bluuebunny/crossref_metadata_2025_split huggingface dataset:
C3H Mouse Mammary Tumor Virus Superantigen Function Requires a Splice Donor Site in the Envelope Gene
Authors: Mustafa Farah , Lozano Mary , Dudley Jaquelin P. 
Published: July 2002

ABSTRACTMouse mammary tumor virus (MMTV) encodes a superantigen (Sag) that is required for efficient milk-borne transmission of virus from mothers to offspring. The mRNA used for Sag expression is controversial, and at least four different promoters (two in the long terminal repeat and two in the envelope gene) forsagmRNA have been reported. To determine which RNA is responsible for Sag function during milk-borne MMTV transmission, we mutated a splice donor site unique to a splicedsagRNA from the 5′ envelope promoter. The splice donor mutation in an infectious provirus was transfected into XC cells and injected into BALB/c mice. Mice injected with wild-type provirus showed Sag activity by the deletion of Sag-specific T cells and induction of mammary tumors in 100% of injected animals. However, mice injected with the splice donor mutant gave sporadic and delayed T-cell deletion and a low percentage of mammary tumors with a long latency, suggesting that the resulting tumors were due to the generation of recombinants with endogenous MMTVs. Third-litter offspring of mice injected with wild-type provirus showed Sag-specific T-cell deletion and developed mammary tumors with kinetics similar to those for mice infected by nursing on MMTV-infected mothers, whereas the third-litter offspring of the splice donor mutant-injected mice did not. One of the fifth-litter progeny of splice donor mutant-injected mice showed C3H Sag activity and had recombinants that repaired the splice donor mutation, thus confirming the necessity for the splice donor site for Sag function. These experiments are the first to show that the splicedsagmRNA from the 5′ envelope promoter is required for efficient milk-borne transmission of C3H MMTV.
Global Analysis of Herpes Simplex Virus Type 1 Transcription Using an Oligonucleotide-Based DNA Microarray
Authors: Stingley S. W. , Ramirez J. J. Garcia , Aguilar S. A. , Simmen K. , Sandri-Goldin R. M. , Ghazal P. , Wagner E. K. 
Published: July 2002

ABSTRACTMore than 100 transcripts of various abundances and kinetic classes are expressed during phases of productive and latent infections by herpes simplex virus (HSV) type 1. To carry out rapid global analysis of variations in such patterns as a function of perturbation of viral regulatory genes and cell differentiation, we have made DNA microchips containing sets of 75-mer oligonucleotides specific for individual viral transcripts. About half of these are unique for single transcripts, while others function for overlapping ones. We have also included probes for 57 human genes known to be involved in some aspect of stress response. The chips efficiently detect all viral transcripts, and analysis of those abundant under various conditions of infection demonstrates excellent correlation with known kinetics of mRNA accumulation. Further, quantitative sensitivity is high. We have further applied global analysis of transcription to an investigation of mRNA populations in cells infected with a mutant virus in which the essential immediate-early α27 (UL54) gene has been functionally deleted. Transcripts expressed at 6 h following infection with this mutant can be classified into three groups: those whose abundance is augmented (mainly immediate-early transcripts) or unaltered, those whose abundance is somewhat reduced, and those where there is a significant reduction in transcript levels. These do not conform to any particular kinetic class. Interestingly, levels of many cellular transcripts surveyed are increased. The high proportion of such transcripts suggests that the α27 gene plays a major role in the early decline in cellular gene expression so characteristic of HSV infection.
High Rate of Recombination throughout the Human Immunodeficiency Virus Type 1 Genome
Authors: Jetzt Amanda E. , Yu Hong , Klarmann George J. , Ron Yacov , Preston Bradley D. , Dougherty Joseph P. 
Published: July 2002

ABSTRACT
The diploid nature of human immunodeficiency virus type 1 (HIV-1) indicates that recombination serves a central function in virus replication and evolution. Previously, while examining the nature of obligatory primer strand transfers during reverse transcription, a high rate of recombination was observed at the ends of the viral genome within the viral long terminal repeats, prompting the following question: does recombination occur at a high rate throughout the genome? To address this question, two vectors based upon different strains of HIV-1 were utilized. The vectors were comprised predominantly of autologous HIV-1 sequence and were approximately the same size as the parental genome. The proviral progeny of heterodimeric virions were analyzed after a single cycle of replication, and the sequence heterogeneity between the two strains allowed direct examination of recombination crossovers. The results obtained indicate that HIV-1 undergoes approximately two to three recombination events per genome per replication cycle. These results imply that both HIV-1 RNAs are typically utilized during reverse transcription and that recombination is an important aspect of HIV-1 replication.
An African Swine Fever Virus
ERV1-ALR
Homologue,
9GL
, Affects Virion Maturation and Viral Growth in Macrophages and Viral Virulence in Swine
Authors: Lewis T. , Zsak L. , Burrage T. G. , Lu Z. , Kutish G. F. , Neilan J. G. , Rock D. L. 
Published: July 2002

