bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression of Respiratory Syncytial Virus-Induced Chemokine Gene Networks in Lower Airway Epithelial Cells Revealed by cDNA Microarrays
Authors: Zhang Yuhong , Luxon Bruce A. , Casola Antonella , Garofalo Roberto P. , Jamaluddin Mohammad , Brasier Allan R. 
Published: July 2002

ABSTRACTTheParamyxovirusrespiratory syncytial virus (RSV) is the primary etiologic agent of serious epidemic lower respiratory tract disease in infants, immunosuppressed patients, and the elderly. Lower tract infection with RSV is characterized by a pronounced peribronchial mononuclear infiltrate, with eosinophilic and basophilic degranulation. Because RSV replication is restricted to airway epithelial cells, where RSV replication induces potent expression of chemokines, the epithelium is postulated to be a primary initiator of pulmonary inflammation in RSV infection. The spectrum of RSV-induced chemokines expressed by alveolar epithelial cells has not been fully investigated. In this report, we profile the kinetics and patterns of chemokine expression in RSV-infected lower airway epithelial cells (A549 and SAE). In A549 cells, membrane-based cDNA macroarrays and high-density oligonucleotide probe-based microarrays identified inducible expression of CC (I-309, Exodus-1, TARC, RANTES, MCP-1, MDC, and MIP-1α and -1β), CXC (GRO-α, -β, and -γ, ENA-78, interleukin-8 [IL-8], and I-TAC), and CX3C (Fractalkine) chemokines. Chemokines not previously known to be expressed by RSV-infected cells were independently confirmed by multiprobe RNase protection assay, Northern blotting, and reverse transcription-PCR. High-density microarrays performed on SAE cells confirmed a similar pattern of RSV-inducible expression of CC chemokines (Exodus-1, RANTES, and MIP-1α and -1β), CXC chemokines (I-TAC, GRO-α, -β, and -γ, and IL-8), and Fractalkine. In contrast, TARC, MCP-1, and MDC were not induced, suggesting the existence of distinct genetic responses for different types of airway-derived epithelial cells. Hierarchical clustering by agglomerative nesting and principal-component analyses were performed on A549-expressed chemokines; these analyses indicated that RSV-inducible chemokines are ordered into three related expression groups. These data profile the temporal changes in expression by RSV-infected lower airway epithelial cells of chemokines, chemotactic proteins which may be responsible for the complex cellular infiltrate in virus-induced respiratory inflammation.
Fv-4: Identification of the Defect in Env and the Mechanism of Resistance to Ecotropic Murine Leukemia Virus
Authors: Taylor Gwen M. , Gao Yi , Sanders David Avram 
Published: July 2002

ABSTRACT

Mice expressing the
Fv-4
gene are resistant to infection by ecotropic murine leukemia viruses (MuLVs). The
Fv-4
gene encodes an envelope (Env) protein whose putative receptor-binding domain resembles that of ecotropic MuLV Env protein. Resistance to ecotropic MuLVs appears to result from viral interference involving binding of the endogenously expressed
Fv-4 env
-encoded protein to the ecotropic receptor, although the immune system also plays a role in resistance. The
Fv-4 env
-encoded protein is processed normally and can be incorporated into virus particles but is unable to promote viral entry. Among the many sequence variations between the transmembrane (TM) subunit of the
Fv-4 env
-encoded protein and the TM subunits of other MuLV Env proteins, there is a substitution of an arginine residue in the
Fv-4 env
-encoded protein for a glycine residue (gly-491 in Moloney MuLV Env) that is otherwise conserved in all of the other MuLVs. This residue is present in the MuLV TM fusion peptide sequence. In this study, gly-491 of Moloney MuLV Env has been replaced with other residues and a mutant Env bearing a substitution for gly-487 was also created. G491R recapitulates the
Fv-4
Env phenotype in cell culture, indicating that this substitution is sufficient for creation of an Env protein that can establish the interference-mediated resistance to ecotropic viruses produced by the
Fv-4
gene. Analysis of the mutant MuLV Env proteins also has implications for an understanding of the role of conserved glycine residues in fusion peptides and for the engineering of organismal resistance to retroviruses.
Expression of Endogenous Betaretroviruses in the Ovine Uterus: Effects of Neonatal Age, Estrous Cycle, Pregnancy, and Progesterone
Authors: Palmarini Massimo , Gray C. Allison , Carpenter Karen , Fan Hung , Bazer Fuller W. , Spencer Thomas E. 
Published: July 2002

