bluuebunny/crossref_metadata_2025_split huggingface dataset:
Role of the Transcription Start Site Core Region and Transcription Factor YY1 in Rous Sarcoma Virus Long Terminal Repeat Promoter Activity
Authors: Mobley Constance M. , Sealy Linda 
Published: December 2019

ABSTRACT

The Rous sarcoma virus (RSV) long terminal repeat (LTR) contains a transcriptionally potent enhancer and promoter that functions in a variety of cell types. Previous studies have identified the viral sequences required for enhancer activity, and characterization of these elements has provided insight into the mechanism of RSV transcriptional activity. The objective of this study was to better define the RSV LTR promoter by examining the transcription start site core (TSSC) region. Deletion of the TSSC resulted in complete loss of transcriptional activity despite the presence of a functional TATA box, suggesting that the TSSC is required for viral expression. Homologies within the TSSC to the DNA binding motif of YY1 suggested that it might regulate promoter activity. YY1 has been shown to regulate transcription in some cellular genes and viral promoters by binding to sites overlapping the transcription start site. Gel shift assays using YY1 antibody identified YY1 as one of three complexes that bound to the TSSC. Mutation of the YY1 binding site reduced RSV transcriptional activity by more than 50%, suggesting that YY1, in addition to other TSSC-binding factors, regulates RSV transcription. Furthermore, in vitro transcription assays performed with
Drosophila
embryo extract (devoid of YY1 activity) showed decreased levels of RSV transcription, while transient transfection experiments overexpressing YY1 demonstrated that YY1 could transactivate the RSV LTR ∼6- to 7-fold. We propose that the TSSC plays a vital role in RSV transcription and that this function is partially carried out by the transcription factor YY1.
T-Cell-Independent Immunoglobulin G Responses In Vivo Are Elicited by Live-Virus Infection but Not by Immunization with Viral Proteins or Virus-Like Particles
Authors: Szomolanyi-Tsuda Eva , Le Quang P. , Garcea Robert L. , Welsh Raymond M. 
Published: December 2019

ABSTRACTImmunoglobulin G (IgG) responses to viruses are generally assumed to be T-cell dependent (TD). Recently, however, polyomavirus (PyV) infection of T-cell-deficient (T-cell receptor β chain [TCR-β] −/− or TCR-β×δ −/−) mice was shown to elicit a protective, T-cell-independent (TI) antiviral IgM and IgG response. A repetitive, highly organized antigenic structure common to many TI antigens is postulated to be important in the induction of antibody responses in the absence of helper T cells. To test whether the repetitive structure of viral antigens is essential and/or sufficient for the induction of TI antibodies, we compared the abilities of three forms of PyV antigens to induce IgM and IgG responses in T-cell-deficient mice: soluble capsid antigens (VP1), repetitive virus-like particles (VLPs), and live PyV. Immunization with each of the viral antigens resulted in IgM production. VLPs and PyV elicited 10-fold-higher IgM titers than VP1, indicating that the highly organized, repetitive antigens are more efficient in IgM induction. Antigen-specific TI IgG responses, however, were detected only in mice infected with live PyV, not in VP1- or VLP-immunized mice. These results suggest that the highly organized, repetitive nature of the viral antigens is insufficient to account for their ability to elicit TI IgG response and that signals generated by live-virus infection may be essential for the switch to IgG production in the absence of T cells. Germinal centers were not observed in T-cell-deficient PyV-infected mice, indicating that the germinal center pathway of B-cell differentiation is TD even in the context of a virus infection.
ORF1a-Encoded Replicase Subunits Are Involved in the Membrane Association of the Arterivirus Replication Complex
Authors: van der Meer Yvonne , van Tol Hans , Krijnse Locker Jacomine , Snijder Eric J. 
Published: December 2019