ABSTRACT

The African swine fever virus (ASFV) genome contains a gene,
9GL
, with similarity to yeast
ERV1
and
ALR
genes.
ERV1
has been shown to function in oxidative phosphorylation and in cell growth, while
ALR
has hepatotrophic activity.
9GL
encodes a protein of 119 amino acids and was highly conserved at both nucleotide and amino acid levels among all ASFV field isolates examined. Monospecific rabbit polyclonal antibody produced to a glutathione
S
-transferase–9GL fusion protein specifically immunoprecipitated a 14-kDa protein from macrophage cell cultures infected with the ASFV isolate Malawi Lil-20/1 (MAL). Time course analysis and viral DNA synthesis inhibitor experiments indicated that p14 was a late viral protein. A
9GL
gene deletion mutant of MAL (Δ9GL), exhibited a growth defect in macrophages of approximately 2 log
10
units and had a small-plaque phenotype compared to either a revertant (9GL-R) or the parental virus. 9GL affected normal virion maturation; virions containing acentric nucleoid structures comprised 90 to 99% of all virions observed in Δ9GL-infected macrophages. The Δ9GL virus was markedly attenuated in swine. In contrast to 9GL-R infection, where mortality was 100%, all Δ9GL-infected animals survived infection. With the exception of a transient fever response in some animals, Δ9GL-infected animals remained clinically normal and exhibited significant 100- to 10,000-fold reductions in viremia titers. All pigs previously infected with Δ9GL survived infection when subsequently challenged with a lethal dose of virulent parental MAL. Thus, ASFV
9GL
gene deletion mutants may prove useful as live-attenuated ASF vaccines.
The Human Cytomegalovirus Major Immediate-Early Distal Enhancer Region Is Required for Efficient Viral Replication and Immediate-Early Gene Expression
Authors: Meier Jeffery L. , Pruessner Jonathan A. 
Published: July 2002

ABSTRACT
The human cytomegalovirus (HCMV) major immediate-early (MIE) genes, encoding IE1 p72 and IE2 p86, are activated by a complex enhancer region (base positions -65 to -550) that operates in a cell type- and differentiation-dependent manner. The expression of MIE genes is required for HCMV replication. Previous studies analyzing functions of MIE promoter-enhancer segments suggest that the distal enhancer region variably modifies MIE promoter activity, depending on cell type, stimuli, or state of differentiation. To further understand the mechanism by which the MIE promoter is regulated, we constructed and analyzed several different recombinant HCMVs that lack the distal enhancer region (-300 to -582, -640, or -1108). In human fibroblasts, the HCMVs without the distal enhancer replicate normally at high multiplicity of infection (MOI) but replicate poorly at low MOI in comparison to wild-type virus (WT) or HCMVs that lack the neighboring upstream unique region and modulator (-582 or -640 to -1108). The growth aberrancy was normalized after restoring the distal enhancer in a virus lacking this region. For HCMVs without a distal enhancer, the impairment in replication at low MOI corresponds to a deficiency in production of MIE RNAs compared to WT or virus lacking the unique region and modulator. An underproduction of viral US3 RNA was also evident at low MOI. Whether lower production of IE1 p72 and IE2 p86 causes a reduction in expression of the immediate-early (IE) class US3 gene remains to be determined. We conclude that the MIE distal enhancer region possesses a mechanism for augmenting viral IE gene expression and genome replication at low MOI, but this regulatory function is unnecessary at high MOI.
Characterization of the Interaction between the Interferon-Induced Protein P56 and the Int6 Protein Encoded by a Locus of Insertion of the Mouse Mammary Tumor Virus
Authors: Guo Jinjiao , Sen Ganes C. 
Published: July 2002

