bluuebunny/crossref_metadata_2025_split huggingface dataset:
Phosphorylation of the Human T-Cell Leukemia Virus Type 1 Transactivator Tax on Adjacent Serine Residues Is Critical for Tax Activation
Authors: Bex Françoise , Murphy Kathy , Wattiez Ruddy , Burny Arsène , Gaynor Richard B. 
Published: December 2019

ABSTRACT
The Tax transactivator protein of human T-cell leukemia virus type 1 (HTLV-1) plays a central role in the activation of viral gene expression. In addition, Tax is capable of activating the expression of specific cellular genes and is involved in the transformation of T-lymphocytes resulting in the development of adult T-cell leukemia. Tax is a phosphoprotein that colocalizes in nuclear bodies with RNA polymerase II, splicing complexes, and specific transcription factors including members of the ATF/CREB and NF-κB families. In this study, we identified adjacent serine residues at positions 300 and 301 in the carboxy terminus of Tax as the major sites for phosphorylation. Phosphorylation of at least one of these serine residues is required for Tax localization in nuclear bodies and for Tax-mediated activation of gene expression via both the ATF/CREB and NF-κB pathways. Introduction of amino acid substitutions which are phosphoserine mimetics at positions 300 and 301 restored the ability of a phosphorylation-defective Tax mutant to form nuclear bodies and to activate gene expression. These studies define sites for regulatory phosphorylation events in Tax which are critical for its ability to activate gene transcription.
Fine Structure and Morphogenesis of Borna Disease Virus
Authors: Kohno Takehiro , Goto Toshiyuki , Takasaki Tomohiko , Morita Chizuko , Nakaya Takaaki , Ikuta Kazuyoshi , Kurane Ichiro , Sano Kouichi , Nakai Masuyo 
Published: December 2019

ABSTRACT

Borna disease virus (BDV), a negative nonsegmented single-stranded RNA virus, has not been fully characterized morphologically. Here we present what is to our knowledge the first data on the fine ultrastructure and morphogenesis of BDV. The supernatant of MDCK cells persistently infected with BDV treated with
n
-butyrate contained many virus-like particles and more BDV-specific RNA than that of untreated samples. The particles were spherical, enveloped, and approximately 130 nm in diameter; had spikes 7 nm in length; and reacted with BDV p40 antibody. A thin nucleocapsid, 4 nm in width, was present peripherally in contrast to the thick nucleocapsid of hemagglutinating virus of Japan. The BDV particles reproduced by budding on the cell surface.
Identical 371-Base-Pair Deletion Mutations in the LAT Genes of Herpes Simplex Virus Type 1 McKrae and 17syn+ Result in Different In Vivo Reactivation Phenotypes
Authors: Loutsch Jeannette M. , Perng Guey-Chuen , Hill James M. , Zheng Xiaodong , Marquart Mary E. , Block Timothy M. , Ghiasi Homayon , Nesburn Anthony B. , Wechsler Steven L. 
Published: December 2019

ABSTRACT

The herpes simplex virus type 1 (HSV-1) LAT gene is the only viral gene abundantly transcribed during latency. LAT null mutants created with strains McKrae and 17syn+ are impaired for both in vivo spontaneous and in vivo-induced reactivation. Thus, LAT is essential for efficient in vivo-induced and spontaneous reactivation. Different investigators have studied two LAT mutants containing a
Sty
I-
Sty
I region deletion corresponding to LAT nucleotides 76 to 447. One mutant,
d
LAT371 (parent strain, McKrae), had parental high frequencies of spontaneous reactivation. In vivo-induced reactivation was not examined. The other mutant, 17ΔSty (parent strain, 17syn+), had parental frequencies of in vitro reactivation following cocultivation of explanted ganglia but reduced frequencies of in vivo-induced reactivation. Spontaneous reactivation frequency was not reported for 17ΔSty. These combined results suggested the possibility that in vivo spontaneous reactivation and in vivo-induced reactivation may map to different regions within the LAT domain. We now report that
d
LAT371 has in vivo-induced reactivation frequencies of the parent and that 17ΔSty has reduced frequencies of in vivo spontaneous reactivation. Thus,
d
LAT371 demonstrated the parental phenotype for both in vivo spontaneous and -induced reactivation while the apparently identical 17ΔSty was impaired for both in vivo spontaneous and -induced reactivation. These results suggest that one or more differences between the genetic backgrounds of McKrae and 17syn+ result in different in vivo reactivation phenotypes of otherwise identical deletion mutations and that McKrae may have compensating sequences sufficient to overcome the loss of the
Sty
I-
Sty
I region of the LAT transcript.
Down Regulation of Gene Expression by the Vaccinia Virus D10 Protein
Authors: Shors Teri , Keck James G. , Moss Bernard 
Published: December 2019

