bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison and validation of three measures of quality of life in patients with pulmonary hypertension
Authors: Chua R. , Keogh A. M. , Byth K. , O’Loughlin A.
Published: October 2006

Background: Pulmonary hypertension, when advanced, markedly limits exercise capacity, activities of daily living and quality of life (QoL). No measure of QoL has yet been validated for the assessment of pulmonary hypertension. The aim of the study was to compare the validity of the Minnesota Living with Heart Failure (MLwHF) questionnaire, the Short Form‐36 (SF‐36) questionnaire and the Australian Quality of Life (AQoL) measure for assessing pulmonary hypertension treatment.

Methods: Eighty‐three patients were enrolled in three studies of pulmonary hypertension treatment (treprostinil, bosentan and sildenafil). They were assessed at baseline and 3 months with the MLwHF questionnaire. Treprostinil and bosentan groups also had 6 and 12 months’ data. Twenty‐one patients in the sildenafil trial completed concurrently, the SF‐36 and AQoL measures at baseline and 3 months. QoL scores were correlated with the 6‐min walk test distance, New York Heart Association functional class and right heart catheter‐derived haemodynamic parameters of the disease for all matching time points and for changes in scores and clinical measurements over time.

Results: The MLwHF and SF‐36 scores correlated well with the 6‐min walk test distance and New York Heart Association functional class, but did not correlate with haemodynamic measurements. MLwHF and SF‐36 scores also correlated with the rate of change of the 6‐min walk test distance and New York Heart Association functional class over time.

Conclusion: The MLwHF questionnaire and SF‐36 are useful tools for the assessment of QoL in pulmonary hypertension and may be useful in the ongoing evaluation of QoL in the treatment and study of pulmonary hypertension.
Acute copper toxicity following copper glycinate injection
Authors: Oon S. , Yap C‐H. , Ihle B. U.
Published: October 2006

We present a patient who developed multi‐organ failure due to severe copper toxicity following attempted suicide by s.c. injection of copper glycinate. Acute copper toxicity is rare in the developed world, although it occurs more frequently in developing world countries, where it is a common mode of suicide. Acute toxicity usually results from oral ingestion and there are several local and systemic effects. Specific management can be difficult as there is little evidence regarding the efficacy of chelating agents in acute toxicity.
Bronchiectasis
Authors: King P. , Holdsworth S. , Freezer N. , Holmes P.
Published: October 2006

Bronchiectasis is generally classified into cystic fibrosis and non‐cystic fibrosis bronchiectasis. This review article describes non‐cystic fibrosis bronchiectasis in adults. Bronchiectasis can be considered a heterogeneous condition characterized by irreversible airway dilatation with chronic bronchial infection/inflammation. It remains a common condition and is a major cause of respiratory morbidity. Many factors are associated with bronchiectasis, but most commonly patients will have idiopathic disease. Important clinical findings include chronic productive cough, rhinosinusitis, fatigue and bi‐basal crackles. Patients have usually had symptoms for many years. Diagnosis is confirmed by high‐resolution computed tomography scanning using standardized criteria. Spirometry shows moderate airflow obstruction and there is a high prevalence of bronchial hyperreactivity. The most common pathogens are non‐typeable Haemophilus influenzae and Pseudomonas aeruginosa. There may be considerable overlap with other chronic airway diseases. Treatment regimens are still not well defined. Patients tend to have ongoing symptoms and decline in respiratory function despite treatment.
Clinical and quality‐of‐life outcomes in women treated by the TVT‐O procedure
Authors: Lim JL , Cornish A , Carey MP
Published: October 2006

Objectives To objectively assess the success rate at 6 months after tension‐free vaginal tape obturator (TVT‐O) procedure. To assess subjective success rates, complications, patient satisfaction, and quality of life (QOL).

Design A prospective observational study.

Setting A tertiary referral urogynaecology practice.

Population A cohort of 100 consecutive women who underwent the TVT‐O procedure between March and October 2004.

Methods The TVT‐O technique was performed as described. Three standardised QOL questionnaires were completed preoperatively at 6 months and 12 months. At 6 months, a urogenital history, visual analogue scale score (VAS) for patient satisfaction, uroflow, and urinary stress test were performed. After 12 months, a urogenital history and patient satisfaction verbal analogue score (VeAS) were obtained by telephone interview.

Main outcome measure Objective success rate of the TVT‐O procedure was measured by negative stress test. Secondary outcomes were subjective success rates at 6 and 12 months, pre‐ and postoperative comparison of urodynamic parameters, complications, postoperative symptomatology, QOL analysis, and patient satisfaction.

Results Mean follow up was 18.5 months. Objective success rate was 95%. Subjective success rates were 92 and 84% at 6 and 12 months. Complications included recurrent urinary tract infection (six), voiding difficulty (two), persistent groin discomfort (three), haematoma (one), wound infection (one), vaginal tape erosion (one), and urethral irritation (one). Prevalence of de novo urge incontinence was 4.1 and 4.8% at 6 and 12 months. QOL analysis showed significant improvements in QOL scores postoperatively. Visual and verbal analogue scores indicated high patient satisfaction (VAS, VeAS ≥ 80%) in 77 and 67% at 6 and 12 months.