ABSTRACT

For determining cellular functions of the interferon-inducible human cytoplasmic protein P56, we undertook a
Saccharomyces cerevisiae
two-hybrid screen that identified Int6 as a P56-interacting protein. That the interaction also occurs in human cells was confirmed by coimmunoprecipitation and the observed cytoplasmic displacement of nuclear Int6 upon coexpression of P56. Because Int6 has been claimed to be both a cytoplasmic and a nuclear protein, we investigated the structural basis of this discrepancy. By mutational analyses, we showed that the Int6 protein contains a bipartite nuclear localization signal and a nuclear export signal at the far end of the amino terminus. The 20 amino-terminal residues of Int6, when they were attached to a different nuclear protein, were sufficient to translocate that protein to the cytoplasm. Within this region, replacement of any of the three leucine residues with alanine destroyed the function of the export signal. The specific domain of P56 that is required for its interaction with Int6 was mapped using the yeast two-hybrid assay and a mammalian coimmunoprecipitation assay. Both assays demonstrated that the C-terminal region of P56 containing three specific tetratricopeptide motifs is required for this interaction. In contrast, removal of an internal domain of P56 enhanced the interaction, as quantified by the two-hybrid assay.
Roscovitine, a Specific Inhibitor of Cellular Cyclin-Dependent Kinases, Inhibits Herpes Simplex Virus DNA Synthesis in the Presence of Viral Early Proteins
Authors: Schang Luis M. , Rosenberg Amy , Schaffer Priscilla A. 
Published: July 2002

ABSTRACT
We have previously shown that two inhibitors specific for cellular cyclin-dependent kinases (cdks), Roscovitine (Rosco) and Olomoucine (Olo), block the replication of herpes simplex virus (HSV). Based on these results, we demonstrated that HSV replication requires cellular cdks that are sensitive to these drugs (L. M. Schang, J. Phillips, and P. A. Schaffer. J. Virol. 72:5626–5637, 1998). We further established that at least two distinct steps in the viral replication cycle require cdks: transcription of immediate-early (IE) genes and transcription of early (E) genes (L. M. Schang, A. Rosenberg, and P. A. Schaffer, J. Virol. 73:2161–2172, 1999). Since Rosco inhibits HSV replication efficiently even when added to infected cells at 6 h postinfection, we postulated that cdks may also be required for viral functions that occur after E gene expression. In the study presented herein, we tested this hypothesis directly by measuring the efficiency of viral replication, viral DNA synthesis, and expression of several viral genes during infections in which Rosco was added after E proteins had already been synthesized. Rosco inhibited HSV replication, and specifically viral DNA synthesis, when the drug was added at the time of release from a 12-h phosphonoacetic acid (PAA)-induced block in viral DNA synthesis. Inhibition of DNA synthesis was not a consequence of inhibition of expression of IE or E genes in that Rosco had no effect on steady-state levels of two E transcripts under the same conditions in which it inhibited viral DNA synthesis. Moreover, viral DNA synthesis was inhibited by Rosco even in the absence of protein synthesis. In a second series of experiments, the replication of four HSV mutants harboring temperature-sensitive mutations in genes essential for viral DNA replication was inhibited when Rosco was added at the time of shift-down from the nonpermissive to the permissive temperature. Viral DNA synthesis was inhibited by Rosco under these conditions, whereas expression of viral E genes was not affected. We conclude that cellular Rosco-sensitive cdks are required for replication of viral DNA in the presence of viral E proteins. This requirement may indicate that HSV DNA synthesis is functionally linked to transcription, which requires cdks, or that both viral transcription and DNA replication, independently, require viral or cellular factors activated by Rosco-sensitive cdks.
Rotavirus Infection Induces an Increase in Intracellular Calcium Concentration in Human Intestinal Epithelial Cells: Role in Microvillar Actin Alteration
Authors: Brunet Jean-Philippe , Cotte-Laffitte Jacqueline , Linxe Catherine , Quero Anne-Marie , Géniteau-Legendre Monique , Servin Alain 
Published: July 2002

