bluuebunny/crossref_metadata_2025 huggingface dataset:
Subgenomic Negative-Strand RNA Function during Mouse Hepatitis Virus Infection
Authors: Baric Ralph S. , Yount Boyd
Published: July 2002

Mouse hepatitis virus (MHV)-infected cells contain full-length and subgenomic-length positive- and negative-strand RNAs. The origin and function of the subgenomic negative-strand RNAs is controversial. In this report we demonstrate that the synthesis and molar ratios of subgenomic negative strands are similar in alternative host cells, suggesting that these RNAs function as important mediators of positive-strand synthesis. Using kinetic labeling experiments, we show that the full-length and subgenomic-length replicative form RNAs rapidly accumulate and then saturate with label, suggesting that the subgenomic-length negative strands are the principal mediators of positive-strand synthesis. Using cycloheximide, which preferentially inhibits negative-strand and to a lesser extent positive-strand synthesis, we demonstrate that cycloheximide treatment equally inhibits full-length and subgenomic-length negative-strand synthesis. Importantly, following treatment, previously transcribed negative strands remain in transcriptionally active complexes even in the absence of new negative-strand synthesis. These findings indicate that the subgenomic-length negative strands are the principal templates of positive-strand synthesis during MHV infection.
Mucosal Delivery of Inactivated Influenza Vaccine Induces B-Cell-Dependent Heterosubtypic Cross-Protection against Lethal Influenza A H5N1 Virus Infection
Authors: Tumpey Terrence M. , Renshaw Mary , Clements John D. , Katz Jacqueline M.
Published: July 2002

Influenza vaccines that induce greater cross-reactive or heterosubtypic immunity (Het-I) may overcome limitations in vaccine efficacy imposed by the antigenic variability of influenza A viruses. We have compared mucosal versus traditional parenteral administration of inactivated influenza vaccine for the ability to induce Het-I in BALB/c mice and evaluated a modifiedEscherichia coliheat-labile enterotoxin adjuvant, LT(R192G), for augmentation of Het-I. Mice that received three intranasal (i.n.) immunizations of H3N2 vaccine in the presence of LT(R192G) were completely protected against lethal challenge with a highly pathogenic human H5N1 virus and had nasal and lung viral titers that were at least 2,500-fold lower than those of control mice receiving LT(R192G) alone. In contrast, mice that received three vaccinations of H3N2 vaccine subcutaneously in the presence or absence of LT(R192G) or incomplete Freund's adjuvant were not protected against lethal challenge and had no significant reductions in tissue virus titers observed on day 5 post-H5N1 virus challenge. Mice that were i.n. administered H3N2 vaccine alone, without LT(R192G), displayed partial protection against heterosubtypic challenge. The immune mediators of Het-I were investigated. The functional role of B and CD8+T cells in Het-I were evaluated by using gene-targeted B-cell (IgH-6−/−)- or β2-microglobulin (β2m−/−)-deficient mice, respectively. β2m−/−but not IgH-6−/−vaccinated mice were protected by Het-I and survived a lethal infection with H5N1, suggesting that B cells, but not CD8+T cells, were vital for protection of mice against heterosubtypic challenge. Nevertheless, CD8+T cells contributed to viral clearance in the lungs and brain tissues of heterotypically immune mice. Mucosal but not parenteral vaccination induced subtype cross-reactive lung immunoglobulin G (IgG), IgA, and serum IgG anti-hemagglutinin antibodies, suggesting the presence of a common cross-reactive epitope in the hemagglutinins of H3 and H5. These results suggest a strategy of mucosal vaccination that stimulates cross-protection against multiple influenza virus subtypes, including viruses with pandemic potential.
Novel Baculovirus Expression Vectors That Provide Sialylation of Recombinant Glycoproteins in Lepidopteran Insect Cells
Authors: Jarvis Donald L. , Howe Dale , Aumiller Jared J.
Published: July 2002

This report describes novel baculovirus vectors designed to express mammalian β1,4-galactosyltransferase and α2,6-sialyltransferase genes at early times after infection. Sf9 cells infected with these viral vectors, unlike cells infected with a wild-type baculovirus, produced a sialylated viral glycoprotein during the late phase of infection. Thus, the two mammalian glycosyltransferases encoded by these viral vectors are necessary and sufficient for sialylation of a foreign glycoprotein in insect cells under the conditions used in this study. While some of the new baculovirus vectors described in this study produced less, one produced wild-type levels of infectious budded virus progeny.
A Nonviral Peptide Can Replace the Entire N Terminus of Zucchini Yellow Mosaic Potyvirus Coat Protein and Permits Viral Systemic Infection
Authors: Arazi T. , Shiboleth Y. M. , Gal-On A.
Published: July 2002