Conclusion The TVT‐O is a safe and effective treatment for female stress urinary incontinence.
Conditioned olfactory responses of a predatory mite, Neoseiulus womersleyi, to volatiles from prey‐infested plants
Authors: Maeda Taro , Liu Yining , Ishiwari Hayato , Shimoda Takeshi
Published: October 2006

To clarify the prey‐finding behavior of the predatory mite Neoseiulus womersleyi (Schicha) (Acari: Phytoseiidae), we studied its olfactory responses to volatiles from the prey‐infested plant on which the mites had been collected. We used a local N. womersleyi population called Kanaya collected from tea (Camellia sinensis L.) (Theaceae) plants infested by Tetranychus kanzawai Kishida (Acari: Tetranychidae) in Kanaya City, Japan. Neoseiulus womersleyi (Kanaya population) were more attracted to volatiles from tea plants infested with five female T. kanzawai per leaf for 7 days than to intact tea leaves in a Y‐tube olfactometer. Tetranychus kanzawai‐induced tea leaf volatiles were identified as (E)‐β‐ocimene, (E)‐4,8‐dimethyl‐1,3,7‐nonatriene, and (E,E)‐α‐farnesene. As olfactory responses are known to differ among local populations of N. womersleyi, we compared the responses of the Kanaya population with those of a Kikugawa population collected from tea plants infested by T. kanzawai in Kikugawa City. To test the influence of previous predation experience, we reared the two populations on tea plants infested by T. kanzawai or on kidney bean plants (Phaseolus vulgaris) infested by Tetranychus urticae Koch. The Kanaya population was more attracted to the volatiles from infested plants on which they had been reared. Because the Kanaya population was not attracted to the plant volatiles they had not previously experienced, the positive response to previously experienced volatiles might be the result of learning. By contrast, the Kikugawa population showed no preference for previously experienced volatiles from infested plants. The implications of this flexibility in foraging behavior are discussed.
Effects of host‐plant species on diapause induction of the Kanzawa spider mite, Tetranychus kanzawai
Authors: Ito Katsura , Saito Yutaka
Published: October 2006

In many herbivorous arthropods, incidence of diapause, which is considered to reflect the timing of diapause, changes depending on the host plants they utilize. Several theoretical studies suggest that the optimal timing of diapause induction depends on life‐history traits; if the development time of the arthropod is short, fecundity is high, or survival rate remains high throughout the season, the optimal timing of diapause induction would be shifted toward the end of the season. For herbivorous arthropods, these life history traits may change among their host plants. Here we examined whether a population of the Kanzawa spider mite, Tetranychus kanzawai Kishida (Acari: Tetranychidae), shows the predicted pattern of diapause induction on two host plants, the kidney bean [Phaseolus vulgaris (Leguminosae)] and Japanese Orixa [Orixa japonica (Rutaceae)], on which the mites show different performances. Rearing conditions were controlled in two ways. In the first experiment, day length and temperature were kept constant throughout the mite lifetime at either of three conditions from mid October to early November. In the second experiment, the conditions were changed from 20 °C and L11.5:D12.5 at immature stages to 18 °C and L11:D13 at adult stage in order to better approximate field conditions. In the first experiment, diapause incidence on P. vulgaris was lower than on O. japonica. This tendency became stronger in the second experiment, suggesting a difference in the timing of diapause induction among host plants. On the other hand, P. vulgaris was proven to allow high performance, i.e., greater lifetime fecundity and shorter development times, although it had no effect on the survival rate. The relationship between diapause incidence and performance is consistent with the prediction of theoretical studies that a short development time or high fecundity delays the timing of diapause induction.
Drosophila melanogaster: the model organism
Authors: Roberts David B.
Published: October 2006

In the 20th century, there were two decades during which Drosophila melanogaster was the most significant model organism and each decade led to the establishment of new scientific disciplines. The first decade was roughly from 1910 and during this period a small group at Columbia University, headed by Thomas Hunt Morgan, established the rules of transmission genetics with which we are all familiar. In the second decade, roughly from 1970, many of the principles and techniques of the earlier period were used to determine the genetic control of basic aspects of the biology of organisms, notably their development and their behaviour. In this review I will show that it was not only the genius of the research workers (five were awarded Nobel Prizes and it has been argued, with justification, that at least one more should have been awarded) but also the special features of D. melanogaster that led to these advances. While Drosophila is still a significant model organism, the advent of molecular biology permits the investigation of organisms less amenable to genetic analysis, but the principles applied in these investigations were in the main principles laid down during the earlier work on Drosophila.
Heterogeneity and Temporal Dynamics of Evolution of G1 Human Rotaviruses in a Settled Population
Authors: Arista Serenella , Giammanco Giovanni M. , De Grazia Simona , Ramirez Stefania , Lo Biundo Concetta , Colomba Claudia , Cascio Antonio , Martella Vito
Published: October 2006

A rotavirus sample collection from 19 consecutive years was used to investigate the heterogeneity and the dynamics of evolution of G1 rotavirus strains in a geographically defined population. Phylogenetic analysis of the VP7 gene sequences of G1P[8] human rotavirus strains showed the circulation of a heterogeneous population comprising three lineages and seven sublineages. Increases in the circulation of G1 rotaviruses were apparently associated with the introduction of novel G1 strains that exhibited multiple amino acid changes in antigenic regions involved in rotavirus neutralization compared to the strains circulating in the previous years. The emergence and/or introduction of G1 antigenic variants might be responsible for the continuous circulation of G1 rotaviruses in the local population, with the various lineages and sublineages appearing, disappearing, or cocirculating in an alternate fashion under the influence of immune-pressure mechanisms. Sequence analysis of VP4-encoding genes of the G1 strains revealed that the older strains were associated with a unique VP4 lineage, while a novel VP4 lineage emerged after 1995. The introduction of human rotavirus vaccines might alter the forces and balances that drive rotavirus evolution and determine the spread of novel strains that are antigenically different from those included in the vaccine formulations. The continuous emergence of VP7-VP4 gene combinations in human rotavirus strains should be taken into consideration when devising vaccination strategies.
PmRab7 Is a VP28-Binding Protein Involved in White Spot Syndrome Virus Infection inShrimp
Authors: Sritunyalucksana Kallaya , Wannapapho Wanphen , Lo Chu Fang , Flegel Timothy W.
Published: October 2006