ABSTRACT

Among the functions of the replicase of equine arteritis virus (EAV; family
Arteriviridae
, order
Nidovirales
) are important viral enzyme activities such as proteases and the putative RNA polymerase and RNA helicase functions. The replicase is expressed in the form of two polyproteins (open reading frame 1a [ORF1a] and ORF1ab), which are processed into 12 nonstructural proteins by three viral proteases. In immunofluorescence assays, the majority of these cleavage products localized to the perinuclear region of the cell. A dense granular and vesicular staining was observed, which strongly suggested membrane association. By using confocal microscopy and double-label immunofluorescence, the distribution of the EAV replicase was shown to overlap with that of PDI, a resident protein of the endoplasmic reticulum and intermediate compartment. An in situ labeling of nascent viral RNA with bromo-UTP demonstrated that the membrane-bound complex in which the replicase subunits accumulate is indeed the site of viral RNA synthesis. A number of ORF1a-encoded hydrophobic domains were postulated to be involved in the membrane association of the arterivirus replication complex. By using various biochemical methods (Triton X-114 extraction, membrane purification, and sodium carbonate treatment), replicase subunits containing these domains were shown to behave as integral membrane proteins and to be membrane associated in infected cells. Thus, contribution to the formation of a membrane-bound scaffold for the viral replication-transcription complex appears to be an important novel function for the arterivirus ORF1a replicase polyprotein.
Viral Ribonucleoprotein Complex Formation and Nucleolar-Cytoplasmic Relocalization of Nucleolin in Poliovirus-Infected Cells
Authors: Waggoner Shelly , Sarnow Peter 
Published: December 2019

ABSTRACT
The poliovirus 3′ noncoding region (3′NCR) is involved in the efficient synthesis of viral negative-stranded RNA molecules. A strong interaction between a 105-kDa host protein and the wild-type 3′NCR, but not with a replication-defective mutant 3′NCR, was detected. This 105-kDa protein was identified as nucleolin which predominantly resides in the nucleolus and has been proposed to function in the folding of rRNA precursor molecules. A functional role for nucleolin in viral genome amplification was examined in a cell-free extract which has been shown to support the assembly of infectious virus from virion RNA. At early times of viral gene expression, extracts depleted of nucleolin produced less infectious virus than extracts depleted of fibrillarin, another resident of the nucleolus, indicating a functional role of nucleolin in the early stages of the viral life cycle in this in vitro system. Immunofluorescence analysis of uninfected and infected cells showed a nucleocytoplasmic relocalization of nucleolin, but not of fibrillarin, in poliovirus-infected cells. Relocalization of nucleolin was not simply a consequence of virally induced inhibition of translation or transcription, because inhibitors of translation or transcription did not induce nucleolar-cytoplasmic relocalization of nucleolin. These findings suggest a novel virus-induced mechanism by which certain nucleolar proteins are selectively redistributed in infected cells.
Human Thymidine Kinase Can Functionally Replace Herpes Simplex Virus Type 1 Thymidine Kinase for Viral Replication in Mouse Sensory Ganglia and Reactivation from Latency upon Explant
Authors: Chen Shun-Hua , Cook W. James , Grove Kristie L. , Coen Donald M. 
Published: December 2019

ABSTRACT
Herpes simplex virus type 1 thymidine kinase exhibits a strikingly broad substrate specificity. It is capable of phosphorylating deoxythymidine and deoxyuridine as does human thymidine kinase, deoxycytidine as does human deoxycytidine kinase, the cytosolic kinase whose amino acid sequence it most closely resembles, and thymidylate as does human thymidylate kinase. Following peripheral inoculation of mice, viral thymidine kinase is ordinarily required for viral replication in ganglia and for reactivation from latency following ganglionic explant. To determine which activity of the viral kinase is important for replication and reactivation in mouse ganglia, recombinant viruses lacking viral thymidine kinase but expressing individual human kinases were constructed. Each recombinant virus expressed the appropriate kinase activity with early kinetics following infection of cultured cells. The virus expressing human thymidine kinase exhibited thymidine phosphorylation activity equivalent to ∼5% of that of wild-type virus in a quantitative plaque autoradiography assay. Nevertheless, it was competent for ganglionic replication and reactivation following corneal inoculation of mice. The virus expressing human thymidylate kinase was partially competent for these activities despite failing to express detectable thymidine kinase activity. The virus expressing human deoxycytidine kinase failed to replicate acutely in neurons or to reactivate from latency. Therefore, it appears that low levels of thymidine phosphorylation suffice to fulfill the role of the viral enzyme in ganglia and that this role can be partially fulfilled by thymidylate kinase activity alone.
Infectious Laryngotracheitis Herpesvirus Expresses a Related Pair of Unique Nuclear Proteins Which Are Encoded by Split Genes Located at the Right End of the ULGenome Region
Authors: Ziemann Katharina , Mettenleiter Thomas C. , Fuchs Walter 
Published: December 2019