Systematic deletion and peptide tagging of the amino-terminal domain (NT, ∼43 amino acids) of an attenuated zucchini yellow mosaic potyvirus (ZYMV-AGII) coat protein (CP) were used to elucidate its role in viral systemic infection. Deletion mutants truncated by 8, 13, and 33 amino acid residues from the CP-NT 5′ end were systemically infectious and produced symptoms similar to those of the AGII virus. Tagging these deletion mutants with either human c-Myc (Myc) or hexahistidine peptides maintained viral infectivity. Similarly, addition of these peptides to the intact AGII CP-NT did not affect viral life cycle. To determine which parts, if any, of the CP-NT are essential for viral systemic infection, a series of Myc-tagged mutants with 8 to 43 amino acids removed from the CP-NT were constructed. All Myc-tagged CP-NT deletion mutants, including those from which virtually all the viral CP-NT had been eliminated, were able to encapsidate and cause systemic infection. Furthermore, chimeric viruses with deletions of up to 33 amino acids from CP-NT produced symptoms indistinguishable from those caused by the parental AGII virus. In contrast to CP-NT Myc fusion, addition of the foot-and-mouth disease virus (FMDV) immunogenic epitope to AGII CP-NT did not permit systemic infection. However, fusion of the Myc peptide to the N terminus of the FMDV peptide restored the capability of the virus to spread systemically. We have demonstrated that all CP-NT fused peptides were exposed on the virion surface, masking natural CP immunogenic determinants. Our findings demonstrate that CP-NT is not essential for ZYMV spread and that it can be replaced by an appropriate foreign peptide while maintaining systemic infectivity.
Insertion of Capsid Proteins from Nonenveloped Viruses into the Retroviral Budding Pathway
Authors: Krishna Neel K. , Wills John W.
Published: July 2002

Retroviral Gag proteins direct the assembly and release of virus particles from the plasma membrane. The budding machinery consists of three small domains, the M (membrane-binding), I (interaction), and L (late or “pinching-off”) domains. In addition, Gag proteins contain sequences that control particle size. For Rous sarcoma virus (RSV), the size determinant maps to the capsid (CA)-spacer peptide (SP) sequence, but it functions only when I domains are present to enable particles of normal density to be produced. Small deletions throughout the CA-SP sequence result in the release of particles that are very large and heterogeneous, even when I domains are present. In this report, we show that particles of relatively uniform size and normal density are released by budding when the size determinant and I domains in RSV Gag are replaced with capsid proteins from two unrelated, nonenveloped viruses: simian virus 40 and satellite tobacco mosaic virus. These results indicate that capsid proteins of nonenveloped viruses can interact among themselves within the context of Gag and be inserted into the retroviral budding pathway merely by attaching the M and L domains to their amino termini. Thus, the differences in the assembly pathways of enveloped and nonenveloped viruses may be far simpler than previously thought.
Herpes Simplex Virus Type 1 Latency-Associated Transcript Gene Promotes Neuronal Survival
Authors: Thompson R. L. , Sawtell N. M.
Published: July 2002

A complex interaction has evolved between the host's peripheral nervous system (PNS) and herpes simplex virus type 1 (HSV-1). Sensory neurons are permissive for viral replication, yet the virus can also enter a latent state in these cells. The interplay of viral and neuronal signals that regulate the switch between the viral lytic and latent states is not understood. The latency-associated transcript (LAT) regulates the establishment of the latent state and is required for >65% of the latent infections established by HSV-1 (R. L. Thompson and N. M. Sawtell, J. Virol. 71:5432–5440, 1997). To further investigate how LAT functions, a 1.9-kb deletion that includes the entire LAT promoter and 827 bp of the 5′ end of the primary LAT mRNA was introduced into strain 17syn+. The wild-type parent, three independently derived deletion mutants, and two independently derived genomically rescued variants of the mutants were analyzed in a mouse ocular model. The number of latent sites established in trigeminal ganglion (TG) neurons was determined using a single-cell quantitative PCR assay for the viral genome on purified TG neurons. It was found that the LAT null mutants established ∼75% fewer latent infections than the number established by the parental strain or rescued variant. The reduced establishment phenotype of LAT null mutants was due at least in part to a dramatic increase in the loss of TG neurons in animals infected with the LAT mutants. Over half of the neurons in the TG were destroyed following infection with the LAT mutants, and this was significantly more than were lost following infection with wild type. This is the first demonstration that the HSV LAT locus prevents the destruction of sensory neurons. The death of these neurons did not appear to be the result of increased apoptosis as measured by a terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling assay. Animals latently infected with the LAT null mutants reactivated less frequently in vivo and this was consistent with the reduction in the number of neurons in which latency was established. Thus, one function of the LAT gene is to protect sensory neurons and enhance the establishment of latency in the PNS.
Bovine Herpesvirus 1 Tegument Protein VP22 Interacts with Histones, and the Carboxyl Terminus of VP22 Is Required for Nuclear Localization
Authors: Ren Xiaodi , Harms Jerome S. , Splitter Gary A.
Published: July 2002

The bovine herpesvirus 1 (BHV-1) UL49 gene encodes a viral tegument protein termed VP22. UL49 homologs are conserved among alphaherpesviruses. Interestingly, the BHV-1 VP22 deletion mutant virus is asymptomatic and avirulent in infected cattle but produces only a slight reduction in titer in vitro. Attenuation of the BHV-1 VP22 deletion mutant virus in vivo suggests that VP22 plays a functional role in BHV-1 replication. In herpes simplex virus type 1, the VP22 homolog was previously shown to interact with another tegument protein,VP16, the α-transinducing factor in vitro. In this report, we show that (i) the nuclear targeting of VP22 is independent of other viral factors, (ii) the carboxyl terminus of VP22 is required for its nuclear localization, (iii) VP22 associates with histones and nucleosomes, (iv) an antihistone monoclonal antibody cross-reacts with VP22, and (v) acetylation of histone H4 is decreased in VP22-expressing cells as well as virus-infected cells. Our data suggest that VP22 may have a modulatory function during BHV-1 infection.
Role of Interleukin-4 (IL-4), IL-12, and Gamma Interferon in Primary and Vaccine-Primed Immune Responses to Friend Retrovirus Infection
Authors: Dittmer Ulf , Peterson Karin E. , Messer Ron , Stromnes Ingunn M. , Race Brent , Hasenkrug Kim J.
Published: July 2002