ABSTRACT
Vaccinia virus genes are expressed in a sequential fashion, suggesting a role for negative as well as positive regulatory mechanisms. A potential down regulator of gene expression was mapped by transfection assays to vaccinia virus open reading frame D10, which encodes a protein with no previously known function. Inhibition was independent of the promoter type used for the reporter gene, indicating that the mechanism did not involve promoter sequence recognition. The inhibition was overcome, however, when the open reading frame of the reporter gene was preceded by the encephalomyocarditis virus internal ribosome entry site, which excludes the possibility of nonspecific metabolic or other antiviral effects and suggests that capped mRNAs or cap-dependent translation might be the target of the D10 product. The inducible overexpression of the D10 gene by a recombinant vaccinia virus severely inhibited viral protein synthesis, decreased the steady-state level of viral late mRNA, and blocked the formation of infectious virus.
Induction and Maintenance of Autonomous Flock House Virus RNA1 Replication
Authors: Johnson Kyle L. , Ball L. Andrew 
Published: December 2019

ABSTRACTThe nodavirus flock house virus (FHV) has a bipartite, positive-sense, RNA genome that encodes the catalytic subunit of the RNA replicase and the viral capsid protein precursor on separate genomic segments (RNA1 and RNA2, respectively). RNA1 can replicate autonomously when transfected into permissive cells, allowing study of the kinetics of RNA1 replication in the absence of either RNA2 or capsid proteins. However, RNA1 replication ceases ca. 3 days after transfection despite the presence of replication-competent RNA. We examined this inhibition by inducing the expression of RNA1 in cells from a cDNA copy that was under the control of a hormone-regulated RNA polymerase II promoter. This system reproduced the shutoff of RNA replication when DNA-templated primary transcription was turned off. Continued primary transcription partially alleviated the shutoff and maintained the rate of RNA replication for several days at a steady-state level approximately one-third that of the peak rate. After shutoff, RNA replication could be restored by transferring the resulting intracellular RNA to fresh cells or by reinducing primary transcription, indicating that cessation of replication occurred despite the competence of both the viral RNA and the cytoplasmic environment. These data suggest that there is a mechanism by which replication is shut off at late times after transfection, which may reflect the natural endpoint of the replicative cycle.
Effects of Stuffer DNA on Transgene Expression from Helper-Dependent Adenovirus Vectors
Authors: Parks Robin J. , Bramson Jonathan L. , Wan Yonghong , Addison Christina L. , Graham Frank L. 
Published: December 2019

ABSTRACTWe have analyzed transgene (lacZ) expression from a first-generation adenovirus (Ad) vector in comparison to helper-dependent (hd) Ads deleted for various portions of the viral coding sequences and generated by using the Cre/loxP helper-dependent system (R. J. Parks et al., Proc. Natl. Acad. Sci. USA 93:13565–13570, 1996). An hd vector deleted for approximately 70% of the Ad genome (AdRP1001) provided levels and durations of transgene expression similar to those of a control first generation Ad vector containing an identical expression cassette. Deletion of all Ad sequences from the hdAd and replacement with a ∼22-kb fragment of lambda DNA resulted in a decrease in the level and duration oflacZexpression which could not be reversed by the inclusion of a matrix attachment region. However, substitution of the lambda stuffer in the fully deleted hdAd with sequences from the human hypoxanthine-guanine phosphoribosyltransferase gene resulted in significantly improved transgene expression. In vitro assays for cytotoxic T lymphocytes (CTL) directed against putative peptides encoded by the vector backbone showed that, although CTL were generated against the vector containing the lambda DNA, no such CTL were generated against the vector containing the hypoxanthine-guanine phosphoribosyltransferase (HPRT) sequences. Surprisingly, the rate of loss of the HPRT- and lambda-containing vectors from mouse liver was similar, despite the differences in expression kinetics, indicating that the lambda stuffer-directed CTL were inefficient at eliminating the transduced cells. Thus, the nature of the DNA backbone of hdAds can have important effects on the functioning of the vector. Since most fully deleted vectors require “stuffer” DNA as part of the vector backbone to maintain optimum vector size, these observations must be taken into account in the design of hdAd vectors.
Human Herpesvirus 6B Genome Sequence: Coding Content and Comparison with Human Herpesvirus 6A
Authors: Dominguez Geraldina , Dambaugh Timothy R. , Stamey Felicia R. , Dewhurst Stephen , Inoue Naoki , Pellett Philip E. 
Published: December 2019

