bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of Novel Subgenomic RNAs and Noncanonical Transcription Initiation Signals of Severe Acute Respiratory Syndrome Coronavirus
Authors: Hussain Snawar , Pan Ji'an , Chen Yu , Yang Yalin , Xu Jing , Peng Yu , Wu Ying , Li Zhaoyang , Zhu Ying , Tien Po , Guo Deyin 
Published: April 2005

ABSTRACT
The expression of the genomic information of severe acute respiratory syndrome coronavirus (SARS CoV) involves synthesis of a nested set of subgenomic RNAs (sgRNAs) by discontinuous transcription. In SARS CoV-infected cells, 10 sgRNAs, including 2 novel ones, were identified, which were predicted to be functional in the expression of 12 open reading frames located in the 3′ one-third of the genome. Surprisingly, one new sgRNA could lead to production of a truncated spike protein. Sequence analysis of the leader-body fusion sites of each sgRNA showed that the junction sequences and the corresponding transcription-regulatory sequence (TRS) are unique for each species of sgRNA and are consistent after virus passages. For the two novel sgRNAs, each used a variant of the TRS that has one nucleotide mismatch in the conserved hexanucleotide core (ACGAAC) in the TRS. Coexistence of both plus and minus strands of SARS CoV sgRNAs and evidence for derivation of the sgRNA core sequence from the body core sequence favor the model of discontinuous transcription during minus-strand synthesis. Moreover, one rare species of sgRNA has the junction sequence AAA, indicating that its transcription could result from a noncanonical transcription signal. Taken together, these results provide more insight into the molecular mechanisms of genome expression and subgenomic transcription of SARS CoV.
Structure and Assembly of a
T
=1 Virus-Like Particle in BK Polyomavirus
Authors: Nilsson Josefina , Miyazaki Naoyuki , Xing Li , Wu Bomu , Hammar Lena , Li Tian Cheng , Takeda Naokazu , Miyamura Tatsuo , Cheng R. Holland 
Published: April 2005

ABSTRACT

In polyomaviruses the pentameric capsomers are interlinked by the long C-terminal arm of the structural protein VP1. The
T
=7 icosahedral structure of these viruses is possible due to an intriguing adaptability of this linker arm to the different local environments in the capsid. To explore the assembly process, we have compared the structure of two virus-like particles (VLPs) formed, as we found, in a calcium-dependent manner by the VP1 protein of human polyomavirus BK. The structures were determined using electron cryomicroscopy (cryo-EM), and the three-dimensional reconstructions were interpreted by atomic modeling. In the small VP1 particle, 26.4 nm in diameter, the pentameric capsomers form an icosahedral
T
=1 surface lattice with meeting densities at the threefold axes that interlinked three capsomers. In the larger particle, 50.6 nm in diameter, the capsomers form a
T
=7 icosahedral shell with three unique contacts. A folding model of the BKV VP1 protein was obtained by alignment with the VP1 protein of simian virus 40 (SV40). The model fitted well into the cryo-EM density of the
T
=7 particle. However, residues 297 to 362 of the C-terminal arm had to be remodeled to accommodate the higher curvature of the
T
=1 particle. The loops, before and after the C-terminal short helix, were shown to provide the hinges that allowed curvature variation in the particle shell. The meeting densities seen at the threefold axes in the
T
=1 particle were consistent with the triple-helix interlinking contact at the local threefold axes in the
T
=7 structure.
Replication-Competent Adenovirus Formation in 293 Cells: the Recombination-Based Rate Is Influenced by Structure and Location of the Transgene Cassette and Not Increased by Overproduction of HsRad51, Rad51-Interacting, or E2F Family Proteins
Authors: Duigou Gregory J. , Young C. S. H. 
Published: April 2005

