronic minigenomes indicated that a significant proportion of abortive replicases continue RNA synthesis to the end of the first gene and then continue in a transcription mode along the remainder of the genome.
Phosphorylation and Proteolytic Cleavage of Gag Proteins in Budded Simian Immunodeficiency Virus
Authors: Rue Sarah M. , Roos Jason W. , Tarwater Patrick M. , Clements Janice E. , Barber Sheila A.
Published: January 2005

The lentiviral Gag polyprotein (Pr55
Gag
) is cleaved by the viral protease during the late stages of the virus life cycle. Proteolytic cleavage of Pr55
Gag
is necessary for virion maturation, a structural rearrangement required for infectivity that occurs in budded virions. In this study, we investigate the relationship between phosphorylation of capsid (CA) domains in Pr55
Gag
and its cleavage intermediates and their cleavage by the viral protease in simian immunodeficiency virus (SIV). First, we demonstrate that phosphorylated forms of Pr55
Gag
, several CA-containing cleavage intermediates of Pr55
Gag
, and the free CA protein are detectable in SIV virions but not in virus-producing cells, indicating that phosphorylation of these CA-containing Gag proteins may require an environment that is unique to the virion. Second, we show that the CA domain of Pr55
Gag
can be phosphorylated in budded virus and that this phosphorylation does not require the presence of an active viral protease. Further, we provide evidence that CA domains (i.e., incompletely cleaved CA) are phosphorylated to a greater extent than free (completely cleaved) CA and that CA-containing Gag proteins can be cleaved by the viral protease in SIV virions. Finally, we demonstrate that Pr55
Gag
and several of its intermediates, but not free CA, are actively phosphorylated in budded virus. Taken together, these data indicate that, in SIV virions, phosphorylation of CA domains in Pr55
Gag
and several of its cleavage intermediates likely precedes the cleavage of these domains by the viral protease.
Genomic and Bioinformatics Analysis of HAdV-4, a Human Adenovirus Causing Acute Respiratory Disease: Implications for Gene Therapy and Vaccine Vector Development
Authors: Purkayastha Anjan , Ditty Susan E. , Su Jing , McGraw John , Hadfield Ted L. , Tibbetts Clark , Seto Donald
Published: January 2005

Human adenovirus serotype 4 (HAdV-4) is a reemerging viral pathogenic agent implicated in epidemic outbreaks of acute respiratory disease (ARD). This report presents a genomic and bioinformatics analysis of the prototype 35,990-nucleotide genome (GenBank accession no.
AY594253
). Intriguingly, the genome analysis suggests a closer phylogenetic relationship with the chimpanzee adenoviruses (simian adenoviruses) rather than with other human adenoviruses, suggesting a recent origin of HAdV-4, and therefore species E, through a zoonotic event from chimpanzees to humans. Bioinformatics analysis also suggests a pre-zoonotic recombination event, as well, between species B-like and species C-like simian adenoviruses. These observations may have implications for the current interest in using chimpanzee adenoviruses in the development of vectors for human gene therapy and for DNA-based vaccines. Also, the reemergence, surveillance, and treatment of HAdV-4 as an ARD pathogen is an opportunity to demonstrate the use of genome determination as a tool for viral infectious disease characterization and epidemic outbreak surveillance: for example, rapid and accurate low-pass sequencing and analysis of the genome. In particular, this approach allows the rapid identification and development of unique probes for the differentiation of family, species, serotype, and strain (e.g., pathogen genome signatures) for monitoring epidemic outbreaks of ARD.
Inhibition of Virus DNA Replication by Artificial Zinc Finger Proteins
Authors: Sera Takashi
Published: January 2005

Prevention of virus infections is a major objective in agriculture and human health. One attractive approach to the prevention is inhibition of virus replication. To demonstrate this concept in vivo, an artificial zinc finger protein (AZP) targeting the replication origin of theBeet severe curly top virus(BSCTV), a model DNA virus, was created. In vitro DNA binding assays indicated that the AZP efficiently blocked binding of the viral replication protein (Rep), which initiates virus replication, to the replication origin. All of the transgenicArabidopsisplants expressing the AZP showed phenotypes strongly resistant to virus infection, and 84% of the transgenic plants showed no symptom. Southern blot analysis demonstrated that BSCTV replication was completely suppressed in the transgenic plants. Since the mechanism of viral DNA replication is well conserved among plants and mammals, this approach could be applied not only to agricultural crop protection but also to the prevention of virus infections in humans.
Increased Efficiency of Phorbol Ester-Induced Lytic Reactivation of Kaposi's Sarcoma-Associated Herpesvirus during S Phase
Authors: McAllister Shane C. , Hansen Scott G. , Messaoudi Ilhem , Nikolich-Zugich Janko , Moses Ashlee V.
Published: January 2005