The immunological resistance of a host to viral infections may be strongly influenced by cytokines such as interleukin-12 (IL-12) and gamma interferon (IFN-γ), which promote T helper type 1 responses, and IL-4, which promotes T helper type 2 responses. We studied the role of these cytokines during primary and secondary immune responses against Friend retrovirus infections in mice. IL-4- and IL-12-deficient mice were comparable to wild-type B6 mice in the ability to control acute and persistent Friend virus infections. In contrast, more than one-third of the IFN-γ-deficient mice were unable to maintain long-term control of Friend virus and developed gross splenomegaly with high virus loads. Immunization with a live attenuated vaccine virus prior to challenge protected all three types of cytokine-deficient mice from viremia and high levels of spleen virus despite the finding that the vaccinated IFN-γ-deficient mice were unable to class switch from immunoglobulin M (IgM) to IgG virus-neutralizing antibodies. The results indicate that IFN-γ plays an important role during primary immune responses against Friend virus but is dispensable during vaccine-primed secondary responses.
C Terminus of Infectious Bursal Disease Virus Major Capsid Protein VP2 Is Involved in Definition of the T Number for Capsid Assembly
Authors: Castón José R. , Martı́nez-Torrecuadrada Jorge L. , Maraver Antonio , Lombardo Eleuterio , Rodrı́guez José F. , Casal J. Ignacio , Carrascosa José L.
Published: July 2002

Infectious bursal disease virus
(IBDV), a member of the
Birnaviridae
family, is a double-stranded RNA virus. The IBDV capsid is formed by two major structural proteins, VP2 and VP3, which assemble to form a T=13 markedly nonspherical capsid. During viral infection, VP2 is initially synthesized as a precursor, called VPX, whose C end is proteolytically processed to the mature form during capsid assembly. We have computed three-dimensional maps of IBDV capsid and virus-like particles built up by VP2 alone by using electron cryomicroscopy and image-processing techniques. The IBDV single-shelled capsid is characterized by the presence of 260 protruding trimers on the outer surface. Five classes of trimers can be distinguished according to their different local environments. When VP2 is expressed alone in insect cells, dodecahedral particles form spontaneously; these may be assembled into larger, fragile icosahedral capsids built up by 12 dodecahedral capsids. Each dodecahedral capsid is an empty T=1 shell composed of 20 trimeric clusters of VP2. Structural comparison between IBDV capsids and capsids consisting of VP2 alone allowed the determination of the major capsid protein locations and the interactions between them. Whereas VP2 forms the outer protruding trimers, VP3 is found as trimers on the inner surface and may be responsible for stabilizing functions. Since elimination of the C-terminal region of VPX is correlated with the assembly of T=1 capsids, this domain might be involved (either alone or in cooperation with VP3) in the induction of different conformations of VP2 during capsid morphogenesis.
Neuroadapted Yellow Fever Virus 17D: Genetic and Biological Characterization of a Highly Mouse-Neurovirulent Virus and Its Infectious Molecular Clone
Authors: Chambers Thomas J. , Nickells Michael
Published: July 2002

A neuroadapted strain of yellow fever virus (YFV) 17D derived from a multiply mouse brain-passaged virus (Porterfield YF17D) was additionally passaged in SCID and normal mice. The virulence properties of this virus (SPYF) could be distinguished from nonneuroadapted virus (YF5.2iv, 17D infectious clone) by decreased average survival time in SCID mice after peripheral inoculation, decreased average survival time in normal adult mice after intracerebral inoculation, and occurrence of neuroinvasiveness in normal mice. SPYF exhibited more efficient growth in peripheral tissues of SCID mice than YF5.2iv, resulting in a more rapid accumulation of virus burden, but with low-titer viremia, at the time of fatal encephalitis. In cell culture, SPYF was less efficient in replication than YF5.2iv in all cell lines tested. The complete nucleotide sequence of SPYF revealed 29 nucleotide substitutions relative to YF5.2iv, and these were distributed throughout the genome. There were a total of 13 predicted amino acid substitutions, some of which correspond to known differences among the Asibi, French viscerotropic virus, French neurotropic vaccine, and YF17D vaccine strains. The envelope (E) protein contained five substitutions, within all three functional domains. Substitutions were also present in regions encoding the NS1, NS2A, NS4A, and NS5 proteins and in the 3′ untranslated region (UTR). Construction of YFV harboring all of the identified coding nucleotide substitutions and those in the 3′ UTR yielded a virus whose cell culture and pathogenic properties, particularly neurovirulence and neuroinvasiveness for SCID mice, generally resembled those of the original SPYF isolate. These findings implicate the E protein and possibly other regions of the genome as virulence determinants during pathogenesis of neuroadapted YF17D virus in mice. The determinants affect replication efficiency in both neural and extraneural tissues of the mouse and confer some limited host-range differences in cultured cells of nonmurine origin.
Moloney Murine Leukemia Virus Integrase Protein Augments Viral DNA Synthesis in Infected Cells
Authors: Lai Lilin , Liu Hongmei , Wu Xiaoyun , Kappes John C.
Published: July 2002