ABSTRACTAvian infectious laryngotracheitis virus (ILTV) possesses an alphaherpesvirus type D DNA genome of ca. 155 kbp. Completion of our previous sequence analyses (W. Fuchs and T. C. Mettenleiter, J. Gen. Virol. 77:2221–2229, 1996) of the right end of the unique long (UL) genome region revealed the presence of two adjacent, presumably ILTV-specific genes, which were named UL0 and UL[−1] because of their location upstream of the conserved UL1 (glycoprotein L) gene. Transcriptional analyses showed that both genes are abundantly expressed during the late phase of the viral replication cycle and that both mRNAs are spliced by the removal of short introns close to their 5′ ends. Furthermore, the deduced gene products exhibit a moderate but significant homology of 28% to each other. The newly identified ILTV genes encode proteins of 63 kDa (UL0) and 73 kDa (UL[−1]), which both are predominantly localized in the nuclei of virus infected chicken cells. In summary, our results indicate that duplication of a spliced ILTV-specific gene encoding a nuclear protein has occurred during evolution of ILTV.
Increased Susceptibility of Diabetic Mice to Influenza Virus Infection: Compromise of Collectin-Mediated Host Defense of the Lung by Glucose?
Authors: Reading Patrick C. , Allison Janette , Crouch Erika C. , Anders E. Margot 
Published: December 2019

ABSTRACT

The influence of diabetes on susceptibility to influenza virus infection was examined in a mouse model in which RIP-K
b
transgenic mice and their nontransgenic littermates were used as the diabetic and nondiabetic hosts, respectively. Influenza virus A/Phil/82 (H3N2) grew to significantly higher titers in the lungs of diabetic than nondiabetic mice. The extent of viral replication in the lungs was proportional to blood glucose levels in the mice at the time of infection, and the enhanced susceptibility of diabetic mice was reversed with insulin. Growth of A/HKx31 (H3N2) virus was also enhanced in diabetic mice, whereas the highly virulent strain A/PR/8/34 (H1N1) showed no difference in virus yields in diabetic and nondiabetic mice, even with low inocula. A/Phil/82 and A/HKx31 are sensitive to neutralization in vitro by the pulmonary collectin surfactant protein D (SP-D), whereas A/PR/8/34 is essentially resistant. Glucose is a ligand for SP-D, and neutralization of A/Phil/82 virus by SP-D was abolished in the presence of glucose at levels commonly found in diabetic mice. These findings suggest that in mice, and perhaps in humans, diabetes predisposes to influenza virus infection through compromise of collectin-mediated host defense of the lung by glucose.
Differential Expression of Nur77 Family Members in Human T-Lymphotropic Virus Type 1-Infected Cells: Transactivation of theTR3/nur77Gene by Tax Protein
Authors: Chen Xiaolin , Zachar Vladimir , Chang Chawnshang , Ebbesen Peter , Liu Xiangdong 
Published: December 2019

ABSTRACTWe analyzed the differential expression and regulation of three members of the Nur77 transcription factor family by the human T-lymphotropic virus type 1 (HTLV-1) Tax protein. We have demonstrated that in both HTLV-1-infected cells and Tax-expressing JPX-9 cells,TR3/nur77is highly expressed, whereas neitherNOR-1norNOTexpression is detectable. Transient transfection analysis further confirmed the Tax transactivation of theTR3/nur77promoter but not theNOR-1promoter in different cell types. Furthermore, expression of a luciferase reporter gene driven by the NGFI-B (rat homolog of TR3/Nur77) response element (NBRE) provided evidence that Tax-mediated transactivation resulted in the induction of a functional protein. Cotransfection assays with theTR3/nur77promoter sequence or the NBRE binding motif together with a series of Tax mutants have shown that Tax-inducedTR3/nur77expression is mediated by CREB/ATF-related transcription factors.
Apoptosis in Feline Panleukopenia Virus-Infected Lymphocytes
Authors: Ikeda Yasuhiro , Shinozuka Junko , Miyazawa Takayuki , Kurosawa Kyoko , Izumiya Yoshihiro , Nishimura Yorihiro , Nakamura Kazuya , Cai Jinshun , Fujita Kentaro , Doi Kunio , Mikami Takeshi 
Published: December 2019

