bluuebunny/crossref_metadata_2025_split huggingface dataset:
By Inhibiting Replication, the Large Hepatitis Delta Antigen Can Indirectly Regulate Amber/W Editing and Its Own Expression
Authors: Sato Shuji , Cornillez-Ty Cromwell , Lazinski David W. 
Published: July 2004

ABSTRACT
Hepatitis delta virus (HDV) expresses two essential proteins with distinct functions. The small hepatitis delta antigen (HDAg-S) is expressed throughout replication and is needed to promote that process. The large form (HDAg-L) is farnesylated, is expressed only at later times via RNA editing of the amber/W site, and is required for virion assembly. When HDAg-L is artificially expressed at the onset of replication, it strongly inhibits replication. However, there is controversy concerning whether HDAg-L expressed naturally at later times as a consequence of editing and replication can similarly inhibit replication. Here, by stabilizing the predicted secondary structure downstream from the amber/W site, a replication-competent HDV mutant that exhibited levels of editing higher than those of the wild type was created. This mutant expressed elevated levels of HDAg-L early during replication, and at later times, its replication aborted prematurely. No further increase in amber/W editing was observed following the cessation of replication, indicating that editing was coupled to replication. A mutation in HDAg-L and a farnesyl transferase inhibitor were both used to abolish the ability of HDAg-L to inhibit replication. Such treatments rescued the replication defect of the overediting mutant, and even higher levels of amber/W editing resulted. It was concluded that when expressed naturally during replication, HDAg-L is able to inhibit replication and thereby inhibit amber/W editing and its own synthesis. In addition, the structure adjacent to the amber/W site is suboptimal for editing, and this creates a window of time in which replication can occur in the absence of HDAg-L.
Role of Nucleolin in Human Parainfluenza Virus Type 3 Infection of Human Lung Epithelial Cells
Authors: Bose Santanu , Basu Mausumi , Banerjee Amiya K. 
Published: July 2004

ABSTRACT
Human parainfluenza virus type 3 (HPIV-3) is an airborne pathogen that infects human lung epithelial cells from the apical (luminal) plasma membrane domain. In the present study, we have identified cell surface-expressed nucleolin as a cellular cofactor required for the efficient cellular entry of HPIV-3 into human lung epithelial A549 cells. Nucleolin was enriched on the apical cell surface domain of A549 cells, and HPIV-3 interacted with nucleolin during entry. The importance of nucleolin during HPIV-3 replication was borne out by the observation that HPIV-3 replication was significantly inhibited following (i) pretreatment of cells with antinucleolin antibodies and (ii) preincubation of HPIV-3 with purified nucleolin prior to its addition to the cells. Moreover, HPIV-3 cellular internalization and attachment assays performed in the presence of antinucleolin antibodies and purified nucleolin revealed the requirement of nucleolin during HPIV-3 internalization but not during attachment. Thus, these results suggest that nucleolin expressed on the surfaces of human lung epithelial A549 cells plays an important role during HPIV-3 cellular entry.
Increased Levels of Wee-1 Kinase in G
2
Are Necessary for Vpr- and Gamma Irradiation-Induced G
2
Arrest
Authors: Yuan Huidong , Kamata Masakazu , Xie Yi-Ming , Chen Irvin S. Y. 
Published: July 2004

ABSTRACT

Human immunodeficiency virus type 1 (HIV-1) Vpr induces cell cycle arrest at the G
2
/M transition and subsequently apoptosis. Here we examined the potential involvement of Wee-1 in Vpr-induced G
2
arrest. Wee-1 is a cellular protein kinase that inhibits Cdc2 activity, thereby preventing cells from proceeding through mitosis. We previously showed that the levels of Wee-1 correlate with Vpr-mediated apoptosis. Here, we demonstrate that Vpr-induced G
2
arrest correlated with delayed degradation of Wee-1 at G
2
/M. Experimental depletion of Wee-1 by a small interfering RNA directed to
wee-1
mRNA alleviated Vpr-induced G
2
arrest and allowed apparently normal progression through M into G
1
. Similar results were observed when cells were arrested at G
2
following gamma irradiation. Thus, Wee-1 is integrally involved as a key cellular regulatory protein in the signal transduction pathway for HIV-1 Vpr-induced cell cycle arrest.
Analysis of Splice Variants of the Immediate-Early 1 Region of Human Cytomegalovirus
Authors: Awasthi Sita , Isler Jennifer A. , Alwine James C. 
Published: July 2004