Mutations in the
IN
domain of retroviral DNA may affect multiple steps of the virus life cycle, suggesting that the IN protein may have other functions in addition to its integration function. We previously reported that the human immunodeficiency virus type 1 IN protein is required for efficient viral DNA synthesis and that this function requires specific interaction with other viral components but not enzyme (integration) activity. In this report, we characterized the structure and function of the Moloney murine leukemia virus (MLV) IN protein in viral DNA synthesis. Using an MLV vector containing green fluorescent protein as a sensitive reporter for virus infection, we found that mutations in either the catalytic triad (D184A) or the HHCC motif (H61A) reduced infectivity by approximately 1,000-fold. Mutations that deleted the entire IN (ΔIN) or 34 C-terminal amino acid residues (Δ34) were more severely defective, with infectivity levels consistently reduced by 10,000-fold. Immunoblot analysis indicated that these mutants were similar to wild-type MLV with respect to virion production and proteolytic processing of the Gag and Pol precursor proteins. Using semiquantitative PCR to analyze viral cDNA synthesis in infected cells, we found the Δ34 and ΔIN mutants to be markedly impaired while the D184A and H61A mutants synthesized cDNA at levels similar to the wild type. The DNA synthesis defect was rescued by complementing the Δ34 and ΔIN mutants in
trans
with either wild-type IN or the D184A mutant IN, provided as a Gag-IN fusion protein. However, the DNA synthesis defect of ΔIN mutant virions could not be complemented with the Δ34 IN mutant. Taken together, these analyses strongly suggested that the MLV IN protein itself is required for efficient viral DNA synthesis and that this function may be conserved among other retroviruses.
Antigenicity and Immunogenicity of Novel Chimeric Hepatitis B Surface Antigen Particles with Exposed Hepatitis C Virus Epitopes
Authors: Netter Hans J. , Macnaughton Thomas B. , Woo Wai-Ping , Tindle Robert , Gowans Eric J.
Published: July 2002

The small envelope protein of hepatitis B virus (HBsAg-S) can self-assemble into highly organized virus like particles (VLPs) and induce an effective immune response. In this study, a restriction enzyme site was engineered into the cDNA of HBsAg-S at a position corresponding to the exposed site within the hydrophilic a determinant region (amino acid [aa] 127–128) to create a novel HBsAg vaccine vector allowing surface orientation of the inserted sequence. We inserted sequences of various lengths from hypervariable region 1 (HVR1) of the hepatitis C virus (HCV) E2 protein containing immunodominant epitopes and demonstrated secretion of the recombinant HBsAg VLPs from transfected mammalian cells. A number of different recombinant proteins were synthesized, and HBsAg VLPs containing inserts up to 36 aa were secreted with an efficiency similar to that of wild-type HBsAg. The HVR1 region exposed on the particles retained an antigenic structure similar to that recognized immunologically during natural infection. VLPs containing epitopes from either HCV-1a or -1b strains were produced that induced strain-specific antibody responses in immunized mice. Injection of a combination of these VLPs induced antibodies against both HVR1 epitopes that resulted in higher titers than were achieved by vaccination with the individual VLPs, suggesting a synergistic effect. This may lead to the development of recombinant particles which are able to induce a broad anti-HCV immune response against the HCV quasispecies or other quasispecies-like infectious agents.
Nuclear Localization and Shuttling of Herpes Simplex Virus Tegument Protein VP13/14
Authors: Donnelly Michelle , Elliott Gillian
Published: July 2002

The herpes simplex virus type 1 gene UL47 encodes the tegument proteins referred to collectively as VP13/14, which are believed to be differentially modified forms of the same protein. Here we show that the major product of the UL47 gene during transient expression is VP14, suggesting that some feature of virus infection is required to produce VP13. We have tagged VP13/14 with green fluorescent protein and have demonstrated that the protein is targeted efficiently to the nucleus, where it often localizes in numerous punctate domains. Furthermore, we show that removal of the N-terminal 127 residues of the protein abrogates nuclear accumulation, and we have identified a 14-amino-acid peptide from this region that is sufficient to function as a nuclear targeting signal and transport a heterologous protein to the nucleus. This short peptide contains two runs of four arginine residues, suggesting that the VP13/14 nuclear localization signal may behave in a manner similar to that of the arginine-rich nuclear localization signals of the retrovirus transactivator proteins Tat, Rev, and Rex. In addition, by using heterokaryon assays, we show that VP13/14 is capable of shuttling between the nucleus and cytoplasm of the cell, a property that may be attributed to three leucine-rich stretches in the C-terminal half of the protein that again bear similarity to the nuclear export signals of Rev and Rex. This is the first demonstration of a tegument protein that is specifically targeted to the nucleus, a feature which may be relevant both during virus entry, when VP13/14 enters the cell as a component of the tegument, and at later times, when large amounts of newly synthesized VP13/14 are present within the cell.
Differences in Cytokine and Chemokine Responses during Neurological Disease Induced by Polytropic Murine Retroviruses Map to Separate Regions of the Viral Envelope Gene
Authors: Peterson Karin E. , Robertson Shelly J. , Portis John L. , Chesebro Bruce
Published: July 2002