ABSTRACT

Rotaviruses, which infect mature enterocytes of the small intestine, are recognized as the most important cause of viral gastroenteritis in young children. We have previously reported that rotavirus infection induces microvillar F-actin disassembly in human intestinal epithelial Caco-2 cells (N. Jourdan, J. P. Brunet, C. Sapin, A. Blais, J. Cotte-Laffitte, F. Forestier, A. M. Quero, G. Trugnan, and A. L. Servin, J. Virol. 72:7228–7236, 1998). In this study, to determine the mechanism responsible for rotavirus-induced F-actin alteration, we investigated the effect of infection on intracellular calcium concentration ([Ca
2+
]
i
) in Caco-2 cells, since Ca
2+
is known to be a determinant factor for actin cytoskeleton regulation. As measured by quin2 fluorescence, viral replication induced a progressive increase in [Ca
2+
]
i
from 7 h postinfection, which was shown to be necessary and sufficient for microvillar F-actin disassembly. During the first hours of infection, the increase in [Ca
2+
]
i
was related only to an increase in Ca
2+
permeability of plasmalemma. At a late stage of infection, [Ca
2+
]
i
elevation was due to both extracellular Ca
2+
influx and Ca
2+
release from the intracellular organelles, mainly the endoplasmic reticulum (ER). We noted that at this time the [Ca
2+
]
i
increase was partially related to a phospholipase C (PLC)-dependent mechanism, which probably explains the Ca
2+
release from the ER. We also demonstrated for the first time that viral proteins or peptides, released into culture supernatants of rotavirus-infected Caco-2 cells, induced a transient increase in [Ca
2+
]
i
of uninfected Caco-2 cells, by a PLC-dependent efflux of Ca
2+
from the ER and by extracellular Ca
2+
influx. These supernatants induced a Ca
2+
-dependent microvillar F-actin alteration in uninfected Caco-2 cells, thus participating in rotavirus pathogenesis.
Development of an Effective Polyvalent Vaccine against both Marek's and Newcastle Diseases Based on Recombinant Marek's Disease Virus Type 1 in Commercial Chickens with Maternal Antibodies
Authors: Sonoda Kengo , Sakaguchi Masashi , Okamura Hiroshi , Yokogawa Kenji , Tokunaga Eiji , Tokiyoshi Sachio , Kawaguchi Yasushi , Hirai Kanji 
Published: July 2002

ABSTRACT
An earlier report (M. Sakaguchi et al., Vaccine 16:472–479, 1998) showed that recombinant Marek's disease virus type 1 (rMDV1) expressing the fusion (F) protein of Newcastle disease virus (NDV-F) under the control of the simian virus 40 late promoter [rMDV1-US10L(F)] protected specific pathogen-free chickens from NDV challenge, but not commercial chickens with maternal antibodies against NDV and MDV1. In the present study, we constructed an improved polyvalent vaccine based on MDV1 against MDV and NDV in commercial chickens with maternal antibodies. The study can be summarized as follows. (i) We constructed rMDV1 expressing NDV-F under the control of the MDV1 glycoprotein B (gB) promoter [rMDV1-US10P(F)]. (ii) Much less NDV-F protein was expressed in cells infected with rMDV1-US10P(F) than in those infected with rMDV1-US10L(F). (iii) The antibody response against NDV-F and MDV1 antigens of commercial chickens vaccinated with rMDV1-US10P(F) was much stronger and faster than with rMDV1-US10L(F), and a high level of antibody against NDV-F persisted for over 80 weeks postvaccination. (iv) rMDV1-US10P(F) was readily reisolated from the vaccinated chickens, and the recovered viruses were found to express NDV-F. (v) Vaccination of commercial chickens having maternal antibodies to rMDV1-US10P(F) completely protected them from NDV challenge. (vi) rMDV1-US10P(F) offered the same degree of protection against very virulent MDV1 as the parental MDV1 and commercial vaccines. These results indicate that rMDV1-US10P(F) is an effective and stable polyvalent vaccine against both Marek's and Newcastle diseases even in the presence of maternal antibodies.
Evidence for Budding of Human Immunodeficiency Virus Type 1 Selectively from Glycolipid-Enriched Membrane Lipid Rafts
Authors: Nguyen Dzung H. , Hildreth James E. K. 
Published: July 2002

