bluuebunny/crossref_metadata_2025 huggingface dataset:
A Conserved Gly436-Trp-Leu-Ala-Gly-Leu-Phe-Tyr Motif in Hepatitis C Virus Glycoprotein E2 Is a Determinant of CD81 Binding and Viral Entry
Authors: Drummer Heidi E. , Boo Irene , Maerz Anne L. , Poumbourios Pantelis
Published: July 2006

The hepatitis C virus (HCV) glycoproteins E1 and E2 form a heterodimer that mediates CD81 receptor binding and viral entry. In this study, we used site-directed mutagenesis to examine the functional role of a conserved G436WLAGLFY motif of E2. The mutants could be placed into two groups based on the ability of mature virion-incorporated E1E2 to bind the large extracellular loop (LEL) of CD81 versus the ability to mediate cellular entry of pseudotyped retroviral particles. Group 1 comprised E2 mutants where LEL binding ability largely correlated with viral entry ability, with conservative and nonconservative substitutions (W437 L/A, L438A, L441V/F, and F442A) inhibiting both functions. These data suggest that Trp-437, Leu-438, Leu-441, and Phe-442 directly interact with the LEL. Group 2 comprised E2 glycoproteins with more conservative substitutions that lacked LEL binding but retained between 20% and 60% of wild-type viral entry competence. The viral entry competence displayed by group 2 mutants was explained by residual binding by the E2 receptor binding domain to cellular full-length CD81. A subset of mutants maintained LEL binding ability in the context of intracellular E1E2 forms, but this function was largely lost in virion-incorporated glycoproteins. These data suggest that the CD81 binding site undergoes a conformational transition during glycoprotein maturation through the secretory pathway. The G436P mutant was an outlier, retaining near-wild-type levels of CD81 binding but lacking significant viral entry ability. These findings indicate that the G436WLAGLFY motif of E2 functions in CD81 binding and in pre- or post-CD81-dependent stages of viral entry.
Compensatory Substitutions Restore Normal Core Assembly in Simian Immunodeficiency Virus Isolates with Gag Epitope Cytotoxic T-Lymphocyte Escape Mutations
Authors: Yeh Wendy W. , Cale Evan M. , Jaru-Ampornpan Pimkwan , Lord Carol I. , Peyerl Fred W. , Letvin Norman L.
Published: July 2006

The evolution of human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) as they replicate in infected individuals reflects a balance between the pressure on the virus to mutate away from recognition by dominant epitope-specific cytotoxic T lymphocytes (CTL) and the structural constraints on the virus' ability to mutate. To gain a further understanding of the strategies employed by these viruses to maintain replication competency in the face of the intense selection pressure exerted by CTL, we have examined the replication fitness and morphological ramifications of a dominant epitope mutation and associated flanking amino acid substitutions on the capsid protein (CA) of SIV/simian-human immunodeficiency virus (SHIV). We show that a residue 2 mutation in the immunodominant p11C, C-M epitope (T47I) of SIV/SHIV not only decreased CA protein expression and viral replication, but it also blocked CA assembly in vitro and virion core condensation in vivo. However, these defects were restored by the introduction of upstream I26V and/or downstream I71V substitutions in CA. These findings demonstrate how flanking compensatory amino acid substitutions can facilitate viral escape from a dominant epitope-specific CTL response through the effects of these associated mutations on the structural integrity of SIV/SHIV.
Psittacid Herpesvirus 1 and Infectious Laryngotracheitis Virus: Comparative Genome Sequence Analysis of Two Avian Alphaherpesviruses
Authors: Thureen Dean R. , Keeler Calvin L.
Published: July 2006

Psittacid herpesvirus 1 (PsHV-1) is the causative agent of Pacheco's disease, an acute, highly contagious, and potentially lethal respiratory herpesvirus infection in psittacine birds, while infectious laryngotracheitis virus (ILTV) is a highly contagious and economically significant avian herpesvirus which is responsible for an acute respiratory disease limited to galliform birds. The complete genome sequence of PsHV-1 has been determined and compared to the ILTV sequence, assembled from published data. The PsHV-1 and ILTV genomes exhibit similar structural characteristics and are 163,025 bp and 148,665 bp in length, respectively. The PsHV-1 genome contains 73 predicted open reading frames (ORFs), while the ILTV genome contains 77 predicted ORFs. Both genomes contain an inversion in the unique long region similar to that observed in pseudorabies virus. PsHV-1 is closely related to ILTV, and it is proposed that it be assigned to the
Iltovirus
genus. These two avian herpesviruses represent a phylogenetically unique clade of alphaherpesviruses that are distinct from the Marek's disease-like viruses (
Mardivirus
). The determination of the complete genomic nucleotide sequences of PsHV-1 and ILTV provides a tool for further comparative and functional analysis of this unique class of avian alphaherpesviruses.
A 38-Amino-Acid Sequence Encompassing the Arm Domain of the
Cucumber Necrosis Virus
Coat Protein Functions as a Chloroplast Transit Peptide in Infected Plants
Authors: Xiang Yu , Kakani Kishore , Reade Ron , Hui Elizabeth , Rochon D'Ann
Published: July 2006