ABSTRACTFeline panleukopenia virus (FPLV) was shown to induce apoptosis to feline lymphoid cells and to reduce the expression of interleukin-2 receptor α on the cells. FPLV-induced apoptosis might be a key element in the pathophysiology of atrophy of lymphoid tissues associated with feline panleukopenia caused by FPLV.
Ilarviruses Encode a Cucumovirus-Like 2b Gene That Is Absent in Other Genera within the
Bromoviridae
Authors: Xin Hong-Wu , Ji Liang-Hui , Scott Simon W. , Symons Robert H. , Ding Shou-Wei 
Published: December 2019

ABSTRACT
We found that RNA 2 of the four ilarviruses sequenced to date encodes an additional conserved open reading frame (ORF), 2b, that overlaps the 3′ end of the previously known ORF, 2a. A novel RNA species of 851 nucleotides was found to accumulate to high levels in plants infected with spinach latent virus (SpLV). Further analysis showed that RNA 4A is a subgenomic RNA of RNA 2 and encodes all of ORF 2b. Moreover, a protein species of the size expected for SpLV ORF 2b was translated in vitro from the RNA 4A-containing virion RNAs. The data support the suggestion that the SpLV 2b protein is translated in vivo. The 2b gene of ilarviruses, which is not encoded by alfamoviruses and bromoviruses, shares several features with the previously reported cucumovirus 2b gene; however, their encoded proteins share no detectable sequence similarities. The evolutionary origin of the 2b gene is discussed.
Recombination in the 5′ Leader of Murine Leukemia Virus Is Accurate and Influenced by Sequence Identity with a Strong Bias toward the Kissing-Loop Dimerization Region
Authors: Mikkelsen Jacob Giehm , Lund Anders H. , Duch Mogens , Pedersen Finn Skou 
Published: December 2019

ABSTRACT
Retroviral recombination occurs frequently during reverse transcription of the dimeric RNA genome. By a forced recombination approach based on the transduction of Akv murine leukemia virus vectors harboring a primer binding site knockout mutation and the entire 5′ untranslated region, we studied recombination between two closely related naturally occurring retroviral sequences. On the basis of 24 independent template switching events within a 481-nucleotide target sequence containing multiple sequence identity windows, we found that shifting from vector RNA to an endogenous retroviral RNA template during minus-strand DNA synthesis occurred within defined areas of the genome and did not lead to misincorporations at the crossover site. The nonrandom distribution of recombination sites did not reflect a bias for specific sites due to selection at the level of marker gene expression. We address whether template switching is affected by the length of sequence identity, by palindromic sequences, and/or by putative stem-loop structures. Sixteen of 24 sites of recombination colocalized with the kissing-loop dimerization region, and we propose that RNA-RNA interactions between palindromic sequences facilitate template switching. We discuss the putative role of the dimerization domain in the overall structure of the reverse-transcribed RNA dimer and note that related mechanisms of template switching may be found in remote RNA viruses.
Neutralization Profiles of Primary Human Immunodeficiency Virus Type 1 Isolates in the Context of Coreceptor Usage
Authors: Cecilia D. , KewalRamani Vineet N. , O’Leary Jeanne , Volsky Barbara , Nyambi Phillipe , Burda Sherri , Xu Serena , Littman Dan R. , Zolla-Pazner Susan 
Published: December 2019

ABSTRACTMost strains of human immunodeficiency virus type 1 (HIV-1) which have only been carried in vitro in peripheral blood mononuclear cells (primary isolates) can be neutralized by antibodies, but their sensitivity to neutralization varies considerably. To study the parameters that contribute to the differential neutralization sensitivity of primary HIV-1 isolates, we developed a neutralization assay with a panel of genetically engineered cell lines (GHOST cells) that express CD4, one of eight chemokine receptors which function as HIV-1 coreceptors, and a Tat-dependent green fluorescent protein reporter cassette which permits the evaluation and quantitation of HIV-1 infection by flow cytometry. All 21 primary isolates from several clades could grow in the various GHOST cell lines, and their use of one or more coreceptors could easily be defined by flow cytometric analysis. Ten of these primary isolates, three that were CXCR4 (X4)-tropic, three that were CCR5 (R5)-tropic, and four that were dual- or polytropic were chosen for study of their sensitivity to neutralization by human monoclonal and polyclonal antibodies. Viruses from the X4-tropic category of viruses were first tested since they have generally been considered to be particularly neutralization sensitive. It was found that the X4-tropic virus group contained both neutralization-sensitive and neutralization-resistant viruses. Similar results were obtained with R5-tropic viruses and with dual- or polytropic viruses. Within each category of viruses, neutralization sensitivity and resistance could be observed. Therefore, sensitivity to neutralization appears to be the consequence of factors that influence the antibody-virus interaction and its sequelae rather than coreceptor usage. Neutralization of various viruses by the V3-specific monoclonal antibody, 447-52D, was shown to be dependent not only on the presence of the relevant epitope but also on its presentation. An epitope within the envelope of a particular virus is not sufficient to render a virus sensitive to neutralization by an antibody that recognizes that epitope. Moreover, conformation-dependent factors may overcome the need for absolute fidelity in the match between an antibody and its core epitope, permitting sufficient affinity between the viral envelope protein and the antibody to neutralize the virus. The studies indicate that the neutralization sensitivity of HIV-1 primary isolates is a consequence of the complex interaction between virus, antibody, and target cell.
Characterization of a Baculovirus-Encoded RNA 5′-Triphosphatase
Authors: Gross Christian H. , Shuman Stewart 
Published: December 2019