ABSTRACT
The major immediate-early (MIE) gene of human cytomegalovirus (HCMV) produces multiple mRNAs through differential splicing and polyadenylation. Reverse transcriptase PCR was used to characterize transcripts from exons 1, 2, 3, and 4 (immediate-early 1 [IE1]). The expected IE72 and IE19 mRNAs were detected, as well as two heretofore-uncharacterized transcripts designated IE17.5 and IE9. The IE72, IE19, and IE17.5 transcripts utilized the same 5′-splice site in exon 3. IE9 utilized a cryptic 5′-splice site within exon 3. The IE19, IE17.5, and IE9 transcripts all used different 3′-splice sites within exon 4. These spliced species occur in infected human foreskin fibroblast (HFF) cells, with accumulation kinetics similar to those of IE72 mRNA. IE19 and IE9 RNAs were much more abundant than IE17.5 RNA. Transfection of CV-1 cells with cDNAs resulted in IE19 and IE17.5 proteins detectable by antibodies to either N-terminal or C-terminal epitopes. No IE9 protein product has been detected. We have not been able to detect IE19, IE17.5, or IE9 proteins during infection of HFF, HEL, or U373MG cells. Failure to detect IE19 protein contrasts with a previous report (M. Shirakata, M. Terauchi, M. Ablikin, K. Imadome, K. Hirai, T. Aso, and Y. Yamanashi, J. Virol. 76:3158-3167, 2002) of IE19 protein expression in HCMV-infected HEL cells. Our analysis suggests that an N-terminal breakdown product of IE72 may be mistaken for IE19. Expression of IE19 or IE17.5 from its respective cDNA results in repression of viral gene expression in infected cells. We speculate that expression of these proteins during infection may be restricted to specific conditions or cell types.
Oral Immunization with RecombinantListeria monocytogenesControls Virus Load after Vaginal Challenge with Feline Immunodeficiency Virus
Authors: Stevens Rosemary , Howard Kristina E. , Nordone Sushila , Burkhard MaryJo , Dean Gregg A. 
Published: July 2004

ABSTRACTRecombinantListeria monocytogeneshas many attractive characteristics as a vaccine vector against human immunodeficiency virus (HIV). Wild-type and attenuatedListeriastrains expressing HIV Gag have been shown to induce long-lived mucosal and systemic T-cell responses in mice. Using the feline immunodeficiency virus (FIV) model of HIV we evaluated recombinantL. monocytogenesin a challenge system. Five cats were immunized with recombinantL. monocytogenesthat expresses the FIV Gag and delivers an FIV Env-expressing DNA vaccine (LMgag/pND14-Lc-env). Control cats were either sham immunized or immunized with wild-typeL. monocytogenes(LM-wt). At 1 year after vaginal challenge, provirus could not be detected in any of the nine tissues evaluated from cats immunized with the recombinant bacteria but was detected in at least one tissue in 8 of 10 control animals. Virus was isolated from bone marrow of four of five LMgag/pND14-Lc-env-immunized cats by use of a stringent coculture system but required CD8+T-cell depletion, indicating CD8+T-cell suppression of virus replication. Control animals had an inverted CD4:CD8 ratio in mesenteric lymph node and were depleted of both CD4+and CD8+intestinal epithelial T cells, while LMgag/pND14-Lc-env-immunized animals showed no such abnormalities. Vaginal FIV-specific immunoglobulin A was present at high titer in three LMgag/pND14-Lc-env-immunized cats before challenge and in all five at 1 year postchallenge. This study demonstrates that recombinantL. monocytogenesconferred some control of viral load after vaginal challenge with FIV.
p53 Targets Simian Virus 40 Large T Antigen for Acetylation by CBP
Authors: Poulin Danielle L. , Kung Andrew L. , DeCaprio James A. 
Published: July 2004