ABSTRACT
The ovine genome contains 15 to 20 copies of endogenous retroviruses (enJSRVs) highly related to the oncogenic jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus. enJSRVs are highly expressed in the endometrial lumenal epithelia (LE) and glandular epithelia (GE) of the ovine uterus. The effects of neonatal age, estrous cycle, pregnancy, and progesterone on expression of enJSRVs in the ovine uterus were determined. Expression of enJSRV RNAs was absent from the uterus of ewes at birth, but enJSRV RNAs were expressed specifically in the LE and developing GE from postnatal day (PND) 7 to PND 56. In adult ewes, enJSRV RNAs were detected only in the epithelia of the uterine endometrium, as well as epithelia of the oviduct, cervix, and vagina. In cyclic ewes, endometrial enJSRV RNA abundance was lowest on day 1, increased 12-fold between days 1 and 13, and then decreased to day 15. In pregnant ewes, levels of endometrial enJSRV RNAs were high on day 11, increased to day 13, and then decreased to day 19. In day 17 and 19 conceptuses, enJSRV RNAs were also detected in binucleate cells of the trophectoderm. Immunoreactive JSRV capsid and envelope proteins were detected in the endometrial LE and GE, as well as in the binucleate cells of the conceptus. In transfection assays utilizing ovine endometrial LE cells, progesterone increased transcriptional activity of several enJSRV long terminal repeats. Collectively, these results indicate that transcription of enJSRVs in the endometrial epithelia of the ovine uterus is increased by progesterone and might support a role for enJSRVs in conceptus-endometrium interactions during the peri-implantation period and early placental morphogenesis.
Base-Pair Substitutions in Avian Sarcoma Virus U5 and U3 Long Terminal Repeat Sequences Alter the Process of DNA Integration In Vitro
Authors: Hindmarsh Patrick , Johnson Michael , Reeves Ray , Leis Jonathan 
Published: July 2002

ABSTRACT

We have described a reconstituted avian sarcoma virus (ASV) concerted DNA integration system with specially designed mini-donor DNA containing a
supF
transcription unit, a supercoiled plasmid acceptor, purified bacterially expressed ASV integrase (IN), and human high-mobility-group protein I(Y). Integration in this system is dependent upon the mini-donor DNA having IN recognition sequences at both ends and upon both ends of the same donor integrating into the acceptor DNA. The integrated DNA product exhibits all of the features associated with integration of viral DNA in vivo (P. Hindmarsh et al., J. Virol., 73:2994–3003, 1999). Individual integrants are isolated from bacteria containing drug-resistant markers with amber mutations. This system was used to evaluate the importance of sequences in the terminal U5 and U3 long terminal repeats at positions 5 and/or 6, adjacent to the conserved CA dinucleotide. Base-pair substitutions introduced at these positions in U5 result in significant reductions in recovered integrants from bacteria, due to increases in one-ended insertion events. Among the recovered integrants from reactions with mutated U5 but not U3 IN recognition sequences were products that contain large deletions in the acceptor DNA. Base-pair substitutions at positions 5 and 6 in U3 mostly reduce the efficiency of integration of the modified donor. Together, these results indicate that sequences directly 5′ to the conserved CA dinucleotide are very important for the process of concerted DNA integration. Furthermore, IN interacts with U3 and U5 termini differently, and aberrant end-processing events leading to nonconcerted DNA integration are more common in U5 than in U3.
Reverse Genetics System for Uukuniemi Virus (
Bunyaviridae
): RNA Polymerase I-Catalyzed Expression of Chimeric Viral RNAs
Authors: Flick Ramon , Pettersson Ralf F. 
Published: July 2002