Experiments to determine the subcellular location of the coat protein (CP) of the tombusvirus
Cucumber necrosis virus
(CNV) have been conducted. By confocal microscopy, it was found that an agroinfiltrated CNV CP-green fluorescent protein (GFP) fusion targets chloroplasts in
Nicotiana benthamiana
leaves and that a 38-amino-acid (aa) region that includes the complete CP arm region plus the first 4 amino acids of the shell domain are sufficient for targeting. Western blot analyses of purified and fractionated chloroplasts showed that the 38-aa region directs import to the chloroplast stroma, suggesting that the CNV arm can function as a chloroplast transit peptide (TP) in plants. Several features of the 38-aa region are similar to features typical of chloroplast TPs, including (i) the presence of an alanine-rich uncharged region near the N terminus, followed by a short region rich in basic amino acids; (ii) a conserved chloroplast TP phosphorylation motif; (iii) the requirement that the CNV 38-aa sequence be present at the amino terminus of the imported protein; and (iv) specific proteolytic cleavage upon import into the chloroplast stroma. In addition, a region just downstream of the 38-aa sequence contains a 14-3-3 binding motif, suggesting that chloroplast targeting requires 14-3-3 binding, as has been suggested for cellular proteins that are targeted to chloroplasts. Chloroplasts of CNV-infected plants were found to contain CNV CP, but only the shell and protruding domain regions were present, indicating that CNV CP enters chloroplasts during infection and that proteolytic cleavage occurs as predicted from agroinfiltration studies. We also found that particles of a CNV CP mutant deficient in externalization of the arm region have a reduced ability to establish infection. The potential biological significance of these findings is discussed.
Oncogenic Activity of Epstein-Barr Virus Latent Membrane Protein 1 (LMP-1) Is Down-Regulated by Lytic LMP-1
Authors: Pandya Jyotsna , Walling Dennis M.
Published: July 2006

The Epstein-Barr virus (EBV) is an oncogenic human herpesvirus. EBV latent membrane protein 1 (LMP-1) is a viral oncogene that manifests its oncogenic phenotype through activation of cellular signaling pathways involved in cell growth, survival, differentiation, and transformation. Lytic LMP-1 (lyLMP-1) is a related EBV gene without oncogenic properties. The lyLMP-1 gene is found in 60% of the EBV strains circulating in nature, but it is not found in EBV strains associated with nasopharyngeal carcinoma. We recently demonstrated that lyLMP-1 down-regulates the half-life of LMP-1 in epithelial cells. Therefore in this study, we tested the hypothesis that lyLMP-1 concomitantly down-regulates LMP-1 oncogenic activity. The results demonstrated that lyLMP-1 inhibits LMP-1-mediated intracellular signaling activation, epithelial cell growth and survival, and fibroblast cell transformation in a dose-dependent manner. Lytic LMP-1 manifested this effect through the promotion of LMP-1 degradation and a reduction in the expressed quantity of LMP-1. Thus, lyLMP-1 functions as a posttranslational negative regulator of LMP-1 oncogenesis. These results support a model of EBV-associated epithelial oncogenesis in which lyLMP-1 may act in vivo to reduce the risk of LMP-1-mediated transformation and is therefore subjected to negative selection in nasopharyngeal carcinoma pathogenesis.
Contribution of Vpu, Env, and Nef to CD4 Down-Modulation and Resistance of Human Immunodeficiency Virus Type 1-Infected T Cells to Superinfection
Authors: Wildum Steffen , Schindler Michael , Münch Jan , Kirchhoff Frank
Published: July 2006

Human immunodeficiency virus type 1 (HIV-1) utilizes Vpu, Env, and Nef to down-modulate its primary CD4 receptor from the cell surface, and this function seems to be critical for the pathogenesis of AIDS. The physiological relevance of CD4 down-modulation, however, is currently not well understood. In the present study, we analyzed the kinetics of CD4 down-modulation and the susceptibility of HIV-1-infected T cells to superinfection using proviral HIV-1 constructs containing individual and combined defects in
vpu
,
env
, and
nef
and expressing red or green fluorescent proteins. T cells infected with HIV-1 mutants containing functional
nef
genes expressed low surface levels of CD4 from the first moment that viral gene expression became detectable. In comparison, Vpu and Env had only minor to moderate effects on CD4 during later stages of infection. Consistent with these quantitative differences, Nef inhibited superinfection more efficiently than Vpu and Env. Notably,
nef
alleles from AIDS patients were more effective in preventing superinfection than those derived from a nonprogressor of HIV-1 infection. Our data suggest that protection against X4-tropic HIV-1 superinfection involves both CD4-independent and CD4-dependent mechanisms of HIV-1 Nef. X4 was effectively down-regulated by simian immunodeficiency virus and HIV-2 but not by HIV-1 Nef proteins. Thus, maximal protection seems to involve an as-yet-unknown mechanism that is independent of CD4 or coreceptor down-modulation. Finally, we demonstrate that superinfected primary T cells show enhanced levels of apoptosis. Accordingly, one reason that HIV-1 inhibits CD4 surface expression and superinfection is to prevent premature cell death in order to expand the period of effective virus production.
Epstein-Barr Virus Nuclear Antigen 2
trans
-Activates the Cellular Antiapoptotic
bfl-1
Gene by a CBF1/RBPJκ-Dependent Pathway
Authors: Pegman Pamela M. , Smith Sinéad M. , D'Souza Brendan N. , Loughran Sinéad T. , Maier Sabine , Kempkes Bettina , Cahill Paul A. , Simmons Matthew J. , Gélinas Céline , Walls Dermot
Published: July 2006