ABSTRACT
Simian virus 40 (SV40) large T antigen (T Ag) interacts with the tumor suppressor p53 and the transcriptional coactivators CBP and p300. Binding of these cellular proteins in a ternary complex has been implicated in T Ag-mediated transformation. It has been suggested that the ability of CBP/p300 to modulate p53 function underlies p53's regulation of cell proliferation and tumorigenesis. In this study, we provide further evidence that CBP activity may be mediated through its synergistic action with p53. We demonstrate that SV40 T Ag is acetylated in vivo in a p53-dependent manner and T Ag acetylation is largely mediated by CBP. The acetylation of T Ag is dependent on its interaction with p53 and on p53's interaction with CBP. We have mapped the site of acetylation on T Ag to the C-terminal lysine residue 697. This acetylation site is conserved between the T antigens of the human polyomaviruses JC and BK, which are also known to interact with p53. We show that both JC and BK T antigens are also acetylated at corresponding sites in vivo. While other proteins are known to be acetylated by CBP/p300, none are known to depend on p53 for acetylation. T Ag acetylation may provide a regulatory mechanism for T Ag binding to a cellular factor or play a role in another aspect of T Ag function.
Common E Protein Determinants for Attenuation of Glycosaminoglycan-Binding Variants of Japanese Encephalitis and West Nile Viruses
Authors: Lee Eva , Hall Roy A. , Lobigs Mario 
Published: July 2004

ABSTRACT
Natural isolates and laboratory strains of West Nile virus (WNV) and Japanese encephalitis virus (JEV) were attenuated for neuroinvasiveness in mouse models for flavivirus encephalitis by serial passage in human adenocarcinoma (SW13) cells. The passage variants displayed a small-plaque phenotype, augmented affinity for heparin-Sepharose, and a marked increase in specific infectivity for SW13 cells relative to the respective parental viruses, while the specific infectivity for Vero cells was not altered. Therefore, host cell adaptation of passage variants was most likely a consequence of altered receptor usage for virus attachment-entry with the involvement of cell surface glycosaminoglycans (GAG) in this process. In vivo blood clearance kinetics of the passage variants was markedly faster and viremia was reduced relative to the parental viruses, suggesting that affinity for GAG (ubiquitously present on cell surfaces and extracellular matrices) is a key determinant for the neuroinvasiveness of encephalitic flaviviruses. A difference in pathogenesis between WNV and JEV, which was reflected in more efficient growth in the spleen and liver of the WNV parent and passage variants, accounted for a less pronounced loss of neuroinvasiveness of GAG binding variants of WNV than JEV. Single gain-of-net-positive-charge amino acid changes at E protein residue 49, 138, 306, or 389/390, putatively positioned in two clusters on the virion surface, define molecular determinants for GAG binding and concomitant virulence attenuation that are shared by the JEV serotype flaviviruses.
Trimeric Hantavirus Nucleocapsid Protein Binds Specifically to the Viral RNA Panhandle
Authors: Mir M. A. , Panganiban A. T. 
Published: July 2004

ABSTRACT

Hantaviruses are tripartite negative-sense RNA viruses and members of the
Bunyaviridae
family. The nucleocapsid (N) protein is encoded by the smallest of the three genome segments (S). N protein is the principal structural component of the viral capsid and is central to the hantavirus replication cycle. We examined intermolecular N-protein interaction and RNA binding by using bacterially expressed Sin Nombre virus N protein. N assembles into di- and trimeric forms. The mono- and dimeric forms exist transiently and assemble into a trimeric form. In contrast, the trimer is highly stable and does not efficiently disassemble into the mono- and dimeric forms. The purified N-protein trimer is able to discriminate between viral and nonviral RNA molecules and, interestingly, recognizes and binds with high affinity the panhandle structure composed of the 3′ and 5′ ends of the genomic RNA. In contrast, the mono- and dimeric forms of N bind RNA to form a complex that is semispecific and salt sensitive. We suggest that trimerization of N protein is a molecular switch to generate a protein complex that can discriminate between viral and nonviral RNA molecules during the early steps of the encapsidation process.
Itinerary of Hepatitis B Viruses: Delineation of Restriction Points Critical for Infectious Entry
Authors: Funk Anneke , Mhamdi Mouna , Lin Li , Will Hans , Sirma Hüseyin 
Published: July 2004