ABSTRACT

Autographa californica
nuclear polyhedrosis virus (AcNPV) encodes a 168-amino-acid polypeptide that contains the signature motif of the superfamily of protein phosphatases that act via a covalent cysteinyl phosphate intermediate. The sequence of the AcNPV phosphatase is similar to that of the RNA triphosphatase domain of the metazoan cellular mRNA capping enzyme. Here, we show that the purified recombinant AcNPV protein is an RNA 5′-triphosphatase that hydrolyzes the γ-phosphate of triphosphate-terminated poly(A); it also hydrolyzes ATP to ADP and GTP to GDP. The phosphatase sediments as two discrete components in a glycerol gradient: a 9.5S oligomer and 2.5S putative monomer. The 2.5S form of the enzyme releases
32
P
i
from 1 μM γ-
32
P-labeled triphosphate-terminated poly(A) with a turnover number of 52 min
−1
and converts ATP to ADP with
V
max
of 8 min
−1
and

K
m

of 25 μM ATP. The 9.5S oligomeric form of the enzyme displays an initial pre-steady-state burst of ADP and P
i
formation, which is proportional to and stoichiometric with the enzyme, followed by a slower steady-state rate of product formation (approximately 1/10 of the steady-state rate of the 2.5S enzyme). We surmise that the oligomeric enzyme is subject to a rate-limiting step other than reaction chemistry and that this step is either distinct from or slower than the rate-limiting step for the 2.5S enzyme. Replacing the presumptive active site nucleophile Cys-119 by alanine abrogates RNA triphosphatase and ATPase activity. Our findings raise the possibility that baculoviruses encode enzymes that cap the 5′ ends of viral transcripts synthesized at late times postinfection by a virus-encoded RNA polymerase.
The Epstein-Barr Virus Major Latent Promoter Qp Is Constitutively Active, Hypomethylated, and Methylation Sensitive
Authors: Tao Qian , Robertson Keith D. , Manns Angela , Hildesheim Allan , Ambinder Richard F. 
Published: December 2019

ABSTRACT
Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) is indispensable for viral DNA replication and episome maintenance in latency. Four promoters, Cp, Wp, Qp, and Fp, are known to drive EBNA1 expression. Here we show that the TATA-less Qp is constitutively active in a variety of EBV-positive [EBV(+)] tumors and cell lines, irrespective of the activities of other EBNA1 promoters, the type of viral latency, and the cell type. The transcription of highly regulated promoters such as the EBV Cp is known to be directly regulated by CpG methylation. To characterize the role of CpG methylation in the regulation of the constitutively active Qp, we performed bisulfite genomic sequencing and functional analyses using a methylation cassette transcriptional reporter assay. Twenty consecutive CpG sites (16 proximal to the Qp initiation site and 4 upstream of the adjacent Fp initiation site) were studied by bisulfite sequencing of DNA extracted from EBV(+) tumors and cell lines. Eighteen EBV(+) tumors of lymphoid (B, T, and NK cell) or epithelial origin and five Burkitt’s lymphoma cell lines were studied. The 16 CpG sites proximal to Qp were virtually all unmethylated, but the 4 CpG sites upstream of the Fp initiation site were variably methylated. The methylation cassette assay showed that in vitro methylation of the Qp cassette (−172 to +32) resulted in strong repression of Qp activity in transient transfection. Thus, Qp is susceptible to repression by methylation but was found to be consistently hypomethylated and expressed in all tumors and tumor-derived cell lines studied.