The human herpesvirus Epstein-Barr virus (EBV) establishes latency and
promotes the long-term survival of its host B cell by targeting the
molecular machinery controlling cell fate decisions. The cellular
antiapoptotic
bfl-1
gene confers protection from apoptosis
under conditions of growth factor deprivation when expressed
ectopically in an EBV-negative Burkitt's lymphoma-derived cell line (B.
D'Souza, M. Rowe, and D. Walls, J. Virol.
74:6652-6658, 2000), and the EBV latent membrane protein 1
(LMP1) and its cellular functional homologue CD40 can both drive
bfl-1
via an NF-κB-dependent enhancer element in the
bfl-1
promoter (B. N. D'Souza, L. C.
Edelstein, P. M. Pegman, S. M. Smith, S.
T. Loughran, A. Clarke, A. Mehl, M. Rowe, C. Gélinas, and D.
Walls, J. Virol. 78:1800-1816, 2004). Here we show
that the EBV nuclear antigen 2 (EBNA2) also upregulates
bfl-1
.
EBNA2
trans
-activation of
bfl-1
requires CBF1 (or
RBP-Jκ), a nuclear component of the Notch signaling pathway,
and there is an essential role for a core consensus CBF1-binding site
on the
bfl-1
promoter.
trans
-activation is dependent
on the EBNA2-CBF1 interaction, is modulated by other EBV gene products
known to interact with the CBF1 corepressor complex, and does not
involve activation of NF-κB.
bfl-1
expression is
induced and maintained at high levels by the EBV growth program in a
lymphoblastoid cell line, and withdrawal of either EBNA2 or LMP1 does
not lead to a reduction in
bfl-1
mRNA levels in this context,
whereas the simultaneous loss of both EBV proteins results in a major
decrease in
bfl-1
expression. These findings are relevant to
our understanding of EBV persistence, its role in malignant disease,
and the B-cell developmental
process.
Isolation and Characterization of Monoclonal Antibodies Which Neutralize Human Metapneumovirus In Vitro and In Vivo
Authors: Ulbrandt Nancy D. , Ji Hong , Patel Nita K. , Riggs Jeffrey M. , Brewah Yambasu A. , Ready Shannon , Donacki Nanci E. , Folliot Karyn , Barnes Arnita S. , Senthil Kannaki , Wilson Susan , Chen Mingzhong , Clarke Lori , MacPhail Mia , Li Jia , Woods Robert M. , Coelingh Kathy , Reed Jennifer L. , McCarthy Michael P. , Pfarr David S. , Osterhaus Albert D. M. E. , Fouchier Ron A. M. , Kiener Peter A. , Suzich JoAnn A.
Published: July 2006

Human metapneumovirus (hMPV) is a recently described member of the
Paramyxoviridae
family/
Pneumovirinae
subfamily and
shares many common features with respiratory syncytial virus (RSV),
another member of the same subfamily. hMPV causes respiratory tract
illnesses that, similar to human RSV, occur predominantly during the
winter months and have symptoms that range from mild to severe cough,
bronchiolitis, and pneumonia. Like RSV, the hMPV virus can be
subdivided into two genetic subgroups, A and B. With RSV, a single
monoclonal antibody directed at the fusion (F) protein can prevent
severe lower respiratory tract RSV infection. Because of the high level
of sequence conservation of the F protein across all the hMPV
subgroups, this protein is likely to be the preferred antigenic target
for the generation of cross-subgroup neutralizing antibodies. Here we
describe the generation of a panel of neutralizing monoclonal
antibodies that bind to the hMPV F protein. A subset of these
antibodies has the ability to neutralize prototypic strains of both the
A and B hMPV subgroups in vitro. Two of these antibodies exhibited
high-affinity binding to the F protein and were shown to protect
hamsters against infection with hMPV. The data suggest that a
monoclonal antibody could be used prophylactically to prevent lower
respiratory tract disease caused by
hMPV.
Viral Determinants of Integration Site Preferences of Simian Immunodeficiency Virus-Based Vectors
Authors: Monse Hella , Laufs Stephanie , Kuate Seraphin , Zeller W. Jens , Fruehauf Stefan , Überla Klaus
Published: July 2006

Preferential integration into transcriptionally active regions of genomes has been observed for retroviral vectors based on gamma-retroviruses and lentiviruses. However, differences in the integration site preferences were detected, which might be explained by differences in viral components of the preintegration complexes. Viral determinants of integration site preferences have not been defined. Therefore, integration sites of simian immunodeficiency virus (SIV)-based vectors produced in the absence of accessory genes or lacking promoter and enhancer elements were compared. Similar integration patterns for the different SIV vectors indicate that
vif
,
vpr
,
vpx
,
nef
,
env
, and promoter or enhancer elements are not required for preferential integration of SIV into transcriptionally active regions of genomes.
Altered Gag Polyprotein Cleavage Specificity of Feline Immunodeficiency Virus/Human Immunodeficiency Virus Mutant Proteases as Demonstrated in a Cell-Based Expression System
Authors: Lin Ying-Chuan , Brik Ashraf , de Parseval Aymeric , Tam Karen , Torbett Bruce E. , Wong Chi-Huey , Elder John H.
Published: July 2006

We have used feline immunodeficiency virus (FIV) protease (PR) as a mutational system to study the molecular basis of substrate-inhibitor specificity for lentivirus PRs, with a focus on human immunodeficiency virus type 1 (HIV-1) PR. Our previous mutagenesis studies demonstrated that discrete substitutions in the active site of FIV PR with structurally equivalent residues of HIV-1 PR dramatically altered the specificity of the mutant PRs in in vitro analyses. Here, we have expanded these studies to analyze the specificity changes in each mutant FIV PR expressed in the context of the natural Gag-Pol polyprotein ex vivo. Expression mutants were prepared in which 4 to 12 HIV-1-equivalent substitutions were made in FIV PR, and cleavage of each Gag-Pol polyprotein was then assessed in pseudovirions from transduced cells. The findings demonstrated that, as with in vitro analyses, inhibitor specificities of the mutants showed increased HIV-1 PR character when analyzed against the natural substrate. In addition, all of the mutant PRs still processed the FIV polyprotein but the apparent order of processing was altered relative to that observed with wild-type FIV PR. Given the importance of the order in which Gag-Pol is processed, these findings likely explain the failure to produce infectious FIVs bearing these mutations.
Influenza A Virus PB1-F2 Protein Contributes to Viral Pathogenesis in Mice
Authors: Zamarin Dmitriy , Ortigoza Mila B. , Palese Peter
Published: July 2006