Infection of the central nervous system (CNS) by several viruses can lead to upregulation of proinflammatory cytokines and chemokines. In immunocompetent adults, these molecules induce prominent inflammatory infiltrates. However, with immunosuppressive retroviruses, such as human immunodeficiency virus (HIV), little CNS inflammation is observed yet proinflammatory cytokines and chemokines are still upregulated in some patients and may mediate pathogenesis. The present study examined expression of cytokines and chemokines in brain tissue of neonatal mice infected with virulent (Fr98) and avirulent (Fr54) polytropic murine retroviruses. While both viruses infect microglia and endothelia primarily in the white matter areas of the CNS, only Fr98 induces clinical CNS disease. The pathology consists of gliosis with minimal morphological changes and no inflammation, similar to HIV. In the present experiments, mice infected with Fr98 had increased cerebellar mRNA levels of proinflammatory cytokines tumor necrosis factor alpha (TNF-α), TNF-β, and interleukin-1α and chemokines macrophage inflammatory protein-1α (MIP-1α), MIP-1β, monocyte chemoattractant protein 1 (MCP-1), gamma-interferon-inducible protein 10 (IP-10), and RANTES compared to mice infected with Fr54 or mock-infected controls. The increased expression of these genes occurred prior to the development of clinical symptoms, suggesting that these cytokines and chemokines might be involved in induction of neuropathogenesis. Two separate regions of the Fr98 envelope gene are associated with neurovirulence. CNS disease associated with the N-terminal portion of the Fr98
env
gene was preceded by upregulation of cytokines and chemokines. In contrast, disease associated with the central region of the Fr98
env
gene showed no upregulation of cytokines or chemokines and thus did not require increased expression of these genes for disease induction.
New World Arenavirus Clade C, but Not Clade A and B Viruses, Utilizes α-Dystroglycan as Its Major Receptor
Authors: Spiropoulou Christina F. , Kunz Stefan , Rollin Pierre E. , Campbell Kevin P. , Oldstone Michael B. A.
Published: July 2002

α-Dystroglycan (α-DG) has been identified as a major receptor for lymphocytic choriomeningitis virus (LCMV) and Lassa virus, two Old World arenaviruses. The situation with New World arenaviruses is less clear: previous studies demonstrated that Oliveros virus also exhibited high-affinity binding to α-DG but that Guanarito virus did not. To extend these initial studies, several additional Old and New World arenaviruses were screened for entry into mouse embryonic stem cells possessing or lacking α-DG. In addition, representative viruses were further analyzed for direct binding to α-DG by means of a virus overlay protein blot assay technique. These studies indicate that Old World arenaviruses use α-DG as a major receptor, whereas, of the New World arenaviruses, only clade C viruses (i.e., Oliveros and Latino viruses) use α-DG as a major receptor. New World clade A and B arenaviruses, which include the highly pathogenic Machupo, Guanarito, Junin, and Sabia viruses, appear to use a different receptor or coreceptor for binding. Previous studies with LCMV have suggested the need for a small aliphatic amino acid at LCMV GP1 glycoprotein amino acid position 260 to allow high-affinity binding to α-DG. As reported herein, this requirement appears to be broadly applicable to the arenaviruses as determined by more extensive analysis of α-DG receptor usage and GP1 sequences of Old and New World arenaviruses. In addition, GP1 amino acid position 259 also appears to be important, since all arenaviruses showing high-affinity α-DG binding possess a bulky aromatic amino acid (tyrosine or phenylalanine) at this position.
Regulation of Hepatocyte Nuclear Factor 1 Activity by Wild-Type and Mutant Hepatitis B Virus X Proteins
Authors: Li Jie , Xu Zhenming , Zheng Yanyan , Johnson Deborah L. , Ou Jing-hsiung
Published: July 2002

The hepatitis B virus (HBV) core promoter regulates the transcription of two related RNA products named precore RNA and core RNA. Previous studies indicate that a double-nucleotide mutation that occurs frequently during chronic HBV infection converts a nuclear receptor binding site in the core promoter to the binding site of the transcription factor hepatocyte nuclear factor-1 (HNF-1) and specifically suppresses the transcription of the precore RNA. This mutation also changes two codons in the overlapping X protein coding sequence. In this report, we demonstrate that the X protein and its mutant X
mt
can physically bind to HNF-1 both in vitro and in vivo. Further analyses indicate that both X and X
mt
can enhance the gene transactivation and the DNA binding activities of HNF-1. This finding demonstrates for the first time that the X protein can stimulate the DNA binding activity of a homeodomain transcription factor. Interestingly, while both X and X
mt
can stimulate the HNF-1 activities, they differ in their effects: a smaller amount of X
mt
is needed to generate greater transactivation and DNA binding activities of HNF-1. This functional difference between X and X
mt
may have important implications in HBV pathogenesis and is apparently why they have different effects on the core promoter bearing the HNF-1 binding site.
Dominance of CD8 Responses Specific for Epitopes Bound by a Single Major Histocompatibility Complex Class I Molecule during the Acute Phase of Viral Infection
Authors: Mothé Bianca R. , Horton Helen , Carter Donald K. , Allen Todd M. , Liebl Max E. , Skinner Pam , Vogel Thorsten U. , Fuenger Sarah , Vielhuber Kathy , Rehrauer William , Wilson Nancy , Franchini Genoveffa , Altman John D. , Haase Ashley , Picker Louis J. , Allison David B. , Watkins David I.
Published: July 2002