Our
aim was to isolate and characterize white spot syndrome virus
(WSSV)-binding proteins from shrimp. After a blot of shrimp hemocyte
membrane proteins was overlaid with a recombinant WSSV envelope protein
(rVP28), the reactive bands on the blot were detected using anti-VP28
antibody. Among three membrane-associated molecules identified by
liquid chromatography-tandem mass spectrometry, there was a 25-kDa
protein that bound to both rVP28 and WSSV. Since it had a primary
structure with high homology to the small GTP-binding protein Rab7, we
named it
Penaeus monodon
Rab7 (PmRab7). The full-length PmRab7
cDNA was obtained, and results from a glutathione
S
-transferase pull-down assay confirmed specific binding to
rVP28. Reverse transcriptase PCR analysis revealed PmRab7 expression in
many tissues, and real-time PCR analysis revealed that expression was
constitutive. Binding of PmRab7 to rVP28 or WSSV occurred in a
dose-dependent manner and was inhibited by anti-Rab7 antibody. In an in
vivo neutralization assay, the number of dead shrimp after challenge
with WSSV plus PmRab7 (15%) or WSSV plus anti-Rab7 antibody (5%) was
significantly lower than after challenge with WSSV alone (95%). In
contrast to the WSSV-injected group, shrimp injected with WSSV plus
PmRab7 or WSSV plus anti-Rab7 showed no WSSV-type histopathology. We
conclude that PmRab7 is involved in WSSV infection in shrimp. This is
the first study to identify a shrimp protein that binds directly to a
major viral envelope protein of
WSSV.
Rotavirus Activates JNK and p38 Signaling Pathways in Intestinal Cells, Leading to AP-1-Driven Transcriptional Responses and Enhanced Virus Replication
Authors: Holloway Gavan , Coulson Barbara S.
Published: October 2006

Rotavirus infection is known to regulate transcriptional changes in many cellular genes. The transcription factors NF-κB and AP-1 are activated by rotavirus infection, but the upstream processes leading to these events are largely unidentified. We therefore studied the activation state during rotavirus infection of c-Jun NH2-terminal kinase (JNK) and p38, which are kinases known to activate AP-1. As assessed by Western blotting using phospho-specific antibodies, infection with rhesus rotavirus (RRV) or exposure to UV-psoralen-inactivated RRV (I-RRV) resulted in the activation of JNK in HT-29, Caco-2, and MA104 cells. Activation of p38 during RRV infection was observed in Caco-2 and MA104 cells but not in HT-29 cells, whereas exposure to I-RRV did not lead to p38 activation in these cell lines. Rotavirus strains SA11, CRW-8, Wa, and UK also activated JNK and p38. Consistent with the activation of JNK, a corresponding increase in the phosphorylation of the AP-1 component c-Jun was shown. The interleukin-8 (IL-8) and c-junpromoters contain AP-1 binding sequences, and these genes have been shown previously to be transcriptionally up-regulated during rotavirus infection. Using specific inhibitors of JNK (SP600125) and p38 (SB203580) and real-time PCR, we showed that maximal RRV-induced IL-8 and c-juntranscription required JNK and p38 activity. This highlights the importance of JNK and p38 in RRV-induced, AP-1-driven gene expression. Significantly, inhibition of p38 or JNK in Caco-2 cells reduced RRV growth but not viral structural antigen expression, demonstrating the potential importance of JNK and p38 activation for optimal rotavirus replication.
Administration of Fludarabine-Loaded Autologous Red Blood Cells in Simian Immunodeficiency Virus-Infected Sooty Mangabeys Depletes pSTAT-1-Expressing Macrophages and Delays the Rebound of Viremia after Suspension of Antiretroviral Therapy
Authors: Cervasi B. , Paiardini M. , Serafini S. , Fraternale A. , Menotta M. , Engram J. , Lawson B. , Staprans S. I. , Piedimonte G. , Perno C. F. , Silvestri G. , Magnani M.
Published: October 2006

A
major limitation of highly active antiretroviral therapy is that it
fails to eradicate human immunodeficiency virus (HIV) infection due to
its limited effects on viral reservoirs carrying replication-competent
HIV, including monocytes/macrophages (M/M). Therefore, therapeutic
approaches aimed at targeting HIV-infected M/M may prove useful in the
clinical management of HIV-infected patients. In previous studies, we
have shown that administration of fludarabine-loaded red blood cells
(RBC) in vitro selectively induces cell death in HIV-infected M/M via a
pSTAT1-dependent pathway. To determine the in vivo efficacy of this
novel therapeutic strategy, we treated six naturally simian
immunodeficiency virus (SIV)-infected sooty mangabeys (SMs) with either
9-[2-(
R
)-(phosphonomethoxy)propyl]adenine (PMPA) only,
fludarabine-loaded RBC only, or PMPA in association with
fludarabine-loaded RBC. The rationale of this treatment was to target
infected M/M with fludarabine-loaded RBC at a time when PMPA is
suppressing viral replication taking place in activated
CD4
+
T cells. In vivo administration of
fludarabine-loaded RBC was well tolerated and did not induce any
discernible side effect. Importantly, addition of fludarabine-loaded
RBC to PMPA delayed the rebound of viral replication after suspension
of therapy, thus suggesting a reduction in the size of SIV reservoirs.
While administrations of fludarabine-loaded RBC did not induce any
change in the CD4
+
or CD8
+
T-cell
compartments, we observed, in chronically SIV-infected SMs, a selective
depletion of M/M expressing pSTAT1. This study suggests that
therapeutic strategies based on the administration of
fludarabine-loaded RBC may be further explored as interventions aimed
at reducing the size of the M/M reservoirs during chronic HIV
infection.
TaxAbolishes Histone H1 Repression of p300 Acetyltransferase Activity at the Human T-Cell Leukemia Virus Type 1Promoter
Authors: Konesky Kasey L. , Nyborg Jennifer K. , Laybourn Paul J.
Published: October 2006