The influenza virus PB1-F2 protein is a novel protein previously shown to be involved in induction of cell death. Here we characterize the expression and the function of the protein within the context of influenza viral infection in tissue culture and a mouse model. We show that the C-terminal region of the protein can be expressed from a downstream initiation codon and is capable of interaction with the full-length protein. Using this knowledge, we generated influenza viruses knocked out for the expression of PB1-F2 protein and its downstream truncation products. Knocking out the PB1-F2 protein had no effect on viral replication in tissue culture but diminished virus pathogenicity and mortality in mice. The viruses replicated to similar levels in mouse lungs by day 3 postinfection, suggesting that the knockout did not impair viral replication. However, while the PB1-F2 knockout viruses were cleared after day 5, the wild-type viruses were detectable in mouse lungs until day 7, implying that expression of PB1-F2 resulted in delayed clearance of the viruses by the host immune system. Based on our findings and on the fact that the PB1 genomic segment was always newly introduced into some pandemic influenza viruses of the last century, we speculate that the PB1-F2 protein plays an important role in pathogenesis of influenza virus infection and may be an important contributor to pathogenicity of pandemic influenza viruses.
Characterization of the Prototype Foamy Virus Envelope Glycoprotein Receptor-Binding Domain
Authors: Duda Anja , Lüftenegger Daniel , Pietschmann Thomas , Lindemann Dirk
Published: July 2006

The
foamy virus (FV) glycoprotein precursor gp130
Env
undergoes a
highly unusual biosynthesis, resulting in the generation of three
particle-associated, mature subunits, leader peptide (LP), surface
(SU), and transmembrane (TM). Little structural and functional
information on the extracellular domains of FV Env is available. In
this study, we characterized the prototype FV (PFV) Env
receptor-binding domain (RBD) by flow cytometric analysis of
recombinant PFV Env immunoadhesin binding to target cells. The
extracellular domains of the C-terminal TM subunit as well as targeting
of the recombinant immunoadhesins by the cognate LP to the secretory
pathway were dispensable for target cell binding, suggesting that the
PFV Env RBD is contained within the SU subunit. N- and C-terminal
deletion analysis of the SU domain revealed a minimal continuous RBD
spanning amino acids (aa) 225 to 555; however, internal deletions
covering the region from aa 397 to 483, but not aa 262 to 300 or aa 342
to 396, were tolerated without significant influence on host cell
binding. Analysis of individual cysteine point mutants in PFV SU
revealed that only most of those located in the nonessential region
from aa 397 to 483 retained residual binding activity. Interestingly,
analysis of various N-glycosylation site mutants suggests an important
role of carbohydrate chain attachment to N
391
, either for
direct interaction with the receptor or for correct folding of the PFV
Env RBD. Taken together, these results suggest that a bipartite
sequence motif spanning aa 225 to 396 and aa 484 to 555 is essential
for formation of the PFV Env RBD, with N-glycosylation site at position
391 playing a crucial role for host cell
binding.
Crystal Structure of Nonstructural Protein 10 from the Severe Acute Respiratory Syndrome Coronavirus Reveals a Novel Fold with Two Zinc-Binding Motifs
Authors: Joseph Jeremiah S. , Saikatendu Kumar Singh , Subramanian Vanitha , Neuman Benjamin W. , Brooun Alexei , Griffith Mark , Moy Kin , Yadav Maneesh K. , Velasquez Jeffrey , Buchmeier Michael J. , Stevens Raymond C. , Kuhn Peter
Published: July 2006

The severe acute respiratory syndrome coronavirus (SARS-CoV) possesses a large 29.7-kb positive-stranded RNA genome. The first open reading frame encodes replicase polyproteins 1a and 1ab, which are cleaved to generate 16 “nonstructural” proteins, nsp1 to nsp16, involved in viral replication and/or RNA processing. Among these, nsp10 plays a critical role in minus-strand RNA synthesis in a related coronavirus, murine hepatitis virus. Here, we report the crystal structure of SARS-CoV nsp10 at a resolution of 1.8 Å as determined by single-wavelength anomalous dispersion using phases derived from hexatantalum dodecabromide. nsp10 is a single domain protein consisting of a pair of antiparallel N-terminal helices stacked against an irregular β-sheet, a coil-rich C terminus, and two Zn fingers. nsp10 represents a novel fold and is the first structural representative of this family of Zn finger proteins found so far exclusively in coronaviruses. The first Zn finger coordinates a Zn
2+
ion in a unique conformation. The second Zn finger, with four cysteines, is a distant member of the “gag-knuckle fold group” of Zn
2+
-binding domains and appears to maintain the structural integrity of the C-terminal tail. A distinct clustering of basic residues on the protein surface suggests a nucleic acid-binding function. Gel shift assays indicate that in isolation, nsp10 binds single- and double-stranded RNA and DNA with high-micromolar affinity and without obvious sequence specificity. It is possible that nsp10 functions within a larger RNA-binding protein complex. However, its exact role within the replicase complex is still not clear.
Dodecamer Structure of Severe Acute Respiratory Syndrome Coronavirus Nonstructural Protein nsp10
Authors: Su Dan , Lou Zhiyong , Sun Fei , Zhai Yujia , Yang Haitao , Zhang Rongguang , Joachimiak Andrzej , Zhang Xuejun C. , Bartlam Mark , Rao Zihe
Published: July 2006