ABSTRACT

Propagation of E1 region replacement adenovirus vectors in 293 cells results in the rare appearance of replication-competent adenovirus (RCA). The RCA genome contains E1 DNA acquired from the 293 cellular genome. The Luria-Delbrück fluctuation test was adapted to measure RCA formation rates. To test if structure affected rate, we measured rates during the production of adenovirus vectors with genomes containing three different expression cassette arrangements. The vectors had different extents of sequence identity with integrated Ad5 DNA of 293 cells and had different distributions of identity flanking the expression cassettes. Empty cassette vector RCA rates ranged from 2.5 × 10
−8
to 5.6 × 10
−10
. The extent of sequence identity was not an accurate RCA rate predictor. The vector with the highest RCA rate also had the least overall sequence identity. To define factors controlling RCA generation, adenovirus vectors expressing E2F family proteins, known to modulate recombination gene expression, and overexpressing the human Rad51 recombination protein were analyzed. Compared to their corresponding empty vectors, RCA rates were not increased but were slightly decreased. Initial results suggested expression cassette orientation and/or transcription direction as potential RCA rate modifiers. Testing adenovirus vectors with identical transgene cassettes oriented in opposite directions suggested that transcription direction was not the basis of these rate differences. Thus, the overall structure and location of the transgene cassette had the largest effect on RCA rate. The RCA fluctuation test should be useful for investigators who require accurate measurements of targeted recombination and the probability of RCA formation during stock production.
Linked Tumor-Selective Virus Replication and Transgene Expression from E3-Containing Oncolytic Adenoviruses
Authors: Zhu Mingzhu , Bristol J. Andrew , Xie Yuefeng , Mina Mervat , Ji Hong , Forry-Schaudies Suzanne , Ennist David L. 
Published: April 2005

ABSTRACT
Historically, the adenoviral E3 region was found to be nonessential for viral replication in vitro. In addition, adenoviruses whose genome was more than approximately 105% the size of the native genome were inefficiently packaged. These profound observations were used experimentally to insert transgenes into the adenoviral backbone. More recently, however, the reintroduction of the E3 region into oncolytic adenoviruses has been found to positively influence antitumor efficacy in preclinical models and clinical trials. In the studies reported here, the granulocyte-macrophage colony-stimulating factor (GM-CSF) cDNA sequence has been substituted for the E3-gp19 gene in oncolytic adenoviruses that otherwise retained the E3 region. Five viruses that differed slightly in the method of transgene insertion were generated and compared to Ar6pAE2fGmF (E2F/GM/ΔE3), a previously described E3-deleted oncolytic adenovirus encoding GM-CSF. In all of the viruses, the human E2F-1 promoter regulated E1A expression and GM-CSF expression was under the control of the adenoviral E3 promoter and the packaging signal was relocated immediately upstream from the right terminal repeat. The E3-gp19-deleted viruses had similar cytolytic properties, as measured in vitro by cytotoxicity assays, but differed markedly in their capacity to express and secrete GM-CSF. Ar15pAE2fGmF (E2F/GM/E3b), the virus that produced the highest levels of GM-CSF and retained the native GM-CSF leader sequence, was selected for further analysis. The E2F/GM/E3b and E2F/GM/ΔE3 viruses exhibited similar cytotoxic activity and GM-CSF production in several tumor cell lines in vitro. However, when compared in vivo in nude mouse xenograft tumor models, E2F/GM/E3b spread through tumors to a greater extent, resulted in higher peak GM-CSF and total exposure levels in both tumor and serum, and was more efficacious than the E3-deleted virus. Using the matched WI-38 (parental) and WI-38-VA13 (simian virus 40 large T antigen transformed) cell pair, GM-CSF was shown to be selectively produced in cells expressing high levels of E2F, indicating that the tumor-selective E2F promoter controlled E1A and GM-CSF expression.
Inhibition of Toll-Like Receptor 7- and 9-Mediated Alpha/Beta Interferon Production in Human Plasmacytoid Dendritic Cells by Respiratory Syncytial Virus and Measles Virus
Authors: Schlender Jörg , Hornung Veit , Finke Stefan , Günthner-Biller Margit , Marozin Sabrina , Brzózka Krzysztof , Moghim Sharareh , Endres Stefan , Hartmann Gunther , Conzelmann Karl-Klaus 
Published: April 2005