Upon
infection of human T-cell leukemia virus type 1 (HTLV-1), the provirus
is integrated into the host cell genome and subsequently packaged into
chromatin that contains histone H1. Consequently, transcriptional
activation of the virus requires overcoming the environment of
chromatin and H1. To efficiently activate transcription, HTLV-1
requires the virally encoded protein Tax and cellular transcription
factor CREB. Together Tax and CREB interact with three
cis
-acting promoter elements called viral cyclic-AMP response
elements (vCREs). Binding of Tax and CREB to the vCREs promotes
association of p300/CBP into the complex and leads to transcriptional
activation. Therefore, to fully understand the mechanism of Tax
transactivation, it is necessary to examine transcriptional activation
from chromatin assembled with H1. Using a DNA template harboring the
complete HTLV-1 promoter sequence and a highly defined recombinant
assembly system, we demonstrate proper incorporation of histone H1 into
chromatin. Addition of H1 to the chromatin template reduces HTLV-1
transcriptional activation through a novel mechanism. Specifically, H1
does not inhibit CREB or Tax binding to the vCREs or p300 recruitment
to the promoter. Rather, H1 directly targets p300 acetyltransferase
activity. Interestingly, in determining the mechanism of H1 repression,
we have discovered a previously undefined function of Tax, overcoming
the repressive effects of H1-chromatin. Tax specifically abrogates the
H1 repression of p300 enzymatic activity in a manner independent of
p300 recruitment and without displacement of H1 from the
promoter.
Sequential Turnover of Human Immunodeficiency Virus Type 1envthroughout the Course of Infection
Authors: Riddle Tara M. , Shire Norah J. , Sherman Marc S. , Franco Kelly F. , Sheppard Haynes W. , Nelson Julie A. E.
Published: October 2006

We examined the rates of variant population turnover of the V1-V2 and V4-V5 hypervariable domains of the human immunodeficiency virus type 1 (HIV-1) gp120 molecule in longitudinal plasma samples from 14 men with chronic HIV-1 infection using heteroduplex tracking assays (HTA). Six men had high rates of CD4+T-cell loss, and eight men had low rates of CD4+T-cell loss over 2.5 to 8 years of infection. We found that V1-V2 and V4-V5envpopulations changed dramatically over time in all 14 subjects; the changes in these regions were significantly correlated with each another over time. The subjects with rapid CD4 loss had significantly less change in theirenvpopulations than the subjects with slow CD4 loss. The two subjects with rapid CD4 loss and sustained low CD4 counts (<150/μl for at least 2 years) showed stabilization of their V1-V2 and V4-V5 populations as reflected by low levels of total change in HTA pattern and low HTA indices (a novel measure of the emergence of new bands and band distribution); this stabilization was not observed in other subjects. The stabilization ofenvvariant populations at low CD4 counts following periods of rapid viral evolution suggests that selective pressure onenv, likely from new immune responses, is minimal when CD4 counts drop dramatically and remain low for extended periods of time.
Latent Kaposi's Sarcoma-Associated Herpesvirus Infection of Endothelial Cells Activates Hypoxia-Induced Factors
Authors: Carroll Patrick A. , Kenerson Heidi L. , Yeung Raymond S. , Lagunoff Michael
Published: October 2006

Kaposi's sarcoma-associated herpesvirus (KSHV or HHV-8) is the etiological agent of Kaposi's sarcoma, a highly vascularized, endothelial-derived tumor. A direct role for KSHV-mediated induction of angiogenesis has been proposed based upon the nature of the neoplasia and various KSHV gene overexpression and infection model systems. We have found that KSHV infection of endothelial cells induces mRNA of hypoxia-induced factor 1α (HIF1α) and HIF2α, two homologous alpha subunits of the heterodimeric transcription factor HIF. HIF is a master regulator of both developmental and pathological angiogenesis, composed of an oxygen-sensitive alpha subunit and a constitutively expressed beta subunit. HIF is classically activated posttranscriptionally with hypoxia, leading to increased protein stability of HIF1α and/or HIF2α. However, we demonstrate that both alpha subunits are up-regulated at the transcript level by KSHV infection. The transcriptional activation of HIF leads to a functional increase in HIF activity under normoxic conditions, as demonstrated by both luciferase reporter assay and the increased expression of vascular endothelial growth factor receptor 1 (VEGFR1), an HIF-responsive gene. KSHV infection synergizes with hypoxia mimics and induces higher expression levels of HIF1α and HIF2α protein, and HIF1α is increased in a significant proportion of the latently infected endothelial cells. Src family kinases are required for the activation of HIF and the downstream gene VEGFR1 by KSHV. We also show that KS lesions, in vivo, express elevated levels of HIF1α and HIF2α proteins. Thus, KSHV stimulates the HIF pathway via transcriptional up-regulation of both HIF alphas, and this activation may play a role in KS formation, localization, and progression.
Identification of a Novel Nonstructural Protein, VP9, from White Spot Syndrome Virus: Its Structure Reveals a Ferredoxin Fold with Specific Metal Binding Sites
Authors: Liu Yang , Wu Jinlu , Song Jianxing , Sivaraman J. , Hew Choy L.
Published: October 2006