The severe acute respiratory syndrome coronavirus (SARS-CoV) nonstructural proteins nsp1 to nsp16 have been implicated by genetic analysis in the assembly of a functional replication/transcription complex. We report the crystal structure of nsp10 from SARS-CoV at 2.1-Å resolution. The nsp10 structure has a novel fold, and 12 identical subunits assemble to form a unique spherical dodecameric architecture. Two zinc fingers have been identified from the nsp10 monomer structure with the sequence motifs C-(X)
2
-C-(X)
5
-H-(X)
6
-C and C-(X)
2
-C-(X)
7
-C-(X)-C. The nsp10 crystal structure is the first of a new class of zinc finger protein three-dimensional structures to be revealed experimentally. The zinc finger sequence motifs are conserved among all three coronavirus antigenic groups, implicating an essential function for nsp10 in all coronaviruses. Based on the structure, we propose that nsp10 is a transcription factor for coronavirus replication/transcription.
Relationships between Hepatitis C Virus Replication and CXCL-8 Production In Vitro
Authors: Koo Bon Chang A. , McPoland Paula , Wagoner Jessica P. , Kane Olivia J. , Lohmann Volker , Polyak Stephen J.
Published: July 2006

The chemokine CXCL-8 (interleukin-8) is induced by many viruses, including hepatitis C virus (HCV). In the current study, we examined CXCL-8 levels in the context of acute and chronic HCV replication in vitro. Two different small interfering RNAs were used to silence CXCL-8 mRNA and protein expression in Huh7 and BB7 replicon cells. HCV RNA synthesis in BB7 cells was inhibited by CXCL-8 knockdown. Furthermore, antibody neutralization of endogenous CXCL-8 activity inhibited HCV replication, while addition of recombinant human CXCL-8 stimulated NS5A protein expression. Moreover, CXCL-8 protein levels correlated positively with HCV RNA levels in four independent subgenomic and genomic replicon lines (R= 0.41,P= 0.0013). However, CXCL-8 mRNA levels correlated inversely with CXCL-8 protein and HCV RNA levels in all replicon lines and in Huh7 cells. Transient replication assays with strongly permissive and weakly permissive Huh7 cells and three independent subgenomic replicons with various replicative capacities revealed that CXCL-8 protein levels were higher in weakly than in strongly permissive cells. The JFH-1 subgenomic replicon, which replicated to high levels in both strongly and weakly permissive Huh7 cells, induced CXCL-8 protein to high levels in both cell types. The data indicate that in the replicon system, CXCL-8 protein levels are positively associated with chronic HCV replication and that CXCL-8 removal inhibits HCV replication. During acute HCV replication, CXCL-8 production may be inhibitory to viruses with low replicative capacity. The data underscore the complex regulation of CXCL-8 mRNA and protein expression and further suggest that in addition to contributing to HCV pathology via proinflammatory actions, CXCL-8 may have opposing antiviral and proviral effects depending on the level of HCV replication, the cellular context, and whether the infection is acute or chronic.
Dissemination of Bovine Leukemia Virus-Infected Cells from a Newly Infected Sheep Lymph Node
Authors: Fulton B. E. , Portella M. , Radke K.
Published: July 2006

To investigate the early establishment of bovine leukemia virus (BLV) infection, we injected BLV-infected or mock-infected allogeneic cells into the shoulder of sheep in which an efferent lymphatic duct of the draining prescapular lymph node had been cannulated. Rare mononuclear cells acting as centers of BLV infection in culture were present within 4 to 6 days in efferent lymph and within 6 to 10 days in blood. Soon after BLV injection, immunoglobulin M
+
(IgM
+
) and CD8
+
cells increased in efferent lymph and oscillated reciprocally in frequency. CD8
+
blasts increased on days 4 to 6, when infectious centers increased 100-fold in lymph. On days 6 and 7, both lymph and blood were enriched with CD8
+
cells that were labeled late on day 5 with an intravenous pulse of 5-bromo-2′-deoxyuridine (BrdU). Lymph, but not blood, was enriched with BrdU
+
B cells on day 7. Capsid-specific antibodies became detectable in efferent lymph on days 6 to 8 and surface glycoprotein-specific antibodies on day 9, preceding their detection in serum by 9 to 14 days. Systemic dissemination of BLV-infected cells was thus accompanied by an increase in proliferating CD8
+
cells and the onset of BLV-specific antibodies in lymph. Infectious centers reached maximum frequencies of 0.2% in lymph by days 11 to 13, and then their frequencies increased by 5- to 40-fold in blood cells, suggesting that many infected blood cells do not recirculate back into lymph. Beginning on days 10 to 13, a subpopulation of B cells having high levels of surface IgM increased sharply in peripheral blood. Such cells were not present in lymph. After a day 16 pulse of BrdU, recently proliferated cells that stained intensely for surface IgM appeared in blood within 15 h. Predominantly B lymphocytes contained the viral capsid protein when lymph and blood cells were cultured briefly to allow BLV expression. However, both early in lymph and later in blood, BrdU
+
B cells greatly exceeded productively infected cells, indicating that new BLV infections stimulate proliferation of two different populations of B cells.
Human CD4
+
CD25
high
Cells Suppress Proliferative Memory Lymphocyte Responses to Herpes Simplex Virus Type 2
Authors: Diaz George A. , Koelle David M.
Published: July 2006

Lymphocytes with the regulatory CD4
+
CD25
+
phenotype frequently suppress memory T-cell responses. Murine herpes simplex virus type 1 (HSV-1) models have shown that CD4
+
CD25
+
cells can limit immunity-mediated corneal damage but slow viral clearance. We investigated the effect of CD4
+
CD25
+
cells from healthy HSV-2-infected humans on recall proliferative (CD4) responses to HSV-2. Depletion and reconstitution experiments were consistent with a suppressive effect of autologous blood-derived CD4
+
CD25
+
cells for whole HSV-2 antigen. Regulatory T cells may modulate human CD4 memory responses to HSV-2 and influence their antiviral and inflammatory functions.
Kaposi's Sarcoma-Associated Herpesvirus Latent Protein LANA Interacts with HIF-1α To Upregulate RTA Expression during Hypoxia: Latency Control under Low Oxygen Conditions
Authors: Cai Qiliang , Lan Ke , Verma Subhash C. , Si Huaxin , Lin Doug , Robertson Erle S.
Published: July 2006