Cytotoxic T-lymphocyte (CTL) responses are thought to control human immunodeficiency virus replication during the acute phase of infection. Understanding the CD8+T-cell immune responses early after infection may, therefore, be important to vaccine design. Analyzing these responses in humans is difficult since few patients are diagnosed during early infection. Additionally, patients are infected by a variety of viral subtypes, making it hard to design reagents to measure their acute-phase immune responses. Given the complexities in evaluating acute-phase CD8+responses in humans, we analyzed these important immune responses in rhesus macaques expressing a common rhesus macaque major histocompatibility complex class I molecule (Mamu-A*01) for which we had developed a variety of immunological assays. We infected eight Mamu-A*01-positive macaques and five Mamu-A*01-negative macaques with the molecularly cloned virus SIVmac239 and determined all of the simian immunodeficiency virus-specific CD8+T-cell responses against overlapping peptides spanning the entire virus. We also monitored the evolution of particular CD8+T-cell responses by tetramer staining of peripheral lymphocytes as well as lymph node cells in situ. In this first analysis of the entire CD8+immune response to autologous virus we show that between 2 and 12 responses are detected during the acute phase in each animal. CTL against the early proteins (Tat, Rev, and Nef) and against regulatory proteins Vif and Vpr dominated the acute phase. Interestingly, CD8+responses against Mamu-A*01-restricted epitopes Tat28-35SL8 and Gag181-189CM9 were immunodominant in the acute phase. After the acute phase, however, this pattern of reactivity changed, and the Mamu-A*01-restricted response against the Gag181-189CM9 epitope became dominant. In most of the Mamu-A*01-positive macaques tested, CTL responses against epitopes bound by Mamu-A*01 dominated the CD8+cellular immune response.
Phylogenetic Analysis of Varicella-Zoster Virus: Evidence of Intercontinental Spread of Genotypes and Recombination
Authors: Muir Winsome Barrett , Nichols Richard , Breuer Judith
Published: July 2002

A heteroduplex mobility assay was used to identify variants of varicella-zoster virus circulating in the United Kingdom and elsewhere. Within the United Kingdom, 58 segregating sites were found out of the 23,266 examined (0.25%), and nucleotide diversity was estimated to be 0.00063. These are an order of magnitude smaller than comparable estimates from herpes simplex virus type 1. Sixteen substitutions were nonsynonymous, the majority of which were clustered within surface-expressed proteins. Extensive genetic correlation between widely spaced sites indicated that recombination has been rare. Phylogenetic analysis of varicella-zoster viruses from four continents distinguished at least three major genetic clades. Most geographical regions contained only one of these three strains, apart from the United Kingdom and Brazil, where two or more strains were found. There was minimal genetic differentiation (one or fewer substitutions in 1,895 bases surveyed) between the samples collected from Africa (Guinea Bissau, Zambia) and the Indian subcontinent (Bangladesh, South India), suggesting recent rapid spread and/or low mutation rates. The geographic pattern of strain distribution would favor a major influence of the former. The genetic uniformity of most virus populations makes recombination difficult to detect. However, at least one probable recombinant between two of the major strains was found among the samples originating from Brazil, where mixtures of genotypes co-occur.
Porcine Endogenous Retrovirus Transmission Characteristics of an Inbred Herd of Miniature Swine
Authors: Oldmixon Beth A. , Wood James C. , Ericsson Thomas A. , Wilson Carolyn A. , White-Scharf Mary E. , Andersson Goran , Greenstein Julia L. , Schuurman Henk-Jan , Patience Clive
Published: July 2002

Here we report the identification of inbred miniature swine that failed to produce human-tropic replication-competent porcine endogenous retroviruses (HTRC PERVs), using in vitro coculture assays. When HTRC PERVs were isolated from transmitting animals, all were recombinant viruses, with the receptor-binding domain of PERV-A combining with PERV-C-related sequences.
Immune-Mediated Protection from Measles Virus-Induced Central Nervous System Disease Is Noncytolytic and Gamma Interferon Dependent
Authors: Patterson Catherine E. , Lawrence Diane M. P. , Echols Lisa A. , Rall Glenn F.
Published: July 2002

Neurons of the mammalian central nervous system (CNS) are an essential and largely nonrenewable cell population. Thus, virus infections that result in neuronal depletion, either by virus-mediated cell death or by induction of the cytolytic immune response, could cause permanent neurological impairment of the host. In a transgenic mouse model of measles virus (MV) infection of neurons, we have previously shown that the host T-cell response was required for resolution of infection in susceptible adult mice. In this report, we show that this protective response did not result in neuronal death, even during the peak of T-cell infiltration into the brain parenchyma. When susceptible mice were intercrossed with specific immune knockout mice, a critical role for gamma interferon (IFN-γ) was identified in protection against MV infection and CNS disease. Moreover, the addition of previously activated splenocytes or recombinant murine IFN-γ to MV-infected primary neurons resulted in the inhibition of viral replication in the absence of neuronal death. Together, these data support the hypothesis that the host immune response can promote viral clearance without concomitant neuronal loss, a process that appears to be mediated by cytokines.
Solubility in the Glycine-H3BO3-H2O System
Authors: Eysseltová Jitka , Dosoudilová Jana
Published: July 2002