White spot syndrome virus (WSSV) is a major pathogen in shrimp aquaculture. VP9, a full-length protein of WSSV, encoded by open reading frame wsv230, was identified for the first time in the infected
Penaeus monodon
shrimp tissues, gill, and stomach as a novel, nonstructural protein by Western blotting, mass spectrometry, and immunoelectron microscopy. Real-time reverse transcription-PCR demonstrated that the transcription of VP9 started from the early to the late stage of WSSV infection as a major mRNA species. The structure of full-length VP9 was determined by both X-ray and nuclear magnetic resonance (NMR) techniques. It is the first structure to be reported for WSSV proteins. The crystal structure of VP9 revealed a ferredoxin fold with divalent metal ion binding sites. Cadmium sulfate was found to be essential for crystallization. The Cd
2+
ions were bound between the monomer interfaces of the homodimer. Various divalent metal ions have been titrated against VP9, and their interactions were analyzed using NMR spectroscopy. The titration data indicated that VP9 binds with both Zn
2+
and Cd
2+
. VP9 adopts a similar fold as the DNA binding domain of the papillomavirus E2 protein. Based on our present investigations, we hypothesize that VP9 might be involved in the transcriptional regulation of WSSV, a function similar to that of the E2 protein during papillomavirus infection of the host cells.
Human T-Cell Leukemia Virus Type 1 Tax Dysregulates β-CateninSignaling
Authors: Tomita Mariko , Kikuchi Akira , Akiyama Tetsu , Tanaka Yuetsu , Mori Naoki
Published: October 2006

Dysregulation
of β-catenin signaling has been implicated in the malignant
transformation of cells. However, the role of β-catenin in the
human T-cell leukemia virus type 1 (HTLV-1)-induced transformation of T
cells is unknown. Here we found that β-catenin protein was
overexpressed in the nucleus and that β-catenin-dependent
transcription was significantly enhanced in Tax-positive
HTLV-1-infected T-cell lines compared to that in Tax-negative
HTLV-1-infected T-cell lines. Transfection withβ
-catenin-specific small interfering RNA inhibited the growth
of the Tax-positive HTLV-1-infected T-cell line HUT-102. Transient
transfection of Tax appeared to enhance β-catenin-dependent
transcription by stabilizing the β-catenin protein via
activation of the cyclic AMP (cAMP) response element-binding protein.
HTLV-1-infected T-cell lines overexpressing β-catenin also
showed increased Akt activity via Tax activation of the cAMP response
element-binding protein, resulting in the phosphorylation and
inactivation of glycogen synthase kinase 3β, which
phosphorylates β-catenin for ubiquitination. The
phosphatidylinositol 3-kinase inhibitor LY294002 reducedβ
-catenin expression in Tax-positive T-cell lines, and
inactivation of glycogen synthase kinase 3β by lithium chloride
restored β-catenin expression in Tax-negative T-cell lines.
Finally, we showed that dominant-negative Akt inhibited Tax-inducedβ
-catenin-dependent transcription. These results indicate that
Tax activates β-catenin through the Akt signaling pathway. Our
findings suggest that activation of β-catenin by Tax may be
important in the transformation of T cells by HTLV-1
infection.
MurineHepatitis Virus Strain 1 Produces a Clinically Relevant Model of Severe Acute Respiratory Syndrome in A/J Mice
Authors: De Albuquerque Nadine , Baig Ehtesham , Ma Xuezhong , Zhang Jianhua , He William , Rowe Andrea , Habal Marlena , Liu Mingfeng , Shalev Itay , Downey Gregory P. , Gorczynski Reginald , Butany Jagdish , Leibowitz Julian , Weiss Susan R. , McGilvray Ian D. , Phillips M. James , Fish Eleanor N. , Levy Gary A.
Published: October 2006

Severe acute respiratory syndrome (SARS) is a life-threatening infectious disease which has been difficult to study and treat because of the lack of a readily available animal model. Intranasal infection of A/J mice with the coronavirus murine hepatitis virus strain 1 (MHV-1) produced pulmonary pathological features of SARS. All MHV-1-infected A/J mice developed progressive interstitial pneumonitis, including dense macrophage infiltrates, giant cells, and hyaline membranes, resulting in death of all animals. In contrast, other mouse strains developed only mild transitory disease. Infected A/J mice had significantly higher cytokine levels, particularly macrophage chemoattractant protein 1 (MCP-1/CCL-2), gamma interferon, and tumor necrosis factor alpha. Furthermore, FGL2/fibroleukin mRNA transcripts and protein and fibrin deposits were markedly increased in the lungs of infected A/J mice. These animals developed a less robust type I interferon response to MHV-1 infection than resistant C57BL/6J mice, and treatment with recombinant beta interferon improved survival. This study describes a potentially useful small animal model of human SARS, defines its pathogenesis, and suggests treatment strategies.
Posttranslational Modification and Cell Type-Specific Degradation of Varicella-Zoster Virus ORF29p
Authors: Stallings Christina L. , Silverstein Saul J.
Published: October 2006