Hypoxia can induce lytic replication of Kaposi's sarcoma-associated herpesvirus (KSHV) in primary effusion lymphoma (PEL) cells. However, the molecular mechanism of lytic reactivation of KSHV by hypoxia remains unclear. Here we show that the latency-associated nuclear antigen (LANA), which plays a crucial role in modulating viral and cellular gene expression, directly associated with a low oxygen responder, hypoxia-inducible factor-1α (HIF-1α). LANA enhanced not only the transcriptional activities of HIF-1α but also its mRNA level. Coimmunoprecipitation and immunofluorescence studies documented a physical interaction between LANA and HIF-1α in transiently transfected 293T cells as well as in PEL cell lines during hypoxia. Through sequence analysis, several putative hypoxia response elements (HRE-1 to -6) were identified in the essential lytic geneRtapromoter. Reporter assays showed that HRE-2 (−1130 to −1123) and HRE-5 and HRE-6 (+234 to +241 and +812 to +820, respectively, within the intron sequence) were necessary and sufficient for the LANA-mediated HIF-1α response. Electrophoretic mobility shift assays showed HIF-1α-dependent binding of a LANA protein complex specifically to the HRE-2, -5, and -6 motifs within the promoter regulatory sequences. This study demonstrates that hypoxia-induced KSHV lytic replication is mediated at least in part through cooperation of HIF-1α with LANA bound to the HRE motifs of theRtapromoter.
Loss of Cell Membrane Integrity in Puumala Hantavirus-Infected PatientsCorrelates with Levels of Epithelial Cell Apoptosis and Perforin
Authors: Klingström Jonas , Hardestam Jonas , Stoltz Malin , Zuber Bartek , Lundkvist Åke , Linder Stig , Ahlm Clas
Published: July 2006

Hemorrhagic fever with renal syndrome and hantavirus pulmonary syndrome are two diseases caused by hantaviruses. Capillary leakage is a hallmark of hantavirus infection. Pathogenic hantaviruses are not cytotoxic, but elevated levels of serum lactate dehydrogenase (LDH), indicative of cellular damage, are observed in patients. We report increased levels of serum perforin, granzyme B, and the epithelial cell apoptosis marker caspase-cleaved cytokeratin-18 during Puumala hantavirus infection. Significant correlation was observed between the levels of LDH and perforin and the levels of LDH and caspase-cleaved cytokeratin-18, suggesting that tissue damage is due to an immune reaction and that epithelial apoptosis contributed significantly to the damage.
Evidence for a Role of the Cellular ND10 Protein PML in Mediating Intrinsic Immunity against Human Cytomegalovirus Infections
Authors: Tavalai Nina , Papior Peer , Rechter Sabine , Leis Martina , Stamminger Thomas
Published: July 2006

Several viruses, including human cytomegalovirus (HCMV), encode proteins that colocalize with a cellular subnuclear structure known as ND10. Since only viral DNA deposited at ND10 initiates transcription, ND10 structures were hypothesized to be essential for viral replication. On the other hand, interferon treatment induces an up-regulation of ND10 structures and viruses have evolved polypeptides that disperse the dot-like accumulation of ND10 proteins, suggesting that ND10 could also be part of an intrinsic defense mechanism. In order to obtain evidence for either a proviral or an antiviral function of ND10, we generated primary human fibroblasts with a stable, short interfering RNA-mediated knockdown (kd) of PML. In these cells, other ND10-associated proteins like hDaxx showed a diffuse nuclear distribution. Interestingly, we observed that HCMV infection induced the de novo formation of ND10-like hDaxx and Sp100 accumulations that colocalized with IE2 and were disrupted, in the apparent absence of PML, in an IE1-dependent manner during the first hours after infection. Furthermore, infection of PML-kd cells with wild-type HCMV at a low multiplicity of infection resulted in enhanced replication. In particular, a significantly increased plaque formation was detected, suggesting that more cells are able to support initiation of replication in the absence of PML. While there was no difference in viral DNA uptake between PML-kd and control cells, we observed a considerable increase in the number of immediate-early (IE) protein-positive cells, indicating that the depletion of PML augments the initiation of viral IE gene expression. These results strongly suggest that PML functions as part of an intrinsic immune mechanism against cytomegalovirus infections.
The Self Primer of the Long Terminal Repeat Retrotransposon Tf1 Is Not Removed during Reverse Transcription
Authors: Atwood-Moore Angela , Yan Kenneth , Judson Robert L. , Levin Henry L.
Published: July 2006

The long terminal repeat retrotransposon Tf1 of
Schizosaccharomyces pombe
uses a unique mechanism of self priming to initiate reverse transcription. Instead of using a tRNA, Tf1 primes minus-strand synthesis with an 11-nucleotide RNA removed from the 5′ end of its own transcript. We tested whether the self primer of Tf1 was similar to tRNA primers in being removed from the cDNA by RNase H. Our analysis of Tf1 cDNA extracted from virus-like particles revealed the surprising observation that the dominant species of cDNA retained the self primer. This suggests that integration of the cDNA relies on mechanisms other than reverse transcription to remove the primer.
Interferon α sensitisation induced fatal renal insufficiency in a patient with chronic myeloid leukaemia: case report and review of literature: Figure 1
Authors: Colovic M , Jurisic V , Jankovic G , Jovanovic D , Nikolic L J , Dimitrijevic J
Published: July 2006