Expression of Kaposi's sarcoma-associated herpesvirus (KSHV) lytic genes is thought to be essential for the establishment and progression of KSHV-induced diseases. The inefficiency of lytic reactivation in various in vitro systems hampers the study of lytic genes in the context of whole virus. We report here increased expression of KSHV lytic genes and increased release of progeny virus when synchronized cultures of body cavity-based lymphoma-1 cells are treated with a phorbol ester during S phase of the cell cycle.
Detection and Partial Characterization of Simian Immunodeficiency Virus SIVsm Strains from Bush Meat Samples from Rural Sierra Leone
Authors: Apetrei Cristian , Metzger Michael J. , Richardson David , Ling Binhua , Telfer Paul T. , Reed Patricia , Robertson David L. , Marx Preston A.
Published: January 2005

Human
immunodeficiency virus type 2 (HIV-2) originated from simian
immunodeficiency viruses (SIVs) that naturally infect sooty mangabeys
(SMs;
Cercocebus atys
). In order to further investigate the
relationship between HIV-2 and SIVsm, the SIV specific to the SM, we
characterized seven new SIVsm strains from SMs sold in Sierra Leone
markets as bush meat. The
gag
,
pol
, and
env
sequences showed that, while the viruses of all seven SMs belonged to
the SIVsm-HIV-2 lineage, they were highly divergent viruses, in
spite of the fact that most of the samples originated from the same
geographical region. They clustered in three lineages, two of which
have been previously reported. Two of the new SIVsm strains clustered
differently in
gag
and
env
phylogenetic trees,
suggesting SIVsm recombination that had occurred in the past. In spite
of the fact that our study doubles the number of known SIVsm strains
from wild SMs, none of the simian strains were close to the groups in
which HIV-2 was epidemic (groups A and
B).
Latent Membrane Protein 1 Deletion Mutants Accumulate in Reed-Sternberg Cells of Human Immunodeficiency Virus-Related Hodgkin's Lymphoma
Authors: Guidoboni Massimo , Ponzoni Maurilio , Caggiari Laura , Lettini Antonia A. , Vago Luca , De Re Valli , Gloghini Annunziata , Zancai Paola , Carbone Antonino , Boiocchi Mauro , Dolcetti Riccardo
Published: January 2005

The origin and biological significance of deletions at the 3′ end of the Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP-1) gene are still controversial. We herein demonstrate that LMP-1 deletion mutants are highly associated with human immunodeficiency virus-related Hodgkin's lymphoma (HIV-HL) of Italian patients (29 of 31 cases; 93.5%), a phenomenon that is not due to a peculiar distribution of EBV strains in this area. In fact, although HIV-HL patients are infected by multiple EBV variants, we demonstrate that LMP-1 deletion mutants preferentially accumulate within neoplastic tissues. Subcloning and sequencing of the 3′ LMP-1 ends of two HIV-HL genes in which both variants were present showed the presence of molecular signatures suggestive of a likely derivation of the LMP-1 deletion mutant from a nondeletion ancestor. This phenomenon likely occurs within tumor cells in vivo, as shown by the detection of both LMP-1 variants in single microdissected Reed-Sternberg cells, and may at least in part explain the high prevalence of LMP-1 deletions associated with HIV-HL.
THE INTERIORS OF GIANT PLANETS: Models and Outstanding Questions
Authors: Guillot Tristan
Published: January 2005

▪ Abstract  We know that giant planets played a crucial role in the making of our Solar System. The discovery of giant planets orbiting other stars is a formidable opportunity to learn more about these objects, what their composition is, how various processes influence their structure and evolution, and most importantly how they form. Jupiter, Saturn, Uranus, and Neptune can be studied in detail, mostly from close spacecraft flybys. We can infer that they are all enriched in heavy elements compared to the Sun, with the relative global enrichments increasing with distance to the Sun. We can also infer that they possess dense cores of varied masses. The intercomparison of presently characterized extrasolar giant planets shows that they are also mainly made of hydrogen and helium, but that they either have significantly different amounts of heavy elements, have had different orbital evolutions, or both. Hence, many questions remain and need to be answered to make significant progress on the origins of planets.