ABSTRACT

We describe here the development of a reverse genetics system for the phlebovirus Uukuniemi virus, a member of the
Bunyaviridae
family, by using RNA polymerase I (pol I)-mediated transcription. Complementary DNAs containing the coding sequence for either chloramphenicol acetyltransferase (CAT) or green fluorescent protein (GFP) (both in antisense orientation) were flanked by the 5′- and 3′-terminal untranslated regions of the Uukuniemi virus sense or complementary RNA derived from the medium-sized (M) RNA segment. This chimeric cDNA (pol I expression cassette) was cloned between the murine pol I promoter and terminator and the plasmid transfected into BHK-21 cells. When such cells were either superinfected with Uukuniemi virus or cotransfected with expression plasmids encoding the L (RNA polymerase), N (nucleoprotein), and NSs (nonstructural protein) viral proteins, strong CAT activity or GFP expression was observed. CAT activity was consistently stronger in cells expressing L plus N than following superinfection. No activity was seen without superinfection, nor was activity detected when either the L or N expression plasmid was omitted. Omitting NSs expression had no effect on CAT activity or GFP expression, indicating that this protein is not needed for viral RNA replication or transcription. CAT activity could be serially passaged to fresh cultures by transferring medium from CAT-expressing cells, indicating that recombinant virus containing the reporter construct had been produced. In summary, we demonstrate that the RNA pol I system, originally developed for influenza virus, which replicates in the nucleus, has strong potential for the development of an efficient reverse genetics system also for
Bunyaviridae
members, which replicate in the cytoplasm.
Cellular Splicing Factor RAF-2p48/NPI-5/BAT1/UAP56 Interacts with the Influenza Virus Nucleoprotein and Enhances Viral RNA Synthesis
Authors: Momose Fumitaka , Basler Christopher F. , O'Neill Robert E. , Iwamatsu Akihiro , Palese Peter , Nagata Kyosuke 
Published: July 2002

ABSTRACT
Previous biochemical data identified a host cell fraction, designated RAF-2, which stimulated influenza virus RNA synthesis. A 48-kDa polypeptide (RAF-2p48), a cellular splicing factor belonging to the DEAD-box family of RNA-dependent ATPases previously designated BAT1 (also UAP56), has now been identified as essential for RAF-2 stimulatory activity. Additionally, RAF-2p48 was independently identified as an influenza virus nucleoprotein (NP)-interacting protein, NPI-5, in a yeast two-hybrid screen of a mammalian cDNA library. In vitro, RAF-2p48 interacted with free NP but not with NP bound to RNA, and the RAF-2p48–NP complex was dissociated following addition of free RNA. Furthermore, RAF-2p48 facilitated formation of the NP-RNA complexes that likely serve as templates for the viral RNA polymerase. RAF-2p48 was shown, in both in vitro binding assays and the yeast two-hybrid system, to bind to the amino-terminal region of NP, a domain essential for RNA binding. Together, these observations suggest that RAF-2p48 facilitates NP-RNA interaction, thus leading to enhanced influenza virus RNA synthesis.
Direct and Indirect Regulation of Cytokine and Cell Cycle Proteins by EBNA-2 during Epstein-Barr Virus Infection
Authors: Spender Lindsay C. , Cornish Georgina H. , Rowland Benjamin , Kempkes Bettina , Farrell Paul J. 
Published: July 2002

ABSTRACT
We have studied the pathways of regulation of cytokine and cell cycle control proteins during infection of human B lymphocytes by Epstein-Barr virus (EBV). Among 30 cytokine RNAs analyzed by the RNase protection assay, tumor necrosis factor alpha (TNF-α), granulocyte colony-stimulating factor, lymphotoxin (LT), and LTβ were found to be regulated within 20 h of EBV infection of primary B cells. Similar results were obtained using the estrogen-regulated EBNA-2 cell line EREB2.5, in which RNAs for LT and TNF-α were induced within 6 h of activation of EBNA-2. Expression of Notch also caused an induction of TNF-α RNA. The induction of TNF-α RNA by EBNA-2 was indirect, and constitutive expression of either LMP-1 or c-myc proteins did not substitute for EBNA-2 in induction of TNF-α RNA. Cyclin D2 is also an indirect target of EBNA-2-mediated transactivation. EBNA-2 was found to activate the cyclin D2 promoter in a transient-transfection assay. A mutant of EBNA-2 that does not bind RBP-Jκ retained some activity in this assay, and activation did not depend on the presence of B-cell-specific factors. Deletion analysis of the cyclin D2 promoter revealed that removal of sequences containing E-box c-myc consensus DNA binding sequences did not reduce EBNA-2-mediated activation of the cyclin D2 promoter in the transient-transfection assay. The results indicate that cytokines are an early target of EBNA-2 and that EBNA-2 can regulate cyclin D2 transcription in EBV-infected cells by mechanisms additional to the c-myc pathway.
Measles Viruses on Throat Swabs from Measles Patients Use Signaling Lymphocytic Activation Molecule (CDw150) but Not CD46 as a Cellular Receptor
Authors: Ono Nobuyuki , Tatsuo Hironobu , Hidaka Yasufumi , Aoki Tomonobu , Minagawa Hiroko , Yanagi Yusuke 
Published: July 2002