ABSTRACT

Human plasmacytoid dendritic cells (PDC) are key sentinels alerting both innate and adaptive immune responses through production of huge amounts of alpha/beta interferon (IFN). IFN induction in PDC is triggered by outside-in signal transduction pathways through Toll-like receptor 7 (TLR7) and TLR9 as well as by recognition of cytosolic virus-specific patterns. TLR7 and TLR9 ligands include single-stranded RNA and CpG-rich DNA, respectively, as well as synthetic derivatives thereof which are being evaluated as therapeutic immune modulators promoting Th1 immune responses. Here, we identify the first viruses able to block IFN production by PDC. Both TLR-dependent and -independent IFN responses are abolished in human PDC infected with clinical isolates of respiratory syncytial virus (RSV), RSV strain A2, and measles virus Schwarz, in contrast to RSV strain Long, which we previously identified as a potent IFN inducer in human PDC (Hornung et al., J. Immunol.
173:
5935-5943, 2004). Notably, IFN synthesis of PDC activated by the TLR7 and TLR9 agonists resiquimod (R848) and CpG oligodeoxynucleotide 2216 is switched off by subsequent infection by RSV A2 and measles virus. The capacity of RSV and measles virus of human PDC to shut down IFN production should contribute to the characteristic features of these viruses, such as Th2-biased immune pathology, immune suppression, and superinfection.
Novel Macula-Like Virus Identified in
Bombyx mori
Cultured Cells
Authors: Katsuma Susumu , Tanaka Shinichiro , Omuro Naoko , Takabuchi Lisa , Daimon Takaaki , Imanishi Shigeo , Yamashita Shuichi , Iwanaga Masashi , Mita Kazuei , Maeda Susumu , Kobayashi Masahiko , Shimada Toru 
Published: April 2005

ABSTRACT

We identified a novel, 6,513-bp-long RNA, termed
Bombyx mori
macula-like latent virus (BmMLV) RNA, which abundantly expressed in
B. mori
cultured BmN cells. BmMLV RNA potentially encodes two proteins, putative RNA replicase and coat protein, which share structural features and sequence similarities with those of a plant RNA virus, the genus
Maculavirus
. Northern blot analysis showed that two transcripts were expressed in BmN cells: a 6.5-kb-long RNA, which contains both putative RNA replicase and coat protein genes, and a 1.2-kb-long RNA, which contains only a coat protein gene. Southern blot analysis showed that BmMLV RNA is not carried by the
B. mori
genome. RT-PCR analysis also revealed the presence of BmMLV RNA in several
B. mori
cell lines other than BmN cells, suggesting that BmMLV RNA latently exists in
B. mori
cultured cells. Infection studies showed that BmMLV virions were able to infect BmMLV-negative
Spodoptera frugiperda
Sf-9 cells and
B. mori
larvae. Electron microscopy and Northern blot analysis of a purified BmMLV revealed that isometric virions appear to be 28 to 30 nm in diameter and contain a 6.5-kb genomic RNA. These results showed that BmMLV is a novel macula-like virus infectious to and replicable in
B. mori
-derived cells.
Macrophage Inflammatory Protein 1α Inhibits Postentry Steps of Human Immunodeficiency Virus Type 1 Infection via Suppression of Intracellular Cyclic AMP
Authors: Amella Carol-Ann , Sherry Barbara , Shepp David H. , Schmidtmayerova Helena 
Published: April 2005

ABSTRACTPrimary isolates of human immunodeficiency virus type 1 (HIV-1) predominantly use chemokine receptor CCR5 to enter target cells. The natural ligands of CCR5, the β-chemokines macrophage inflammatory protein 1α (MIP-1α), MIP-1β, and RANTES, interfere with HIV-1 binding to CCR5 receptors and decrease the amount of virions entering cells. Although the inhibition of HIV-1 entry by β-chemokines is well documented, their effects on postentry steps of the viral life cycle and on host cell components that control the outcome of infection after viral entry are not well defined. Here, we show that all three β-chemokines, and MIP-1α in particular, inhibit postentry steps of the HIV-1 life cycle in primary lymphocytes, presumably via suppression of intracellular levels of cyclic AMP (cAMP). Productive HIV-1 infection of primary lymphocytes requires cellular activation. Cell activation increases intracellular cAMP, which is required for efficient synthesis of proviral DNA during early steps of viral infection. Binding of MIP-1α to cognate receptors decreases activation-induced intracellular cAMP levels through the activation of inhibitory G proteins. Furthermore, inhibition of one of the downstream targets of cAMP, cAMP-dependent PKA, significantly inhibits synthesis of HIV-1-specific DNA without affecting virus entry. These data reveal that β-chemokine-mediated inhibition of virus replication in primary lymphocytes combines inhibitory effects at the entry and postentry levels and imply the involvement of β-chemokine-induced signaling in postentry inhibition of HIV-1 infection.
Cell Cycle Arrest in G
2
/M Promotes Early Steps of Infection by Human Immunodeficiency Virus
Authors: Groschel Bettina , Bushman Frederic 
Published: April 2005