ABSTRACT

Little is known about cellular determinants essential for human hepatitis B virus infection. Using the duck hepatitis B virus as a model, we first established a sensitive binding assay for both virions and subviral particles and subsequently elucidated the characteristics of the early viral entry steps. The infection itinerary was found to initiate with the attachment of viral particles to a low number of binding sites on hepatocytes (about 10
4
per cell). Virus internalization was fully accomplished in less than 3 h but was then followed by a period of unprecedented length, about 14 h, until completion of nuclear import of the viral genome. Steps subsequent to virus entry depended on both intact microtubules and their dynamic turnover but not on actin cytoskeleton. Notably, cytoplasmic trafficking of viral particles and emergence of nuclear covalently closed circular DNA requires microtubules during entry only at and for specific time periods. Taken together, these data disclose for the first time a series of steps and their kinetics that are essential for the entry of hepatitis B viruses into hepatocytes and are different from those of any other virus reported so far.
Coiled-Coil Domains in Glycoproteins B and H Are Involved in Human Cytomegalovirus Membrane Fusion
Authors: Lopper Matthew , Compton Teresa 
Published: July 2004

ABSTRACT
Human cytomegalovirus (CMV) utilizes a complex route of entry into cells that involves multiple interactions between viral envelope proteins and cellular receptors. Three conserved viral glycoproteins, gB, gH, and gL, are required for CMV-mediated membrane fusion, but little is known of how these proteins cooperate during entry (E. R. Kinzler and T. Compton, submitted for publication). The goal of this study was to begin defining the molecular mechanisms that underlie membrane fusion mediated by herpesviruses. We identified heptad repeat sequences predicted to form alpha-helical coiled coils in two glycoproteins required for fusion, gB and gH. Peptides derived from gB and gH containing the heptad repeat sequences inhibited virus entry when introduced coincident with virus inoculation onto cells or when mixed with virus prior to inoculation. Neither peptide affected binding of CMV to fibroblasts, suggesting that the peptides inhibit membrane fusion. Both gB and gH coiled-coil peptides blocked entry of several laboratory-adapted and clinical strains of human CMV, but neither peptide affected entry of murine CMV or herpes simplex virus type 1 (HSV-1). Although murine CMV and HSV-1 gB and gH have heptad repeat regions, the ability of human CMV gB and gH peptides to inhibit virus entry correlates with the specific residues that comprise the heptad repeat region. The ability of gB and gH coiled-coil peptides to inhibit virus entry independently of cell contact suggests that the coiled-coil regions of gB and gH function differently from those of class I, single-component fusion proteins. Taken together, these data support a critical role for alpha-helical coiled coils in gB and gH in the entry pathway of CMV.
Effect of Vaccine Use in the Evolution of Mexican Lineage H5N2 Avian Influenza Virus
Authors: Lee Chang-Won , Senne Dennis A. , Suarez David L. 
Published: July 2004

ABSTRACT
An outbreak of avian influenza (AI) caused by a low-pathogenic H5N2 type A influenza virus began in Mexico in 1993 and several highly pathogenic strains of the virus emerged in 1994-1995. The highly pathogenic virus has not been reported since 1996, but the low-pathogenic virus remains endemic in Mexico and has spread to two adjacent countries, Guatemala and El Salvador. Measures implemented to control the outbreak and eradicate the virus in Mexico have included a widespread vaccination program in effect since 1995. Because this is the first case of long-term use of AI vaccines in poultry, the Mexican lineage virus presented us with a unique opportunity to examine the evolution of type A influenza virus circulating in poultry populations where there was elevated herd immunity due to maternal and active immunity. We analyzed the coding sequence of the HA1 subunit and the NS gene of 52 Mexican lineage viruses that were isolated between 1993 and 2002. Phylogenetic analysis indicated the presence of multiple sublineages of Mexican lineage isolates at the time vaccine was introduced. Further, most of the viruses isolated after the introduction of vaccine belonged to sublineages separate from the vaccine's sublineage. Serologic analysis using hemagglutination inhibition and virus neutralization tests showed major antigenic differences among isolates belonging to the different sublineages. Vaccine protection studies further confirmed the in vitro serologic results indicating that commercial vaccine was not able to prevent virus shedding when chickens were challenged with antigenically different isolates. These findings indicate that multilineage antigenic drift, which has not been observed in AI virus, is occurring in the Mexican lineage AI viruses and the persistence of the virus in the field is likely aided by its large antigenic difference from the vaccine strain.
Human Cytomegalovirus-Encoded G Protein-Coupled Receptor US28 Mediates Smooth Muscle Cell Migration through Gα12
Authors: Melnychuk Ryan M. , Streblow Daniel N. , Smith Patricia P. , Hirsch Alec J. , Pancheva Dora , Nelson Jay A. 
Published: July 2004