ABSTRACTBoth CD46 and signaling lymphocytic activation molecule (SLAM) have been shown to act as cellular receptors for measles virus (MV). The viruses on throat swabs from nine patients with measles in Japan were titrated on Vero cells stably expressing human SLAM. Samples from all but two patients produced numerous plaques on SLAM-expressing Vero cells, whereas none produced any plaques on Vero cells endogenously expressing CD46. The Edmonston strain of MV, which can use either CD46 or SLAM as a receptor, produced comparable titers on these two types of cells. The results strongly suggest that the viruses in the bodies of measles patients use SLAM but probably not CD46 as a cellular receptor.
Macaques with Rapid Disease Progression and Simian Immunodeficiency Virus Encephalitis Have a Unique Cytokine Profile in Peripheral Lymphoid Tissues
Authors: Orandle Marlene S. , Williams Kenneth C. , MacLean Andrew G. , Westmoreland Susan V. , Lackner Andrew A. 
Published: July 2002

ABSTRACTThe influence of host cytokine response on viral load, disease progression, and neurologic lesions was investigated in the simian immunodeficiency virus (SIV)-infected macaque model of AIDS. Cytokine gene expression (interleukin-1β [IL-1β], IL-2, IL-6, IL-10, gamma interferon [IFN-γ], and tumor necrosis factor alpha [TNF-α]) and viral loads were evaluated by semiquantitative reverse transcription-PCR in lymph nodes of 5 control animals and 28 animals infected with SIVmac251 at the terminal stages of AIDS. Infected animals showed higher expression of IFN-γ, IL-6, and IL-10 mRNAs compared with controls. Levels of all cytokines were comparable between animals with rapid (survival, <200 days) or slow/normal (survival, >200 days) disease progression. However, among rapid progressors, the eight animals with SIV encephalitis had a unique cytokine profile (increased IL-2, IL-6, and IFN-γ) that was associated with higher viral loads. These observations provide evidence that host cytokine responses may influence SIV neuropathogenesis independent of disease progression.
Ebola Virus VP40 Drives the Formation of Virus-Like Filamentous Particles Along with GP
Authors: Noda Takeshi , Sagara Hiroshi , Suzuki Emiko , Takada Ayato , Kida Hiroshi , Kawaoka Yoshihiro 
Published: July 2002

ABSTRACT
Using biochemical assays, it has been demonstrated that expression of Ebola virus VP40 alone in mammalian cells induced production of particles with a density similar to that of virions. To determine the morphological properties of these particles, cells expressing VP40 and the particles released from the cells were examined by electron microscopy. VP40 induced budding from the plasma membrane of filamentous particles, which differed in length but had uniform diameters of approximately 65 nm. When the Ebola virus glycoprotein (GP) responsible for receptor binding and membrane fusion was expressed in cells, we found pleomorphic particles budding from the plasma membrane. By contrast, when GP was coexpressed with VP40, GP was found on the filamentous particles induced by VP40. These results demonstrated the central role of VP40 in formation of the filamentous structure of Ebola virions and may suggest an interaction between VP40 and GP in morphogenesis.
Interaction of the Coronavirus Nucleoprotein with Nucleolar Antigens and the Host Cell
Authors: Chen Hongying , Wurm Torsten , Britton Paul , Brooks Gavin , Hiscox Julian A. 
Published: July 2002

ABSTRACTCoronavirus nucleoproteins (N proteins) localize to the cytoplasm and the nucleolus, a subnuclear structure, in both virus-infected primary cells and in cells transfected with plasmids that express N protein. The nucleolus is the site of ribosome biogenesis and sequesters cell cycle regulatory complexes. Two of the major components of the nucleolus are fibrillarin and nucleolin. These proteins are involved in nucleolar assembly and ribosome biogenesis and act as chaperones for the import of proteins into the nucleolus. We have found that fibrillarin is reorganized in primary cells infected with the avian coronavirus infectious bronchitis virus (IBV) and in continuous cell lines that express either IBV or mouse hepatitis virus N protein. Both N protein and a fibrillarin-green fluorescent protein fusion protein colocalized to the perinuclear region and the nucleolus. Pull-down assays demonstrated that IBV N protein interacted with nucleolin and therefore provided a possible explanation as to how coronavirus N proteins localize to the nucleolus. Nucleoli, and proteins that localize to the nucleolus, have been implicated in cell growth-cell cycle regulation. Comparison of cells expressing IBV N protein with controls indicated that cells expressing N protein had delayed cellular growth. This result could not to be attributed to apoptosis. Morphological analysis of these cells indicated that cytokinesis was disrupted, an observation subsequently found in primary cells infected with IBV. Coronaviruses might therefore delay the cell cycle in interphase, where maximum translation of viral mRNAs can occur.
Association of the Caveola Vesicular System with Cellular Entry by Filoviruses
Authors: Empig Cyril J. , Goldsmith Mark A. 
Published: July 2002