Pourquoi l'azur muet et l'espace insondable?

Pourquoi les astres d'or fourmillant comme un sable?

Arthur Rimbaud—Soleil et chair
Molecular mechanisms underlying K+ current downregulation in canine tachycardia-induced heart failure
Authors: Akar Fadi G. , Wu Richard C. , Juang George J. , Tian Yanli , Burysek Mirka , DiSilvestre Deborah , Xiong Wei , Armoundas Antonis A. , Tomaselli Gordon F.
Published: January 2005

Heart failure (HF) is characterized by marked prolongation of action potential duration and reduction in cellular repolarization reserve. These changes are caused in large part by HF-induced K+ current downregulation. Molecular mechanisms underlying these changes remain unclear. We determined whether downregulation of K+ currents in a canine model of tachycardia-induced HF is caused by altered expression of underlying K+ channel α- and β-subunits encoding these currents. K+ channel subunit expression was quantified in normal and failing dogs at the mRNA and protein levels in epicardial (Epi), midmyocardial (Mid), and endocardial (Endo) layers of left ventricle. Analysis of mRNA and protein levels of candidate genes encoding the transient outward K+ current ( Ito) revealed marked reductions in canine cKv4.3 expression in HF in Epi (44% mRNA, 39% protein), Mid (52% mRNA, 34% protein), and Endo (49% mRNA, 73% protein) layers and a paradoxical enhancement (41% Epi, 97% Mid, 113% Endo) in cKv1.4 protein levels, without significant changes in Kv channel-interacting protein cKChIP2 expression. Expression of cKir2.1, the gene underlying inward rectifier K+ current ( IK1), was unaffected by HF at mRNA and protein levels despite significant reduction in IK1, whereas canine ether-à-go-go-related gene (cERG), which encodes the rapidly activating component of the delayed rectifier current ( IK), exhibited increased protein expression. HF was not accompanied by significant changes in cKvLQT1 or cMinK mRNA and protein levels. These data indicate that 1) downregulation of Ito in HF is associated with decreased cKv4.3 and not cKv1.4 or cKChIP2, and 2) alterations in both the rapidly activating and slowly activating components of IK as well as IK1 in nonischemic dilated cardiomyopathy are not caused by changes in either transcript or immunoreactive protein levels of relevant channel subunits, which suggests posttranslational modification of these currents by HF.
Overexpression of elastin fragments in infarcted myocardium attenuates scar expansion and heart dysfunction
Authors: Mizuno Tomohiro , Mickle Donald A. G. , Kiani Chris G. , Li Ren-Ke
Published: January 2005

Ventricular dilation after myocardial infarction can cause heart failure. Increasing strength and elasticity in the infarct region might prevent ventricular dilation. Because elastin provides strength, extensibility, and resilience to tissues and maintains tissue architecture, we studied the effect of elastin expression in the infarct on scar expansion and heart function. COS-7 cells transfected with a plasmid with an elastin gene fragment or a vector were seeded into a Gelfoam mesh and cultured. Mechanical stretch test ( n = 5/group) showed that the elastin mesh was more elastic ( P < 0.05) and tensile ( P < 0.05) than the vector mesh. In an in vivo study in rats, 6 days after left anterior descending coronary artery ligation, COS-7 cells (Cell group, n = 7) or COS-7 cells with elastin gene (Elastin group, n = 9) or vector (Vector group, n = 9) were transplanted into the infarct; infarcted rats served as controls ( n = 7). Over 8 wk the Cell group did not demonstrate effects on scar expansion and deterioration of heart function vs. controls. In contrast, infarct expansion was smaller and heart function was better maintained in the Elastin group vs. the Vector group ( P < 0.05). At 8 wk after cell transplantation Langendorff data showed that the Elastin group had greater ( P < 0.01) developed pressure and a smaller left ventricular volume than the Vector group. Western blot and histology showed accumulated elastin in the Elastin group infarct. Changing the extracellular matrix composition of a myocardial infarct by increasing elastin fragment content attenuated scar expansion, ventricular dilation, and onset of heart dysfunction.
Structural adaptation increases predicted perfusion capacity after vessel obstruction in arteriolar arcade network of pig skeletal muscle
Authors: Gruionu Gabriel , Hoying James B. , Gruionu Lucian G. , Laughlin M. Harold , Secomb Timothy W.
Published: January 2005