Renal dysfunction as a sequel to extended interferon alfa (IFNα) treatment in chronic myeloid leukaemia (CML) has been reported previously in six patients. An additional patient is presented with Philadelphia chromosome positive (Ph+) CML and nephrotic syndrome in whom initial renal insufficiency developed after only one month of low dose IFNα therapy. The renal biopsy showed a focal segmental mononuclear cell infiltration, basal membrane thickening, and deposits of immunoglobulins (IgG; IgAGM IC3). In spite of discontinuation of IFNα, renal function deteriorated and the patient died six months later. This case represents an instance of fatal kidney insufficiency as an untoward effect of sensitisation to the IFNα, confirmed by modified Coombs assay.
My approach to the interpretation of endometrial biopsies and curettings
Authors: McCluggage W G
Published: July 2006

A major proportion of the workload in many histopathology laboratories is accounted for by endometrial biopsies, either curettage specimens or outpatient biopsy specimens. The increasing use of pipelle and other methods of biopsy not necessitating general anaesthesia has resulted in greater numbers of specimens with scant tissue, resulting in problems in assessing adequacy and in interpreting artefactual changes, some of which appear more common with outpatient biopsies. In this review, the criteria for adequacy and common artefacts in endometrial biopsies, as well as the interpretation of endometrial biopsies in general, are discussed, concentrating on areas that cause problems for pathologists. An adequate clinical history, including knowledge of the age, menstrual history and menopausal status, and information on the use of exogenous hormones and tamoxifen, is necessary for the pathologist to critically evaluate endometrial biopsies. Topics such as endometritis, endometrial polyps, changes that are induced by hormones and tamoxifen within the endometrium, endometrial metaplasias and hyperplasias, atypical polypoid adenomyoma, adenofibroma, adenosarcoma, histological types of endometrial carcinoma and grading of endometrial carcinomas are discussed with regard to endometrial biopsy specimens rather than hysterectomy specimens. The value of ancillary techniques, especially immunohistochemistry, is discussed where appropriate.
Immunohistochemical markers of prognosis in non-small cell lung cancer: a review and proposal for a multiphase approach to marker evaluation
Authors: Zhu C-Q , Shih W , Ling C-H , Tsao M-S
Published: July 2006

Characteristics of the tumour that affect and predict the survival outcome of patients with cancer are prognostic markers for cancer. In non-small cell lung carcinoma (NSCLC), stage is the main determinant of prognosis and the basis for deciding options for treatment. Patients with early-stage tumour are treated by complete surgical resection, which is curative in 40–70% of patients. That there are other factors important in determining the biology of these tumours, especially genes that have a role in metastasis, is indicated. Such factors could potentially be used to further classify patients into groups according to substages that may be treated differently. During the past decade, a large number of proteins that are putatively important in carcinogenesis and cancer biology have been studied for their prognostic value in NSCLC, but none of them have been proved to be sufficiently useful in clinical diagnosis. Several markers (epidermal growth factor receptor, human epidermal growth factor receptor 2, Ki-67, p53 and Bcl-2) have been studied exhaustively. Ki-67, p53 and Bcl-2 are suggested to be important but weak prognostic markers, by meta-analyses of the results. Cyclin E, vascular endothelial growth factor A, p16INK4A, p27kip1and β-catenin are promising candidates, but require further study in large randomised clinical trial samples by using standardised assays and scoring systems. Some issues and inconsistencies in the reported studies to date are highlighted and discussed. A guideline for a multi-phase approach for conducting future studies on prognostic immunohistochemistry markers is proposed here.
Lymphoproliferative lesions of the skin
Authors: Cerroni L
Published: July 2006

Diagnosis and differential diagnosis of cutaneous lymphoproliferative disorders is one of the most difficult areas in dermatopathology, and biopsies are often taken to rule out a cutaneous lymphoma in patients with “unclear” or “therapy-resistant” skin lesions. Histopathological features alone often enable a given case to be classified to a diagnostic group (eg, epidermotropic lymphomas), but seldom allow a definitive diagnosis to be made. Performing several biopsies from morphologically different lesions is suggested, especially in patients with suspicion of mycosis fungoides. Immunohistochemistry is often crucial for proper classification of the cases, but in some instances is not helpful (eg, early lesions of mycosis fungoides). Although molecular techniques provide new, powerful tools for diagnosing cutaneous lymphoproliferative disorders, results of molecular methods should always be interpreted with the clinicopathological features, keeping in mind the possibility of false positivity and false negativity. In many cases, a definitive diagnosis can be made only on careful correlation of the clinical with the histopathological, immunophenotypical and molecular features.
Best practice in primary care pathology: review 3
Authors: Smellie W S A , Forth J , Bareford D , Twomey P , Galloway M J , Logan E C M , Smart S R S , Reynolds T M , Waine C
Published: July 2006

This best practice review examines four series of common primary care questions in laboratory medicine: (i) “minor” blood platelet count and haemoglobin abnormalities; (ii) diagnosis and monitoring of anaemia caused by iron deficiency; (iii) secondary hyperlipidaemia and hypertriglyceridaemia; and (iv) glycated haemoglobin and microalbumin use in diabetes. The review is presented in question–answer format, referenced for each question series. The recommendations represent a précis of guidance found using a standardised literature search of national and international guidance notes, consensus statements, health policy documents and evidence-based medicine reviews, supplemented by Medline Embase searches to identify relevant primary research documents. They are not standards, but form a guide to be set in the clinical context. Most of the recommendations are based on consensus rather than evidence. They will be updated periodically to take account of new information.
MORE ON GENERAL p-BRANE SOLUTIONS
Authors: GAL'TSOV D. , KLEVTSOV S. , ORLOV D. , CLÉMENT G.
Published: July 2006