ABSTRACT

Human herpesvirus 6 variants A and B (HHV-6A and HHV-6B) are closely related viruses that can be readily distinguished by comparison of restriction endonuclease profiles and nucleotide sequences. The viruses are similar with respect to genomic and genetic organization, and their genomes cross-hybridize extensively, but they differ in biological and epidemiologic features. Differences include infectivity of T-cell lines, patterns of reactivity with monoclonal antibodies, and disease associations. Here we report the complete genome sequence of HHV-6B strain Z29 [HHV-6B(Z29)], describe its genetic content, and present an analysis of the relationships between HHV-6A and HHV-6B. As sequenced, the HHV-6B(Z29) genome is 162,114 bp long and is composed of a 144,528-bp unique segment (U) bracketed by 8,793-bp direct repeats (DR). The genomic sequence allows prediction of a total of 119 unique open reading frames (ORFs), 9 of which are present only in HHV-6B. Splicing is predicted in 11 genes, resulting in the 119 ORFs composing 97 unique genes. The overall nucleotide sequence identity between HHV-6A and HHV-6B is 90%. The most divergent regions are DR and the right end of U, spanning ORFs U86 to U100. These regions have 85 and 72% nucleotide sequence identity, respectively. The amino acid sequences of 13 of the 17 ORFs at the right end of U differ by more than 10%, with the notable exception of U94, the adeno-associated virus type 2
rep
homolog, which differs by only 2.4%. This region also includes putative
cis
-acting sequences that are likely to be involved in transcriptional regulation of the major immediate-early locus. The catalog of variant-specific genetic differences resulting from our comparison of the genome sequences adds support to previous data indicating that HHV-6A and HHV-6B are distinct herpesvirus species.
Target Structures of the CD8+-T-Cell Response to Human Cytomegalovirus: the 72-Kilodalton Major Immediate-Early Protein Revisited
Authors: Kern Florian , Surel Ingolf Pascal , Faulhaber Nicole , Frömmel Claudia , Schneider-Mergener Jens , Schönemann Constanze , Reinke Petra , Volk Hans-Dieter 
Published: December 2019

ABSTRACTCell-mediated immunity plays an essential role in the control of infection with the human cytomegalovirus (HCMV). However, only a few CD8+-T-cell epitopes are known, with the majority being contained in the pp65 phosphoprotein, which is believed to dominate the CD8+-T-cell response to HCMV. Here, we have readdressed the issue of CD8+T cells specific for the 72-kDa major immediate-early protein (IE-1), which is nonstructural but is found very early and throughout the replicative cycle. Using a novel flow-cytometric assay, we were able to identify CD8+-T-cell epitopes (by IE-1 peptide-specific induction of cytokine synthesis) and simultaneously measure the frequency of cells directed against them. For this purpose, 81 pentadecamer peptides covering the complete 491-amino-acid sequence of IE-1 were tested on peripheral blood mononuclear cells of anti-HCMV immunoglobulin G-seropositive donors. At least 10 new epitopes were identified, and the fine specificity and presenting HLA molecule of the first of them was determined. The frequencies of CD8+T cells directed against IE-1 were similar to those directed against pp65 in donors tested with known pp65-derived peptides. Importantly, additional testing of a corresponding set of peptides covering the complete sequence of pp65 on 10 of these donors identified individuals whose CD8+T cells recognized IE-1 but not pp65 and vice versa, clearly illustrating that either protein may be a major target. In summary, our results suggest that IE-1 is far more important as a CD8+-T-cell target than current opinion suggests.
Genetic Analysis of
Gv1
, a Gene Controlling Transcription of Endogenous Murine Polytropic Proviruses
Authors: Oliver Peter L. , Stoye Jonathan P. 
Published: December 2019

ABSTRACT

Gv1
is a genetic locus that coordinately regulates the expression of multiple murine leukemia virus-related endogenous proviral sequences. A quantitative nuclease protection assay for typing
Gv1
inheritance has been developed. Use of this assay demonstrates that
Gv1
controls transcription of polytropic but not of modified polytropic endogenous proviruses. A combination of genetic techniques were used to map
Gv1
; these analyses demonstrate that
Gv1
lies approximately 37 centimorgans from the centromeric end of mouse chromosome 13.
Diminished Rev-Mediated Stimulation of Human Immunodeficiency Virus Type 1 Protein Synthesis Is a Hallmark of Human Astrocytes
Authors: Ludwig Eva , Silberstein Francesca Ceccherini- , van Empel Jutta , Erfle Volker , Neumann Markus , Brack-Werner Ruth 
Published: December 2019