ABSTRACT
Coupling of G proteins to ligand-engaged chemokine receptors is the paramount event in G-protein-coupled receptor signal transduction. Previously, we have demonstrated that the human cytomegalovirus-encoded chemokine receptor US28 mediates human vascular smooth muscle cell (SMC) migration in response to either RANTES or monocyte chemoattractant protein 1. In this report, we identify the G proteins that couple with US28 to promote vascular SMC migration and identify other signaling molecules that play critical roles in this process. US28-mediated cellular migration was enhanced with the expression of the G-protein subunits Gα12 and Gα13, suggesting that US28 may functionally couple to these G proteins. In correlation with this observation, US28 was able to activate RhoA, a downstream effector of Gα12 and Gα13 in cell types with these G proteins but not in those without them and activation of RhoA was dependent on US28 stimulation with RANTES. In addition, inactivation of RhoA or the RhoA-associated kinase p160ROCK with a dominant-negative mutant of RhoA or the small molecule inhibitor Y27632, respectively, abrogated US28-induced SMC migration. The data presented here suggest that US28 functionally signals through Gα12 family G proteins and RhoA in a ligand-dependent manner and these signaling molecules are important for the ability of US28 to induce cellular migration.
Is virtual bronchoscopy useful for physicians practising in a district general hospital?
Authors: Dheda K , Roberts C M , Partridge M R , Mootoosamy I 
Published: July 2004

Abstract
Background: Virtual bronchoscopy software is now available to district general hospitals (DGHs). There is limited information on the clinical utility of virtual bronchoscopy and whether it offers any additional information over conventional axial computed tomography in the setting of a busy DGH chest unit.
Methods: Virtual bronchoscopy and computed tomography findings were compared in all patients who had a virtual bronchoscopy study over a 12 month period.
Results: Eighteen consecutive patients had virtual bronchoscopy for a specific clinical indication over the study period. Additional information was conveyed by virtual bronchoscopy in five patients (in four patients the airways distal to an obstruction were better visualised thereby influencing decisions about airway stenting and in one patient the virtual bronchoscopy study showed an endobronchial lesion missed on computed tomography). In nine patients who were unfit for fibreoptic bronchoscopy (FOB) the radiologist was more confident in excluding an obstructive airway lesion. The main indication for performing a virtual bronchoscopy study was to rule out an obstructive airway lesion in patients who were unfit for FOB (n = 11).
Conclusion: Virtual bronchoscopy is feasible and useful in the management of a few selected patients in a DGH chest unit. Virtual bronchoscopy may convey additional information over computed tomography when the distal airways need to be visualised and for discrete endoluminal lesions.
Iron deficiency anaemia in general practice: clinical outcomes over three years and factors influencing diagnostic investigations
Authors: Yates J M , Logan E C M , Stewart R M 
Published: July 2004

Abstract
Background: Iron deficiency anaemia (IDA) may be a sign of significant gastrointestinal disease, and delayed diagnosis may result in chronic morbidity. Studies in patients referred to hospital for investigation of their anaemia have shown that 5%–15% have a gastrointestinal cancer but there are few studies of patients presenting to primary care. Factors influencing further investigation in these patients have not previously been identified.
Patients and methods: A cohort of patients presenting to their general practitioners (GPs) with IDA was identified and clinical outcomes recorded. Logistic regression was used to determine which factors influenced GPs to investigate the anaemia.
Results: 43% of patients had investigations within three months and serious pathology was found in 30% of these; 13% of patients were considered unfit for further investigation and 8% refused to have any. Independent predictors of non-investigation were a mild anaemia (odds ratio (OR) 0.38, confidence interval (CI) 0.23 to 0.61, p&lt;0.001), female gender (OR 0.49, CI 0.3 to 0.8, p = 0.004), a previous history of anaemia (OR 0.39, CI 0.24 to 0.64, p&lt;0.001), and age &lt;65 years (OR 0.44, CI 0.26 to 0.74, p = 0.002). During the entire study period gastrointestinal cancer was diagnosed in 48 patients (11%); 17% of men had colorectal cancer. Of 263 patients alive at 12 months without a confirmed diagnosis, 113 (43%) had recurrent or persistent anaemia during the study period.
Conclusion: Although the overall prevalence of gastrointestinal cancer in patients presenting to primary care is similar to that seen in secondary care, the diagnosis may be delayed due to lack of appropriate investigations resulting in significant morbidity.