Recently it was found that the complete integration of the Einstein-dilaton-antisymmetric form equations depending on one variable and describing static singly charged p-branes leads to two and only two classes of solutions: the standard asymptotically flat black p-brane and the asymptotically nonflat p-brane approaching the linear dilaton background at spatial infinity. Here we analyze this issue in more details and generalize the corresponding uniqueness argument to the case of partially delocalized branes. We also consider the special case of codimension one and find, in addition to the standard domain wall, the black wall solution. Explicit relations between our solutions and some recently found p-brane solutions "with extra parameters" are presented.
SCALE AND CONFORMAL INVARIANCE AND VARIATION OF THE METRIC
Authors: SADEGHI J. , TOFIGHI A. , BANIJAMALI A.
Published: July 2006

We consider the relation between scale invariance and conformal invariance. In our analysis the variation of the metric is taken into account. By imposing some conditions on the trace of the energy–momentum tensor and on the variation of the action, we find that the scale dimensions of the fields are not affected. We also obtain the conserved currents. We find that the conditions for conformal invariance are stronger than for scale invariance.
q-DEFORMED KINK SOLUTIONS
Authors: DE LIMA A. F. , DE LIMA RODRIGUES R.
Published: July 2006

The q-deformed kink of the λϕ4-model is obtained via the normalizable ground state eigenfunction of a fluctuation operator associated with the q-deformed hyperbolic functions. From such a bosonic zero-mode the q-deformed potential in 1+1 dimensions is found, and we show that the q-deformed kink solution is a kink displaced away from the origin.
SCALAR MESONS AND LOW-MASS SIGMA: DOES THE σ REVEAL THE CONFINEMENT SINGULARITY?
Authors: ANISOVICH V. V.
Published: July 2006

We present a short review of the current understanding of the scalar meson sector, with special attention to the problem of the low-mass σ. The dispersion relation N/D-method used for the restoration of the low-energy ππ (IJ PC = 00++)-wave amplitude is discussed. The low-energy ππ amplitude was determined from the data in the energy region 280–500 MeV and it was sewn with the previously found K-matrix solution for the region 450–1950 MeV. The N/D-amplitude has a pole on the second sheet of the complex-s plane, near the ππ threshold at [Formula: see text], that corresponds to the low-mass σ-meson. We discuss the hypothesis that this pole may be related to the confinement forces, thus being the eyewitness of confinement.
COORDINATE-INVARIANT PATH INTEGRAL METHODS IN CONFORMAL FIELD THEORY
Authors: VAN TONDER ANDRÉ
Published: July 2006

We present a coordinate-invariant approach, based on a Pauli–Villars measure, to the definition of the path integral in two-dimensional conformal field theory. We discuss some advantages of this approach compared to the operator formalism and alternative path integral approaches. We show that our path integral measure is invariant under conformal transformations and field reparametrizations, in contrast to the measure used in the Fujikawa calculation, and we show the agreement, despite different origins, of the conformal anomaly in the two approaches. The natural energy–momentum in the Pauli–Villars approach is a true coordinate-invariant tensor quantity, and we discuss its nontrivial relationship to the corresponding nontensor object arising in the operator formalism, thus providing a novel explanation within a path integral context for the anomalous Ward identities of the latter. We provide a direct calculation of the nontrivial contact terms arising in expectation values of certain energy–momentum products, and we use these to perform a simple consistency check confirming the validity of the change of variables formula for the path integral. Finally, we review the relationship between the conformal anomaly and the energy–momentum two-point functions in our formalism.
COARSE-GRAINING 1/2 BPS GEOMETRIES OF TYPE IIB SUPERGRAVITY
Authors: BUCHEL ALEX
Published: July 2006

Recently Lin, Lunin and Maldacena (LLM)1 explicitly mapped 1/2 BPS excitations of type IIB supergravity on AdS 5 × S 5 into free fermion configurations. We discuss thermal coarse-gaining of LLM geometries by explicitly mapping the corresponding equilibrium finite temperature fermion configuration into supergravity. Following Mathur conjecture, a prescription of this sort should generate a horizon in the geometry. We did not find a horizon in finite temperature equilibrium LLM geometry. This most likely is due to the fact that coarse-graining is performed only in a half-BPS sector of the full Hilbert space of type IIB supergravity. For temperatures much less than the AdS curvature scale the equilibrium background corresponding to nearly degenerate dual Fermi-gas is found analytically.
A REVIEW OF DISTRIBUTIONS ON THE STRING LANDSCAPE
Authors: KUMAR JASON
Published: July 2006

We review some basic flux vacua counting techniques and results, focusing on the distributions of properties over different regions of the landscape of string vacua and assessing the phenomenological implications. The topics we discuss include: an overview of how moduli are stabilized and how vacua are counted; the applicability of effective field theory; the uses of and differences between probabilistic and statistical analysis (and the relation to the anthropic principle); the distribution of various parameters on the landscape, including cosmological constant, gauge group rank, and supersymmetry-breaking scale; "friendly landscapes;" open string moduli; the (in)finiteness of the number of phenomenologically viable vacua; etc. At all points, we attempt to connect this study to the phenomenology of vacua which are experimentally viable.
AXIAL SEGREGATION IN UNSTEADY DIFFUSION-DOMINATED SOLIDIFICATION IN A FINITE AMPOULE
Authors: NECULAE ADRIAN , BALINT AGNETA M.
Published: July 2006

The paper deals with axial segregation in unsteady diffusion dominated directional solidification from melt of a binary alloy in a finite cylindrical ampoule. The study focuses on the influence of the finite length of the ampoule on the concentration profile in a crystal obtained in a microgravity environment. The results are compared to those reported by Kim et al. [J. Crystal Growth183, 490 (1998)] for an infinite crystal. Numerical simulations are performed for two different situations: the initial transient solidification when the melt is uniform in composition, and the multipass solidification when the crystal is grown from the melt, formed by re-melting a crystal with an initial axial composition profile.