ABSTRACT

Astrocytes are target cells for human immunodeficiency virus type 1 (HIV-1) in the central nervous system with attenuated virus replication in vivo and in vitro. In infected astrocytes, viral gene expression is restricted mainly to nonstructural (early) viral components like Nef, suggesting inhibition of Rev-dependent posttranscriptional processes in these cells. Because of the heterogeneity of astrocytic cells, the objective of this study was to determine whether restriction of HIV-1 Rev-associated activities is a common property of human astrocytes. To this end, we compared the
trans
activation capacity and intracellular distribution of Rev in four astrocytoma cell lines previously shown to be infectible by HIV-1 and in primary human fetal astrocytes from different sources with Rev-permissive nonglial control cell lines. In all astrocytic cell cultures, the Rev response was reduced to about 10% of that of Rev-permissive control cells. Rev was apparent both in cytoplasmic and in nuclear compartments of living astrocytes, in contrast to the typical nuclear and/or nucleolar localization of Rev in permissive control cells. Nuclear accumulation of Rev in astrocytes was restored by blocking export of Rev. The
trans
activation capacity and nuclear localization of Tat were not affected in astrocytes. These results demonstrate that inhibition of Rev-dependent posttranscriptional regulation of HIV-1 is a hallmark of human astrocytes and may contribute to suppression of HIV-1 production in these HIV-1 reservoirs. Astrocytes constitute the first example of a human cell type showing an impaired Rev response, indicating that posttranscriptional control of HIV-1 gene expression can be modulated in a cell-dependent manner.
Alternative Translation Initiation of Theiler’s Murine Encephalomyelitis Virus
Authors: Yamasaki Kenji , Weihl Conrad C. , Roos Raymond P. 
Published: December 2019

ABSTRACTDA strain and other members of the TO subgroup of Theiler’s murine encephalomyelitis virus (TMEV) produce a chronic demyelinating disease in which the virus persists but has a restricted expression. We previously reported that TO subgroup strains, in addition to synthesizing the picornaviral polyprotein, use an alternative initiation codon just downstream from the polyprotein’s AUG to translate an 18-kDa protein called L* that is out of frame with the polyprotein (H. H. Chen et al., Nat. Med. 1:927–931, 1995; W. P. Kong and R. P. Roos, J. Virol. 65:3395–3399, 1991). L* is critically important for virus persistence and the induction of the demyelinating disease (Chen et al., 1995; G. D. Ghadge et al. J. Virol. 72:8605–8612, 1998). We have proposed that variations in the amount of translation initiation from the L* AUG versus the polyprotein AUG may occur in different cell types and therefore affect the degree of expression of viral capsid proteins. We now demonstrate that ribosomal translation initiation at the polyprotein’s initiation codon affects initiation at the L* AUG, suggesting that ribosomes land at the polyprotein’s initiation codon before scanning downstream and initiating at the L* AUG. We also find that the viral 5′ untranslated region affects utilization of the L* AUG. Surprisingly, mutant DA cDNAs were found to be infectious despite the presence of mutations of the polyprotein initiation codon or placement of a stop codon upstream of the L* AUG in the polyprotein’s reading frame. Sequencing studies showed that these viruses had a second site mutation, converting the reading frame of L* into the polyprotein’s reading frame; the results suggest that translation of the polyprotein during infection of these mutant viruses can be initiated at the L* AUG. These data are important in our understanding of translation initiation of TMEV and other RNAs that contain an internal ribosome entry site.
African Swine Fever Virus Replication in the Midgut Epithelium Is Required for Infection of
Ornithodoros
Ticks
Authors: Kleiboeker S. B. , Scoles G. A. , Burrage T. G. , Sur J.-H. 
Published: December 2019