Arteriolar arcades provide alternate pathways for blood flow after obstruction of arteries or arterioles such as occurs in stroke and coronary and peripheral vascular disease. When obstruction is prolonged, remaining vessels adjust their diameters chronically in response to altered hemodynamic and metabolic conditions. Here, the effectiveness of arcades in maintaining perfusion both immediately following obstruction and after structural adaptation was examined. Morphometric data from a vascular casting of the pig triceps brachii muscle and published data were used to develop a computational model for the hemodynamics and structural adaptation of the arcade network between two feed artery branches, FA1 and FA2. The predicted total flow to capillaries (Q̇TA) in the region initially supplied by FA2 decreased to 26% of the normal value immediately after FA2 obstruction but was restored to 78% of the normal value after adaptation. After obstruction of 1–10 randomly selected arcade segments, Q̇TAwas on average 18% higher in the arcade network than in a corresponding two-tree network without arcades. Structural adaptation increased Q̇TAby an additional 16% in the arcade network but had almost no effect in the two-tree network. These results indicate that arcades can partially maintain blood flow after vascular blockage and that this effect is substantially enhanced by structural adaptation.
Mechanistic model of cardiac energy metabolism predicts localization of glycolysis to cytosolic subdomain during ischemia
Authors: Zhou Lufang , Salem Jennifer E. , Saidel Gerald M. , Stanley William C. , Cabrera Marco E.
Published: January 2005

A new multidomain mathematical model of cardiac cellular metabolism was developed to simulate metabolic responses to reduced myocardial blood flow. The model is based on mass balances and reaction kinetics that describe transport and metabolic processes of 31 key chemical species in cardiac tissue. The model has three distinct domains (blood, cytosol, and mitochondria) with interdomain transport of chemical species. In addition to distinguishing between cytosol and mitochondria, the model includes a subdomain in the cytosol to account for glycolytic metabolic channeling. Myocardial ischemia was induced by a 60% reduction in coronary blood flow, and model simulations were compared with experimental data from anesthetized pigs. Simulations with a previous model without compartmentation showed a slow activation of glycogen breakdown and delayed lactate production compared with experimental results. The addition of a subdomain for glycolysis resulted in simulations showing faster rates of glycogen breakdown and lactate production that closely matched in vivo experimental data. The dynamics of redox (NADH/NAD+) and phosphorylation (ADP/ATP) states were also simulated. These controllers are coupled to energy transfer reactions and play key regulatory roles in the cytosol and mitochondria. Simulations showed a similar dynamic response of the mitochondrial redox state and the rate of pyruvate oxidation during ischemia. In contrast, the cytosolic redox state displayed a time response similar to that of lactate production. In conclusion, this novel mechanistic model effectively predicted the rapid activation of glycogen breakdown and lactate production at the onset of ischemia and supports the concept of localization of glycolysis to a subdomain of the cytosol.
Reduction of heart rate by chronic β1-adrenoceptor blockade promotes growth of arterioles and preserves coronary perfusion reserve in postinfarcted heart
Authors: Dedkov Eduard I. , Christensen Lance P. , Weiss Robert M. , Tomanek Robert J.
Published: January 2005