ABSTRACT
The filoviruses Ebola Zaire virus and Marburg virus are believed to infect target cells through endocytic vesicles, but the details of this pathway are unknown. We used a pseudotyping strategy to investigate the cell biology of filovirus entry. We observed that specific inhibitors of the caveola system, including cholesterol-sequestering drugs and phorbol esters, inhibited the entry of filovirus pseudotypes into human cells. We also measured slower cell entry kinetics for both filovirus pseudotypes than for pseudotypes of vesicular stomatitis virus (VSV), which has been recognized to exploit the clathrin-mediated entry pathway. Finally, visualization by immunofluorescence and confocal microscopy revealed that the filovirus pseudotypes colocalized with the caveola protein marker caveolin-1 but that VSV pseudotypes did not. Collectively, these results provide evidence suggesting that filoviruses use caveolae to gain entry into cells.
Folding and Dimerization of Tick-Borne Encephalitis Virus Envelope Proteins prM and E in the Endoplasmic Reticulum
Authors: Lorenz Ivo C. , Allison Steven L. , Heinz Franz X. , Helenius Ari 
Published: July 2002

ABSTRACT

Flavivirus envelope proteins are synthesized as part of large polyproteins that are co- and posttranslationally cleaved into their individual chains. To investigate whether the interaction of neighboring proteins within the precursor protein is required to ensure proper maturation of the individual components, we have analyzed the folding of the flavivirus tick-borne encephalitis (TBE) virus envelope glycoproteins prM and E by using a recombinant plasmid expression system and virus-infected cells. When expressed in their polyprotein context, prM and E achieved their native folded structures with half-times of approximately 4 min for prM and about 15 min for E. They formed heterodimeric complexes within a few minutes after synthesis that were required for the final folding of E but not for that of prM. Heterodimers could also be formed in
trans
when these proteins were coexpressed from separate constructs. When expressed without prM, E could form disulfide bonds but did not express a specific conformational epitope and remained sensitive to reduction by dithiothreitol. This is consistent with a chaperone-like role for prM in the folding of E. PrM was able to achieve its native folded structure without coexpression of E, but signal sequence cleavage at the N terminus was delayed. Our results show that prM is an especially rapidly folding viral glycoprotein, that polyprotein cleavage and folding of the TBE virus envelope proteins occurs in a coordinated sequence of processing steps, and that proper and efficient maturation of prM and E can only be achieved by cosynthesis of these two proteins.
Inhibition of Tumor Necrosis Factor Alpha-Induced NF-κB Activation by the Adenovirus E3-10.4/14.5K Complex
Authors: Friedman Joshua M. , Horwitz Marshall S. 
Published: July 2002

ABSTRACT
Recombinant adenoviruses (Ads) are useful tools in gene transfer because they are able to infect a wide variety of tissues and cell types and do not require a replicating target cell. However, transgene expression is only transient due to host innate and acquired immune responses to the virus. Most recombinant Ads have deletions of early region 3 (E3) genes, allowing more space for insertion of the transgene. Although the E3 region is not necessary for infection, it has been observed that these “nonessential” genes have immunomodulatory properties. We demonstrate here that the E3 region of Ad inhibits the activation of NF-κB induced by tumor necrosis factor alpha (TNF-α) and interleukin-1. Ad E3 is able to prevent NF-κB from entering the nucleus, where it is normally active. Ad E3 also appears to function by preventing the activation of the kinase complex, IKK, which is responsible for phosphorylation of IκB that retains NF-κB in the cytoplasm in an inactive state. The prevention of NF-κB activation has been mapped to a complex of two of the seven E3 products, E3-10.4K and E3-14.5K (RIDα/β). These and other studies indicate that, by using Ad vectors containing the E3 region, it may be possible to reduce the harmful proinflammatory effects of TNF-α and other cytokines that compromise the use of Ad gene therapy vectors in vivo.