ABSTRACT

We have identified four small molecules that boost transduction of cells by human immunodeficiency virus (HIV) and investigated their mechanism of action. These molecules include etoposide and camptothecin, which induce DNA damage by inhibiting religation of cleaved topoisomerase-DNA complexes, taxol, which interferes with the function of microtubules, and aphidicolin, which inhibits DNA polymerases. All four compounds arrest the cell cycle at G
2
/M, though in addition high concentrations of aphidicolin arrest in G
1
. We find that early events of HIV replication, including synthesis of late reverse transcription products, two-long terminal repeat circles, and integrated proviruses, were increased after treatment of cells with concentrations of each compound that arrested in G
2
/M. Stimulation was seen for both transformed cell lines (293T and HeLa cells) and primary cells (IMR90 lung fibroblasts). Arrest in G
1
with high concentrations of aphidicolin boosted transduction, though not much as with lower concentrations that arrested in G
2
/M. Arrest of IMR90 cells in G
1
by serum starvation and contact inhibition reduced transduction. Previously, the proteasome inhibitor MG132 was reported to increase HIV infection—here we investigated the effects of combinations of the cell cycle inhibitors with MG132 and obtained data suggesting that MG132 may also boost transduction by causing G
2
/M cell cycle arrest. These data document that cell cycle arrest in G
2
/M boosts the early steps of HIV infection and suggests methods for increasing transduction with HIV-based vectors.
Evaluation of the Conformational Switch Model for Alfalfa Mosaic Virus RNA Replication
Authors: Petrillo Jessica E. , Rocheleau Gail , Kelley-Clarke Brenna , Gehrke Lee 
Published: April 2005

ABSTRACT
Key elements of the conformational switch model describing regulation of alfalfa mosaic virus (AMV) replication (R. C. Olsthoorn, S. Mertens, F. T. Brederode, and J. F. Bol, EMBO J. 18:4856-4864, 1999) have been tested using biochemical assays and functional studies in nontransgenic protoplasts. Although comparative sequence analysis suggests that the 3′ untranslated regions of AMV and ilarvirus RNAs have the potential to fold into pseudoknots, we were unable to confirm that a proposed pseudoknot forms or has a functional role in regulating coat protein-RNA binding or viral RNA replication. Published work has suggested that the pseudoknot is part of a tRNA-like structure (TLS); however, we argue that the canonical sequence and functional features that define the TLS are absent. We suggest here that the absence of the TLS correlates directly with the distinctive requirement for coat protein to activate replication in these viruses. Experimental data are evidence that elevated magnesium concentrations proposed to stabilize the pseudoknot structure do not block coat protein binding. Additionally, covarying nucleotide changes proposed to reestablish pseudoknot pairings do not rescue replication. Furthermore, as described in the accompanying paper (L. M. Guogas, S. M. Laforest, and L. Gehrke, J. Virol. 79:5752-5761, 2005), coat protein is not, by definition, inhibitory to minus-strand RNA synthesis. Rather, the activation of viral RNA replication by coat protein is shown to be concentration dependent. We describe the 3′ organization model as an alternate model of AMV replication that offers an improved fit to the available data.
Rhesus Cytomegalovirus Contains Functional Homologues of US2, US3, US6, and US11
Authors: Pande Nupur T. , Powers Colin , Ahn Kwangseog , Früh Klaus 
Published: April 2005

ABSTRACTHuman cytomegalovirus (HCMV) is a paradigm for mechanisms subverting antigen presentation by major histocompatibility complex (MHC) molecules. Due to its limited host range, HCMV cannot be studied in animals. Thus, the in vivo importance of inhibiting antigen presentation for the establishment and maintenance of infection with HCMV is unknown. Rhesus cytomegalovirus (RhCMV) is an emerging animal model that shares many of the features of HCMV infection. The recent completion of the genomic sequence of RhCMV revealed a significant degree of homology to HCMV. Strikingly, RhCMV contains several genes with low homology to the HCMV US6 gene family of inhibitors of the MHC I antigen presentation pathway. Here, we examine whether the RhCMV US6 homologues (open reading frames Rh182, -184, -185, -186, -187, and -189) interfere with the MHC I antigen-processing pathway. We demonstrate that Rh182 and Rh189 function similarly to HCMV US2 and US11, respectively, mediating the proteasomal degradation of newly synthesized MHC I. The US3 homologue, Rh184, delayed MHC I maturation. Unlike US3, MHC I molecules eventually escaped retention by Rh184, so that steady-state surface levels of MHC I remained unchanged. Rh185 acted similarly to US6 and inhibited peptide transport by TAP and, consequently, peptide loading of MHC I molecules. Thus, despite relatively low sequence conservation, US6 family-related genes in RhCMV are functionally closely related to the conserved structural features of HCMV immunomodulators. The conservation of these mechanisms implies their importance for immune evasion in vivo, a question that can now be addressed experimentally.
ULTRASONOGRAPHIC DIAGNOSIS IN THE EVALUATION OF SHOULDER PAIN
Authors: Blankstein Alexander , Mirovski Yigal , Givon Uri , Chechick Aharon , Adunsky Abraham , Ganel Avraham 
Published: April 2005