Solubility isotherms in the title system were measured at 0, 25, 50, and 70 °C. Regression straight line equations interrelating the solubilities of boric acid and glycine were derived. The activity of water in saturated solutions of the system at 25 °C was determined by vapour pressure measurements.
TPSR, TPR, and TPO Studies of Pd-V2O5/Al2O3 Catalysts in Complete Catalytic Oxidation of Benzene
Authors: Dancheva Snejana , Ilieva Liuba , Kotsev Nikolay , Andreev Atanas
Published: July 2002

Alumina-supported palladium vanadium-pentoxide catalysts were studied by means of temperature-programmed surface reaction, temperature-programmed reduction, temperature-programmed oxidation, and by ESR spectroscopy in the complete oxidation of benzene. The surface reaction measurements indicated that 0.5% Pd - 30% V2O5 / Al2O3 produced most oxidation products, compared to 0.5% Pd / Al2O3 or 30% V2O5 / Al2O3 samples. A drastic fall of peak maxima in temperature-programmed oxidation and reduction spectra indicates the decisive role of palladium in the mixed metal metal oxide system in facilitating the vanadium redox transitions. During initial period of catalysts operation, there proceeds formation of vanadium oxide phases which contain clustered V4+ ions in a matrix of vanadium pentoxide. These ions are responsible for the high activity of the palladium vanadium pentoxide catalyst.
Study of the Electrochemically Initiated Cyanomethylation of Azomethines by Means of the MNDO and AM1 Semiempirical Methods
Authors: Toušek Jaromír , Komenda Jaro , Hess Ulrich
Published: July 2002

The semiempirical methods (MNDO and AM1) have been used in the quantum chemical study of electrochemically initiated cyanomethylation of azomethines. Reaction steps leading to the main product of the studied reaction, β-aminonitrile, have been described. The stationary points on the reaction path have been found and the activation enthalpies of reaction steps have been determined. Three possible mechanisms for the abstraction of a proton from acetonitrile have been described (by azomethine radical anion, by β-aminonitrile anion, and by hydrogenated azomethine anion). At the MNDO level, the abstraction of a proton by azomethine radical anion has the highest activation enthalpy (∆H = 197 kJ mol-1) and the abstraction of a proton by hydrogenated azomethine anion has the lowest activation enthalpy (∆H = 146 kJ mol-1). The activation enthalpy of the next reaction step (the reaction of acetonitrile anion with azomethine) is low compared to the activation enthalpies of the previous reation steps (∆H= 12 kJ mol-1).
Determination of Estimates of Parameters of Calibration Regression Straight Line with Objective Elimination of Remote Measurements
Authors: Jehlička Vladimír , Mach Vladimír
Published: July 2002

An alternative algorithm of calculation of estimates of parameters of calibration regression straight line with elimination of remote measurements has been elaborated for a function f(x) composed of linear-nonlinear or nonlinear-linear parts or, as the case may be, of nonlinear-linear-nonlinear parts. This algorithm makes it possible to objectively eliminate remote measurements and determine the interval  in which is located the linear part of dependence of output measured quantity y = f(x) with normal distribution N(f(x), σ2) on the input, i.e., independent variable x. For the procedure used for testing of remoteness of experimental points, a relation has been derived for calculation of the critical value of deviation of the point tested. The algorithm is finished by the calculation of parameters of the corresponding regression straight line and other statistical characteristics. On the basis of the algorithm suggested, a program has been assembled whose reliability was verified on a series of both model and practical examples.
New Perspectives for the Treatment of HIV Infections
Authors: De Clercq Erik
Published: July 2002

The HIV replicative cycle reveals several virus-specific events that could function as targets for chemotherapeutic intervention. The compounds that are presently available as anti-HIV drugs are targeted at either the substrate binding site of the reverse transcriptase (zidovudine, didanosine, zalcitabine, stavudine, lamivudine) or a non-substrate binding site of the reverse transcriptase (nevirapine, delavirdine), or the viral protease (saquinavir, ritonavir, indinavir, nelfinavir). Remarkable clinical efficacy has been observed with combinations of different reverse transcriptase inhibitors and protease inhibitors. It may be anticipated that with the advent of newer and more efficient compounds the effectiveness of HIV inhibition could still be improved upon and the prospects for a definitive cure of the disease may be accomplished. An account with 107 references.
Modelling of DNA Complexes with Distamycin Analogues Using an ab initio Continuum Solvent Model
Authors: Bouř Petr , Král Vladimír
Published: July 2002

Model systems related to non-covalent minor groove DNA complexes with distamycin analogues were investigated using the Turbomole and Gaussian quantum chemical packages. The role of molecular shape, electrostatic field and conformer energies in the complex formation was discussed. The ab initio calculations included the COSMO solvent model. If compared to vacuum computations, polar solvent significantly destabilizes such complexes and increases conformational flexibility of distamycin. The DNA complex formation appears to be driven mainly by entropy lowering and complementarity of molecular shapes. The NH moiety of the amide group preferably points to the base pair according to the computations, in agreement with experimental data.
Synthesis and Extraction Properties of 1,2-Bis(amidoxime) Derivatives
Authors: Gup Ramazan , Alpoguz H. Korkmaz , Beduk A. Dincer
Published: July 2002

Four 1,2-bis(amidoxime)s bearing benzoate groups were synthesized. Their complexing properties were studied by the liquid-liquid extraction of selected alkali (Na+ and K+) and transition metals (Hg2+, Cu2+, Pb2+ and Ni2+). It has been observed that all ligands show a high affinity to Cu2+ and Hg2+ ions, whereas almost no affinity to alkali metals.
Hyperammonaemia in critically ill septic infants
Authors: McEwan P , Simpson D , Kirk J M , Barr D G D , McKenzie K J
Published: July 2002

Three infants with subphrenic abscess, pyonephrosis, and obstructive ureterocoele respectively had grossly increased concentrations of plasma ammonia. This was considered to be a result of infections with urea splitting organisms. All died in spite of intensive care support, including specific measures to reduce plasma ammonia.
The slow component of V̇o2in professional cyclists
Authors: Lucía Alejandro , Hoyos Jesús , Chicharro José L
Published: July 2002

Objectives—To analyse the slow component of oxygen uptake (V̇o2) in professional cyclists and to determine whether this phenomenon is due to altered neuromuscular activity, as assessed by surface electromyography (EMG).