The ORF29 gene of varicella-zoster virus encodes a single-stranded DNA binding protein that is predominantly nuclear during lytic infection but appears to be restricted to the cytoplasm of latently infected neurons. Following reactivation, ORF29p accumulates in the nuclei of neurons, suggesting that its confinement to the cytosol may be critical for maintaining quiescence. When autonomously expressed, ORF29p accumulates in the nuclei of fibroblasts and the cytoplasm of cells (guinea pig enteric neurons) and cell lines (U373MG) of neuronal origin. Inhibition of the 26S proteasome redirects the accumulation of ORF29p to the nucleus in cells of neuronal origin. Here, we show that ORF29p is ubiquitinated and sumoylated in 293T cells and subsequently degraded from the N terminus. Ubiquitinated ORF29p accumulates in both the nuclei and the cytoplasm of fibroblasts, but degradation products are seen primarily in the cytoplasm. Modification and degradation of ORF29p occurs in 293T, U373MG, and MeWo cells. Therefore, these processes are ubiquitous; however, the robustness of the degradation process is cell type specific. The proteasome-mediated mechanism of nuclear exclusion in U373MG cells is an active process that is not specific for the endogenous ORF29p nuclear localization signal but can be saturated by protein stabilization or overexpression, which leads to nuclear accumulation of ORF29p. The evidence for ORF29p ubiquitination and previous data regarding the effect of proteasome inhibitors on the abundance and distribution of ORF29p implicate the 26S proteasome in influencing the protein's cell type-specific localization.
The Unique C Termini of Orthopoxvirus Gamma Interferon Binding Proteins Are Essential for Ligand Binding
Authors: Nuara Anthony A. , Bai Hongdong , Chen Nanhai , Buller R. Mark L. , Walter Mark R.
Published: October 2006

The orthopoxviruses ectromelia virus (ECTV) and vaccinia virus (VACV) express secreted gamma interferon binding proteins (IFN-γBPs) with homology to the ligand binding domains of the host's IFN-γ receptor (IFN-γR1). Homology between these proteins is limited to the extracellular portions of the IFN-γR1 and the first ∼200 amino acids of the IFN-γBPs. The remaining 60 amino acids at the C termini of the IFN-γBPs contain a single cysteine residue shown to be important in covalent dimerization of the secreted proteins. The function of the remaining C-terminal domain (CTD) has remained elusive, yet this region is conserved within all orthopoxvirus IFN-γBPs. Using a series of C-terminal deletion constructs, we have determined that the CTD is essential for IFN-γ binding despite having no predicted homology to the IFN-γR1. Truncation of the ECTV IFN-γBP by more than two amino acid residues results in a complete loss of binding activity for both murine IFN-γ and human IFN-γ (hIFN-γ), as measured by surface plasmon resonance (SPR) and bioassay. Equivalent truncation of the VACV IFN-γBP resulted in comparable loss of hIFN-γ binding activity by SPR. Full-length IFN-γBPs were observed to form higher-ordered structures larger than the previously reported dimers. Mutants that were unable to bind IFN-γ with high affinity in SPR experiments failed to assemble into these higher-ordered structures and migrated as dimers. We conclude that the unique CTD of orthopoxvirus IFN-γBPs is important for the assembly of covalent homodimers as well as the assembly of higher-ordered structures essential for IFN-γ binding.
Antilipopolysaccharide Factor Interferes with White Spot Syndrome Virus Replication In Vitro and In Vivo in the Crayfish
Pacifastacus leniusculus
Authors: Liu Haipeng , Jiravanichpaisal Pikul , Söderhäll Irene , Cerenius Lage , Söderhäll Kenneth
Published: October 2006

In a study of genes expressed differentially in the freshwater crayfish
Pacifastacus leniusculus
infected experimentally with the white spot syndrome virus (WSSV), one protein, known as antilipopolysaccharide factor (ALF), was chosen, among those whose transcript levels increased upon viral infection, for further studies. ALF RNA interference (RNAi) experiments in whole animals and in cell cultures indicated that ALF can protect against WSSV infection, since knockdown of ALF by RNAi specifically resulted in higher rates of viral propagation. In a cell culture of hematopoietic tissue (Hpt) from
P. leniusculus
, quantitative PCR showed that knockdown of ALF by RNAi resulted into WSSV levels that were about 10-fold higher than those treated with control double-stranded RNA (dsRNA). In addition, RNAi experiments with other crayfish genes that had been found to be up-regulated by a WSSV infection did not result in any changes of viral loads. Thus, the cell culture does not respond to dsRNA in a similar manner, as shown earlier for dsRNA injected into shrimp, which gave a higher degree of resistance to WSSV infection. If ALF transcription in whole animals was stimulated by the administration of UV-treated WSSV, a partial protection against a subsequent challenge with the active virus was conferred to the host. This is the first crustacean gene product identified with the capacity to interfere with replication of this important pathogen.
Herpes Simplex Virus Type 1 ICP27-Dependent Activation of NF-κB
Authors: Hargett Danna , Rice Stephen , Bachenheimer Steven L.
Published: October 2006