Adequate growth of coronary vasculature in the remaining left ventricular (LV) myocardium after myocardial infarction (post-MI) is a crucial factor for myocyte survival and performance. We previously demonstrated that post-MI coronary angiogenesis can be stimulated by bradycardia induced with the ATP-sensitive K+channel antagonist alinidine. In this study, we tested the hypothesis that heart rate reduction with β-blockade may also induce coronary growth in the post-MI heart. Transmural MI was induced in 12-mo-old male Sprague-Dawley rats by occlusion of the left anterior descending coronary artery. Bradycardia was induced by administration of the β-adrenoceptor blocker atenolol (AT) via drinking water (30 mg/day). Three groups of rats were compared: 1) control/sham (C/SH), 2) MI, and 3) MI + AT. In the MI + AT rats, heart rate was consistently reduced by 25–28% compared with C/SH rats. At 4 wk after left anterior descending coronary ligation, infarct size was similar in MI and MI + AT rats (67.1 and 61.5%, respectively), whereas a greater ventricular hypertrophy occurred in bradycardic rats, as indicated by a higher ventricular weight-to-body weight ratio (3.4 ± 0.1 vs. 2.8 ± 0.1 mg/g in MI rats). Analysis of LV function revealed a smaller drop in ejection fraction in the MI + AT than in the MI group (∼24 vs. ∼35%). Furthermore, in MI + AT rats, maximal coronary conductance and coronary perfusion reserve were significantly improved compared with the MI group. The better myocardial perfusion indexes in MI + AT rats were associated with a greater increase in arteriolar length density than in the MI group. Thus chronic reduction of heart rate induced with β-selective blockade promotes growth of coronary arterioles and, thereby, facilitates regional myocardial perfusion in post-MI hearts.
MnSOD in mouse heart: acute responses to ischemic preconditioning and ischemia-reperfusion injury
Authors: Jin Zhu-Qiu , Zhou Hui-Zhong , Cecchini Gary , Gray Mary O. , Karliner Joel S.
Published: January 2005

Manganese superoxide dismutase (MnSOD) is one of the main antioxidant enzymes that protects the heart against ischemia-reperfusion (I/R) injury. Ischemic preconditioning (IPC) is a short period of ischemia-reperfusion that reduces subsequent prolonged I/R injury. Although MnSOD localizes in mitochondria, the immediate subcellular distribution of MnSOD in heart after IPC and I/R has not been studied. In a Langendorff mouse heart model, IPC significantly improved cardiac function and reduced the infarction size induced by I/R. Immunoblotting and double immunostaining in fresh preparations revealed that I/R resulted in an increase in cytosolic MnSOD content accompanied by the release of cytochrome c. In contrast, IPC increased mitochondrial MnSOD and reduced cytosolic MnSOD and cytochrome c release induced by I/R. We found that compared with freshly prepared fractions, the freeze-thaw approach results in mitochondrial integrity disruption and release of large amounts of MnSOD into the cytosol along with mitochondrial markers even in the absence of I/R. In contrast, fresh preparations exhibit early MnSOD release into the cytosol after I/R that is prevented by IPC and cyclosporin A administration.
Oxygen release from low and normal P50Hb vesicles in transiently occluded arterioles of the hamster window model
Authors: Sakai Hiromi , Cabrales Pedro , Tsai Amy G. , Tsuchida Eishun , Intaglietta Marcos
Published: January 2005

A phospholipid vesicle encapsulating Hb [Hb vesicle (HbV)] has been developed as a transfusion alternative. One characteristic of HbV is that the O2affinity [Po2at which Hb is 50% saturated (P50)] of Hb can be easily regulated by the amount of the coencapsulated allosteric effector pyridoxal 5′-phosphate. In this study, we prepared two HbVs with different P50s (8 and 29 mmHg, termed HbV8and HbV29, respectively) and observed their O2-releasing behavior from an occluded arteriole in a hamster skinfold window model. Conscious hamsters received HbV8or HbV29at a dose rate of 7 ml/kg. In the microscopic view, an arteriole (diameter: 53.0 ± 6.6 μm) was occluded transcutaneously by a glass pipette on a manipulator, and the reduction of the intra-arteriolar Po2100 μm down from the occlusion was measured by the phosphorescence quenching of preinfused Pd-porphyrin. The baseline arteriolar Po2(50–52 mmHg) decreased to about 5 mmHg for all the groups. Occlusion after HbV8infusion showed a slightly slower rate of Po2reduction compared with that after HbV29infusion. The arteriolar O2content was calculated at each reducing Po2in combination with the O2equilibrium curves of HbVs, and it was clarified that HbV8showed a significantly slower rate of O2release compared with HbV29and was a primary source of O2(maximum fraction, 0.55) overwhelming red blood cells when the Po2was reduced (e.g., <10 mmHg) despite a small dosage of HbV. This result supports the possible utilization of Hb-based O2carriers with lower P50for oxygenation of ischemic tissues.