Methods—The following variables were measured during 20 minute cycle ergometer tests performed at about 80% of V̇o2maxin nine professional road cyclists (mean (SD) age 26 (2) years; V̇o2max 72.6 (2.2) ml/kg/min): heart rate (HR), gas exchange variables (V̇o2, ventilation (V̇e), tidal volume (VT), breathing frequency (fb), ventilatory equivalents for oxygen and carbon dioxide (V̇e/V̇o2and V̇e/V̇co2respectively), respiratory exchange ratio (RER), and end tidal Po2and Pco2(Peto2and Petco2respectively)), blood variables (lactate, pH, and [HCO3−]) and EMG data (root mean from square voltage (rms-EMG) and mean power frequency (MPF)) from the vastus lateralis muscle.

Results—The mean magnitude of the slow component (from the end of the third minute to the end of exercise) was 130 (0.04) ml in 17 minutes or 7.6 ml/min. Significant increases from three minute to end of exercise values were found for the following variables: V̇o2(p<0.01), HR (p<0.01), V̇e(p<0.05), fb(p<0.01), V̇e/V̇o2(p<0.05), V̇e/V̇co2(p<0.01), Peto2(p<0.05), and blood lactate (p<0.05). In contrast, rms-EMG and MPF showed no change (p>0.05) throughout the exercise tests.

Conclusions—A significant but small V̇o2slow component was shown in professional cyclists during constant load heavy exercise. The results suggest that the primary origin of the slow component is not neuromuscular factors in these subjects, at least for exercise intensities up to 80% of V̇o2max.
Views of chartered physiotherapists on the psychological content of their practice: a preliminary study in the United Kingdom
Authors: Hemmings B , Povey L
Published: July 2002

Background: Although research into the psychological aspects of sports injury is increasing and psychological interventions have been identified as important in the rehabilitation process, few studies have focused on how sports medicine practitioners deal with psychological problems.

Objective: To investigate the perceptions of English chartered physiotherapists on the psychological content of their practice.

Methods: The Physiotherapist and Sport Psychology Questionnaire (PSPQ) was adapted slightly from the Athletic Trainer and Sport Psychology Questionnaire and used in this study. A survey package comprising a PSPQ, introductory letter, and self addressed envelope was mailed to 179 chartered physiotherapists registered in the England Eastern Region Sports Medicine Directory.

Results: In total, 90 (50% response rate) questionnaires were returned. The sample consisted of 67 women and 23 men with a mean (SD) age of 40.1 (5.4) years and 9.2 (3.1) years of experience as chartered physiotherapists. Descriptive statistical and qualitative analysis showed that physiotherapists believed athletes were often psychologically affected by injury. The physiotherapists also reported often using psychological techniques when treating injured athletes, but few reported having access to a sport psychologist for referral.

Conclusions: This research indicates that future physiotherapy education may need more emphasis on the psychological aspect of injury, and seek to increase knowledge on the potential of using psychological interventions within a physiotherapy rehabilitation programme. Furthermore, some form of referral network should be established between chartered physiotherapists and sport psychologists.
The morality of abortion and the deprivation of futures
Authors: Brown Mark T
Published: July 2002

In an influential essay entitled Why abortion is wrong, Donald Marquis argues that killing actual persons is wrong because it unjustly deprives victims of their future; that the fetus has a future similar in morally relevant respects to the future lost by competent adult homicide victims, and that, as consequence, abortion is justifiable only in the same circumstances in which killing competent adult human beings is justifiable.1The metaphysical claim implicit in the first premise, that actual persons have a future of value, is ambiguous. The Future Like Ours argument (FLO) would be valid if “future of value” were used consistently to mean either “potential future of value” or “self-represented future of value”, and FLO would be sound if one or the other interpretation supported both the moral claim and the metaphysical claim, but if, as I argue, any interpretation which makes the argument valid renders it unsound, then FLO must be rejected. Its apparent strength derives from equivocation on the concept of “a future of value”.
Asbestosis and lobar site of lung cancer: Table 1
Authors: Weiss William
Published: July 2002

OBJECTIVE

To assess the evidence for the hypothesis that lung cancer has a predilection for the lower lobes in workers with asbestosis.

METHOD

A review of the available literature with relevant information.

RESULTS

Six published reports were analysed. In four studies limited to series of cases with diagnoses of asbestosis, three showed lower lobe predominance of lung cancer whereas the fourth study included cases in which the radiographic readings did not meet the usual criterion of profusion for asbestosis. One cohort study showed lower lobe predominance; the other reported only 33% lower lobe cancers compared with 20% in unexposed controls.

CONCLUSION

There is some support for the hypothesis but more studies are needed.