The ability of herpes simplex virus type 1 (HSV-1) to activate NF-κB has been well documented. Beginning at 3 to 5 h postinfection, HSV-1 induces a robust and persistent nuclear translocation of an NF-κB-dependent (p50/p65 heterodimer) DNA binding activity, as measured by electrophoretic mobility shift assay. Activation requires virus binding and entry, as well as de novo infected-cell protein synthesis, and is accompanied by loss of both IκBα and IκBβ. In this study, we identified loss of IκBα as a marker of NF-κB activation, and infection with mutants with individual immediate-early (IE) regulatory proteins deleted indicated that ICP27 was necessary for IκBα loss. Analysis of both N-terminal and C-terminal mutants of ICP27 identified the region from amino acids 21 to 63 as being necessary for IκBα loss. Additional experiments with mutant viruses with combinations of IE genes deleted revealed that the ICP27-dependent mechanism of NF-κB activation may be augmented by functional ICP4. We also analyzed two additional markers for NF-κB activation, phosphorylation of the p65 subunit on Ser276 and Ser536. Phosphorylation of both serines was induced upon HSV infection and required functional ICP4 and ICP27. Pharmacological inhibitor studies revealed that both IκBα and Ser276 phosphorylation were dependent on Jun N-terminal protein kinase activity, while Ser536 phosphorylation was not affected during inhibitor treatment. These results demonstrate that there are several layers of regulation of NF-κB activation during HSV infection, highlighting the important role that NF-κB may play in infection.
Packaging Determinants in the UL11 Tegument Protein of Herpes Simplex Virus Type 1
Authors: Loomis Joshua S. , Courtney Richard J. , Wills John W.
Published: October 2006

The UL11 gene of herpes simplex virus type 1 encodes a 96-amino-acid tegument protein that is myristylated, palmitylated, and phosphorylated and is found on the cytoplasmic faces of nuclear, Golgi apparatus-derived, and plasma membranes of infected cells. Although this protein is thought to play a role in virus budding, its specific function is unknown. Purified virions were found to contain ∼700 copies of the UL11 protein per particle, making it an abundant component of the tegument. Moreover, comparisons of cell-associated and virion-associated UL11 showed that packaging is selective for underphosphorylated forms, as has been reported for several other tegument proteins. Although the mechanism by which UL11 is packaged is unknown, previous studies have identified several sequence motifs in the protein that are important for membrane binding, intracellular trafficking, and interaction with UL16, another tegument protein. To ascertain whether any of these motifs are needed for packaging, a transfection/infection-based assay was used in which mutant forms of the protein must compete with the wild type. In this assay, the entire C-terminal half of UL11 was found to be dispensable. In the N-terminal half, the sites of myristylation and palmitylation, which enable membrane-binding and Golgi apparatus-specific targeting, were found to be essential for efficient packaging. The acidic cluster motif, which is not needed for Golgi apparatus-specific targeting but is involved in recycling the protein from the plasma membrane and for the interaction with UL16, was found to be essential, too. Thus, something other than mere localization of UL11 to Golgi apparatus-derived membranes is needed for packaging. The critical factor is unlikely to be the interaction with UL16 because other mutants that fail to bind this protein (due to removal of the dileucine-like motif or substitutions with foreign acidic clusters) were efficiently packaged. Collectively, these results suggest that UL11 packaging is not driven by a passive mechanism but instead requires trafficking through a specific pathway.
Identification of Major Histocompatibility Complex Class II-Restricted Antigens and Epitopes of the Epstein-Barr Virus by a Novel Bacterial Expression Cloning Approach
Authors: Milosevic Slavoljub , Behrends Uta , Adhikary Dinesh , Mautner Josef
Published: October 2006

Epstein-Barr virus (EBV)-specific T cells have been successfully used to
treat or prevent EBV-positive lymphoproliferative disease in
hematopoietic stem cell transplant recipients, but the antigens
recognized by the infused CD4
+
T cells have remained
unknown. Here, we describe a simple procedure that permits the
identification of viral T-helper (T
H
)-cell antigens and
epitopes. This direct antigen identification method is based on the
random expression of viral polypeptides fused to chloramphenicol
acetyltransferase (CAT) in bacteria, which are subsequently fed to
major histocompatibility complex class II
+
antigen-presenting cells and probed with antigen-specific T cells. The
fusion of antigenic fragments to CAT offers several advantages. First,
chloramphenicol treatment allows the selection of bacteria expressing
antigen-CAT fusion proteins in frame, which greatly reduces the number
of colonies to be screened. Second, antigenic fragments fused to CAT
are expressed at high levels, even when derived from proteins that are
toxic to bacteria. Third, the uniformly high expression level of
antigen-CAT fusion proteins permits the establishment of large and
representative pool sizes. Finally, antigen identification does not
require knowledge of the restriction element and often leads directly
to the identification of the T-cell epitope. Using this approach, the
BALF4 and BNRF1 proteins were identified as targets of the EBV-specific
T-helper-cell response, demonstrating that lytic cycle antigens are a
relevant component of the EBV-specific T
H
-cell
response.
Human Cytomegalovirus IE86 Attenuates Virus- and Tumor Necrosis Factor Alpha-Induced NFκB-Dependent Gene Expression
Authors: Taylor R. Travis , Bresnahan Wade A.
Published: October 2006

Human cytomegalovirus (HCMV) infection regulates a number of genes involved in the host antiviral response. We have previously reported that HCMV attenuates the expression of beta interferon (IFN-β) and a number of proinflammatory chemokines, and this attenuation is mediated by the HCMV immediate-early protein IE86. The present study seeks to identify the mechanism by which IE86 blocks IFN-β expression. We demonstrate that the induction of IFN-β during HCMV infection requires the activation of both the IRF-3 and the NFκB pathways. Therefore, IE86 may target either pathway to block IFN-β expression. Our results show that IE86 does not block IRF-3 phosphorylation, dimerization, nuclear translocation, or target gene expression. However, using gel shift analysis, we demonstrate that IE86 efficiently inhibits virus-induced binding of NFκB to the IFN-β promoter, resulting in attenuation of IFN-β and NFκB-dependent gene expression. Furthermore, IE86 expression inhibits tumor necrosis factor alpha-induced NFκB DNA binding and target gene expression. Together, these results identify IE86 as a NFκB antagonist, which results in the suppression of NFκB-dependent cytokine and chemokine gene expression.
Methylation Status of theEpstein-Barr Virus (EBV) BamHI W Latent Cycle Promoter and Promoter Activity: Analysis with Novel EBV-Positive Burkitt and LymphoblastoidCell Lines
Authors: Hutchings Isabel A. , Tierney Rosemary J. , Kelly Gemma L. , Stylianou Julianna , Rickinson Alan B. , Bell Andrew I.
Published: October 2006