ABSTRACTA number of recent studies have demonstrated the significance of detergent-insoluble, glycolipid-enriched membrane domains or lipid rafts, especially in regard to activation and signaling in T lymphocytes. These domains can be viewed as floating rafts composed of sphingolipids and cholesterol which sequester glycosylphosphatidylinositol (GPI)-linked proteins, such as Thy-1 and CD59. CD45, a 200-kDa transmembrane phosphatase protein, is excluded from these domains. We have found that human immunodeficiency virus type 1 (HIV-1) particles produced by infected T-cell lines acquire the GPI-linked proteins Thy-1 and CD59, as well as the ganglioside GM1, which is known to partition preferentially into lipid rafts. In contrast, despite its high expression on the cell surface, CD45 was poorly incorporated into virus particles. Confocal fluorescence microscopy revealed that HIV-1 proteins colocalized with Thy-1, CD59, GM1, and a lipid raft-specific fluorescent lipid, DiIC16(3), in uropods of infected Jurkat cells. CD45 did not colocalize with HIV-1 proteins and was excluded from uropods. Dot immunoassay of Triton X-100-extracted membrane fractions revealed that HIV-1 p17 matrix protein and gp41 were present in the detergent-resistant fractions and that [3H]myristic acid-labeled HIV Gag showed a nine-to-one enrichment in lipid rafts. We propose a model for the budding of HIV virions through lipid rafts whereby host cell cholesterol, sphingolipids, and GPI-linked proteins within these domains are incorporated into the viral envelope, perhaps as a result of preferential sorting of HIV Gag to lipid rafts.
Impaired Antiviral Response and Alpha/Beta Interferon Induction in Mice Lacking Beta Interferon
Authors: Deonarain Raj , Alcamí Antonio , Alexiou Maria , Dallman Margaret J. , Gewert Dirk R. , Porter Andrew C. G. 
Published: July 2002

ABSTRACTWe have generated mice lacking the gene for beta interferon and report that they are highly susceptible to vaccinia virus infection. Furthermore, in cultured embryo fibroblasts, viral induction of alpha interferon and of 2-5A synthetase genes is impaired. We also show that beta interferon does not prime its own expression.
Cultured Cell Sublines Highly Susceptible to Prion Infection
Authors: Bosque Patrick J. , Prusiner Stanley B. 
Published: July 2002

ABSTRACT

Cultured cell lines infected with prions produce an abnormal isoform of the prion protein (PrP
Sc
). In order to derive cell lines producing sufficient quantities of PrP
Sc
for most studies, it has been necessary to subclone infected cultures and select the subclones producing the largest amounts of PrP
Sc
. Since postinfection cloning can introduce differences between infected and uninfected cell lines, we sought an approach to generate prion-infected cell lines that would avoid clonal artifacts. Using an improved cell blot technique, which permits sensitive and rapid comparison of PrP
Sc
levels in multiple independent cell cultures, we discovered marked heterogeneity with regard to prion susceptibility in tumor cell sublines. We exploited this heterogeneity to derive sublines which are highly susceptible to prion infection and used these cells to generate prion-infected lines without further subcloning. These infected sublines can be compared to the cognate uninfected cultures without interference from cloning artifacts. We also used susceptible cell lines and our modified cell blot procedure to develop a sensitive and reproducible quantitative cell culture bioassay for prions. We found that the sublines were at least 100-fold more susceptible to strain RML prions than to strain ME7 prions. Comparisons between scrapie-susceptible and -resistant cell lines may reveal factors that modulate prion propagation.
The Human Immunodeficiency Virus Type 1gagGene Encodes an Internal Ribosome Entry Site
Authors: Buck Christopher B. , Shen Xuefei , Egan Michael A. , Pierson Theodore C. , Walker Christopher M. , Siliciano Robert F. 
Published: July 2002

ABSTRACTSeveral retroviruses have recently been shown to promote translation of theirgaggene products by internal ribosome entry. In this report, we show that mRNAs containing the human immunodeficiency virus type 1 (HIV-1)gagopen reading frame (ORF) exhibit internal ribosome entry site (IRES) activity that can promote translational initiation of Pr55gag. Remarkably, this IRES activity is driven by sequences within thegagORF itself and is not dependent on the nativegag5′-untranslated region (UTR). This cap-independent mechanism for Pr55gagtranslation may help explain the high levels of translation of this protein in the face of major RNA structural barriers to scanning ribosomes found in thegag5′ UTR. ThegagIRES activity described here also drives translation of a novel 40-kDa Gag isoform through translational initiation at an internal AUG codon found near the amino terminus of the Pr55gagcapsid domain. Our findings suggest that this low-abundance Gag isoform may be important for wild-type replication of HIV-1 in cultured cells. The activities of the HIV-1gagIRES may be an important feature of the HIV-1 life cycle and could serve as a novel target for antiretroviral therapeutic strategies.