The benefit of adding sonography to the examination of patients suffering from shoulder pain, was examined in this retrospective evaluation. A patient suffering from shoulder pain consults generally his family physician, who sends his patient first to a radiography and not to sonography. In 75% of the patients examined by sonography, shoulder lesions were diagnosed, demonstrated in only 33% of radiographs. Most patients had more than one lesion: 38% had partial or full-thickness tears of the supraspinatus tendon, about one third bicipital tendonitis, 18% irregularities of the humeral greater tuberosity, 10% subdeltoid bursitis and about 10% calcified tendonitis. In about a quarter of the patients, no pathology was found. Many patients had more than one lesion. In conclusion, clinicians encountering shoulder pain should not be content with conventional radiography and are encouraged to make sonographic examinations part of the early diagnostic protocol.
The Role of Boundary Conditions in AMIP-2 Simulations of the NAO
Authors: Cohen Judah , Frei Allan , Rosen Richard D. 
Published: April 2005

Abstract
The simulated North Atlantic Oscillation (NAO) teleconnection patterns and their interannual variability are evaluated from a suite of atmospheric models participating in the second phase of the Atmospheric Model Intercomparison Project (AMIP-2). In general the models simulate the observed spatial pattern well, although there are important differences among models. The NAO response to interannual variations in sea surface temperature (SST) and snow-cover boundary forcings are also evaluated. The simulated NAO indices are not correlated with the observed NAO index, despite being forced with observed SSTs, indicating that SSTs are not driving NAO variability in the models. Similarly, although a number of studies have identified a link between Eurasian snow extent and the phase of the NAO, no such link is apparent in the AMIP-2 results. It appears that, within the framework of the AMIP-2 experiments, the NAO is an internal mode of atmospheric variability and that impacts of SSTs and Eurasian snow cover on the phase of the NAO are not discernable. However, these conclusions do not necessarily apply to decadal-scale and longer variability or to coupled atmosphere–ocean models.
Ocean Dynamics and the Nature of Air–Sea Interactions over the North Atlantic at Decadal Time Scales
Authors: Park W. , Latif M. 
Published: April 2005

Abstract
The dependence of the air–sea interactions over the North Atlantic on the ocean dynamics is explored by analyzing multicentury integrations with two different coupled ocean–atmosphere models. One is a coupled general circulation model (CGCM), in which both the atmospheric and the oceanic components are represented by general circulation models (GCMs). The second coupled model employs the same atmospheric GCM, but the oceanic GCM is replaced by a fixed-depth mixed layer model, so that variations of the ocean dynamics are excluded. The coupled model including active ocean dynamics simulates strong multidecadal variability in the sea surface temperature (SST) of the North Atlantic, with a monopolar spatial structure. In contrast, the coupled model that employs an oceanic mixed layer model and thus does not carry active ocean dynamics simulates a tripolar SST anomaly pattern at decadal time scales. The tripolar SST anomaly pattern is characterized by strong horizontal gradients and is by definition the result of the action of surface heat flux anomalies on the oceanic mixed layer.
The differences in the spatial structures of the dominant decadal SST anomaly patterns yield rather different atmospheric responses. While the response to the monopolar SST anomaly pattern is shallow and thermal, the response to the tripolar SST anomaly pattern involves changes in the transient eddy statistics. The latter can be explained by the strong horizontal SST gradients that affect the surface baroclinicity, which in turn affects the growth rate of the transient eddies. The differences in the atmospheric response characteristics yield completely different response patterns. In the coupled run with active ocean dynamics, the sea level pressure (SLP) anomalies exhibit a rather homogeneous pattern that resembles somewhat the East Atlantic Pattern (EAP), while a dipolar (North Atlantic Oscillation) NAO-like SLP anomaly pattern is simulated in the coupled run without active ocean dynamics.