The
Epstein-Barr virus (EBV) latent cycle promoter Wp, present in each
tandemly arrayed copy of the BamHI W region in the EBV genome, drives
expression of the EB viral nuclear antigens (EBNAs) at the initiation
of virus-induced B-cell transformation. Thereafter, an alternative EBNA
promoter, Cp, becomes dominant, Wp activity declines dramatically, and
bisulfite sequencing of EBV-transformed lymphoblastoid cell lines
(LCLs) shows extensive Wp methylation. Despite this, Wp is never
completely silenced in LCLs. Here, using a combination of bisulfite
sequencing and methylation-specific PCR, we show that in standard LCLs
transformed with wild-type EBV isolates, some Wp copies always remain
unmethylated, and in LCLs transformed with a recombinant EBV carrying
just two BamHI W copies, Wp is completely unmethylated. Furthermore, we
have analyzed rare LCLs, recently established using wild-type EBV
isolates, and rare Burkitt lymphoma (BL) cell clones, recently
established from tumors carrying EBNA2-deleted EBV genomes, which
express EBNAs exclusively from Wp-initiated transcripts. Here, in sharp
contrast to standard LCL and BL lines, all resident copies of Wp appear
to be predominantly hypomethylated. Thus, studies of B cells with
atypical patterns of Wp usage emphasize the strong correlation between
the presence of unmethylated Wp sequences and promoter
activity.
LMP1 Transmembrane Domain 1 and 2 (TM1-2) FWLY Mediates Intermolecular Interactions with TM3-6 To Activate NF-κB
Authors: Soni Vishal , Yasui Teruhito , Cahir-McFarland Ellen , Kieff Elliott
Published: October 2006

The Epstein-Barr virus oncoprotein LMP1 has six transmembrane domains (TMs) that enable intermolecular aggregation and constitutive signaling through two C-terminal cytosolic domains. Expression of both TMs 1 and 2 without the C terminus (TM1-2ΔC) and TMs 3 to 6 fused to the C terminus (TM3-6) results in partial association, which is substantially decreased by TM1 F
38
WLY
41
mutation to A
38
ALA
41
. We now investigate whether TM1-2ΔC can functionally interact with TM3-6. TM1-2ΔC induced TM3-6 to mediate NF-κB activation at 59% of LMP1 levels, and the effect was dependent on TM1-2 F
38
WLY
41
. TM1-2ΔC even induced TM3-4 C terminus-mediated NF-κB activation to 44% of LMP1 levels. Surprisingly, this effect was TM1 F
38
WLY
41
independent, indicative of a role for TMs 5 and 6 in TM1 F
38
WLY
41
effects. TM3 W
98
was also important for TM1-2ΔC induction of TM3-6-mediated NF-κB activation, for association, and for TM1 F
38
WLY
41
dependence on C-terminal NF-κB activation. These data support models in which the TM1 F
38
WLY
41
effects are at least partially dependent on TM3 W
98
and a residue(s) in TMs 5 and 6.
CD8 T Cells Control Cytomegalovirus Latency by Epitope-Specific Sensing of Transcriptional Reactivation
Authors: Simon Christian O. , Holtappels Rafaela , Tervo Hanna-Mari , Böhm Verena , Däubner Torsten , Oehrlein-Karpi Silke A. , Kühnapfel Birgit , Renzaho Angélique , Strand Dennis , Podlech Jürgen , Reddehase Matthias J. , Grzimek Natascha K. A.
Published: October 2006

During
murine cytomegalovirus (mCMV) latency in the lungs, most of the viral
genomes are transcriptionally silent at the major immediate-early
locus, but rare and stochastic episodes of desilencing lead to the
expression of IE1 transcripts. This low-frequency but perpetual
expression is accompanied by an activation of lung-resident
effector-memory CD8 T cells specific for the antigenic peptide
168-YPHFMPTNL-176, which is derivedfrom the IE1 protein. These molecular and immunological findings were
combined in the “silencing/desilencing and immune sensing
hypothesis” of cytomegalovirus latency and reactivation. This
hypothesis proposes that IE1 gene expression proceeds to cell surface
presentation of the IE1 peptide by the major histocompatibility complex
(MHC) class I molecule L
d
and that its recognition by CD8 T
cells terminates virus reactivation. Here we provide experimental
evidence in support of this hypothesis. We generated mutant virus
mCMV-IE1-L176A, in which the antigenic IE1 peptide is functionally
deleted by a point mutation of the C-terminal MHC class I anchor
residue Leu into Ala. Two revertant viruses, mCMV-IE1-A176L and the
wobble nucleotide-marked mCMV-IE1-A176L*, in which Leu is restored by
back-mutation of Ala codon GCA into Leu codons CTA and CTT,
respectively, were constructed. Pulmonary latency of the mutant virus
was found to be associated with an increased prevalence of IE1
transcription and with events of IE3 transactivator splicing. In
conclusion, IE1-specific CD8 T cells recognize and terminate virus
reactivation in vivo at the first opportunity in the reactivated gene
expression program. The perpetual gene expression and antigen
presentation might represent the driving molecular force in
CMV-associated
immunosenescence.