ABSTRACT

Although the Malawi Lil20/1 (MAL) strain of African swine fever virus (ASFV) was isolated from
Ornithodoros
sp. ticks, our attempts to experimentally infect ticks by feeding them this strain failed. Ten different collections of
Ornithodorus porcinus porcinus
ticks and one collection of
O. porcinus domesticus
ticks were orally exposed to a high titer of MAL. At 3 weeks postinoculation (p.i.), <25% of the ticks contained detectable virus, with viral titers of <4 log
10
50% hemadsorbing doses/ml. Viral titers declined to undetectability in >90% of the ticks by 5 weeks p.i. To further study the growth defect,
O. porcinus porcinus
ticks were orally exposed to MAL and assayed at regular intervals p.i. Whole-tick viral titers dramatically declined (>1,000-fold) between 2 and 6 days p.i., and by 18 days p.i., viral titers were below the detection limit. In contrast, viral titers of ticks orally exposed to a tick-competent ASFV isolate, Pretoriuskop/96/4/1 (Pr4), increased 10-fold by 10 days p.i. and 50-fold by 14 days p.i. Early viral gene expression, but not extensive late gene expression or viral DNA synthesis, was detected in the midguts of ticks orally exposed to MAL. Ultrastructural analysis demonstrated that progeny virus was rarely present in ticks orally exposed to MAL and, when present, was associated with extensive cytopathology of phagocytic midgut epithelial cells. To determine if viral replication was restricted only in the midgut epithelium, parenteral inoculations into the hemocoel were performed. With inoculation by this route, a persistent infection was established although a delay in generalization of MAL was detected and viral titers in most tissues were typically 10- to 1,000-fold lower than those of ticks injected with Pr4. MAL was detected in both the salivary secretion and coxal fluid following feeding but less frequently and at a lower titer compared to Pr4. Transovarial transmission of MAL was not detected after two gonotrophic cycles. Ultrastructural analysis demonstrated that, when injected, MAL replicated in a number of cell types but failed to replicate in midgut epithelial cells. In contrast, ticks injected with Pr4 had replicating virus in midgut epithelial cells. Together, these results indicate that MAL replication is restricted in midgut epithelial cells. This finding demonstrates the importance of viral replication in the midgut for successful ASFV infection of the arthropod host.
Initial Binding of Murine Leukemia Virus Particles to Cells Does Not Require Specific Env-Receptor Interaction
Authors: Pizzato Massimo , Marlow Susan A. , Blair Edward D. , Takeuchi Yasuhiro 
Published: December 2019

ABSTRACT
The initial step of virus-cell interaction was studied by immunofluorescence microscopy. Single particles of murine leukemia virus (MLV) vectors and human immunodeficiency virus (HIV) were visualized by immunofluorescence. Fluorescent dots representing single virions could be localized by staining of capsid proteins (CA) or surface envelope proteins (SU) after fixation of virus supernatants. This technique can be used to determine particle concentration in viral supernatants and also to study virus-cell interaction. We investigated the role of the Env-receptor interaction for the initial binding event between the cell and the viral particles. Ecotropic MLV vector particles were shown to bind to human cells which do not express the specific viral receptor. In addition, MLV particles defective for Env were shown to bind the cells similarly to infectious MLV. Time course experiments of virus-cell binding and dissociation showed identical profiles for infectious and Env-defective MLV particles and suggested that MLV Env is not involved in the early phases of attachment of virus to cells. The possible implication of cellular factors in enhancing viral binding and infectivity is discussed.
Comparative Genomics of
Streptococcus thermophilus
Phage Species Supports a Modular Evolution Theory
Authors: Lucchini Sacha , Desiere Frank , Brüssow Harald 
Published: December 2019

ABSTRACT

The comparative analysis of five completely sequenced
Streptococcus thermophilus
bacteriophage genomes demonstrated that their diversification was achieved by a combination of DNA recombination events and an accumulation of point mutations. The five phages included lytic and temperate phages, both
pac
site and
cos
site, from three distinct geographical areas. The units of genetic exchange were either large, comprising the entire morphogenesis gene cluster, excluding the putative tail fiber genes, or small, consisting of one or maximally two genes or even segments of a gene. Many indels were flanked by DNA repeats. Differences in a single putative tail fiber gene correlated with the host ranges of the phages. The predicted tail fiber protein consisted of highly conserved domains containing conspicuous glycine repeats interspersed with highly variable domains. As in the T-even coliphage adhesins, the glycine-containing domains were recombinational hot spots. Downstream of a highly conserved DNA replication region, all lytic phages showed a short duplication; in three isolates the origin of replication was repeated. The lytic phages could conceivably be derived from the temperate phages by deletion and multiple rearrangement events in the lysogeny module, giving rise to occasional selfish phages that defy the superinfection control systems of the corresponding temperate phages.