bluuebunny/crossref_metadata_2025_split huggingface dataset:
Human Parainfluenza Virus Type 1 Matrix and Nucleoprotein Genes Transiently Expressed in Mammalian Cells Induce the Release of Virus-Like Particles Containing Nucleocapsid-Like Structures
Authors: Coronel Elizabeth C. , Murti K. Gopal , Takimoto Toru , Portner Allen 
Published: December 2019

ABSTRACT
The matrix (M) protein plays an essential role in the assembly and budding of some enveloped RNA viruses. We expressed the human parainfluenza virus type 1 (hPIV-1) M and/or NP genes into 293T cells using the mammalian expression vector pCAGGS. Biochemical and electron microscopic analyses of transfected cells showed that the M protein alone can induce the budding of virus-like particles (vesicles) from the plasma membrane and that the NP protein can assemble into intracellular nucleocapsid-like (NC-like) structures. Furthermore, the coexpression of both the M and NP genes resulted in the production of vesicles enclosing NC-like structures, suggesting that the hPIV-1 M protein has the intrinsic ability to induce membrane vesiculation and to incorporate NC-like structures into these budding vesicles.
The Integration Machinery of ZAM, a Retroelement from
Drosophila melanogaster
, Acts as a Sequence-Specific Endonuclease
Authors: Leblanc Pascal , Dastugue Bernard , Vaury Chantal 
Published: December 2019

ABSTRACT

Retroviruses and retrotransposons insert into the host genome with no obvious sequence specificity. We examined the target sites of the retroelement ZAM by sequencing each host-ZAM junction in the genome of
Drosophila melanogaster
. Our overall data provide compelling evidence that ZAM integration machinery recognizes and leads to ZAM insertion into the sequence 5′-GCGCGCg-3′. This unique property of ZAM will facilitate the development of new tools to study the integration process of retroelements.
Expression of the Coxsackievirus and Adenovirus Receptor in Cultured Human Umbilical Vein Endothelial Cells: Regulation in Response to Cell Density
Authors: Carson Steven D. , Hobbs Justin T. , Tracy Steven M. , Chapman Nora M. 
Published: December 2019

ABSTRACT
Primary cultures of human umbilical vein endothelial cells (HUVEC) express the human coxsackievirus and adenovirus receptor (HCAR). Whereas HCAR expression in HeLa cells was constant with respect to cell density, HCAR expression in HUVEC increased with culture confluence. HCAR expression in HUVEC was not quantitatively altered by infection with coxsackievirus B.
Interaction of Human Immunodeficiency Virus-Derived Vectors with Wild-Type Virus in Transduced Cells
Authors: Bukovsky Anatoly A. , Song Jin-Ping , Naldini Luigi 
Published: December 2019

ABSTRACT
The interaction of human immunodeficiency virus (HIV)-derived vectors with wild-type virus was analyzed in transduced cells. Vector transcripts upregulated by infection had no measurable effect on HIV type 1 (HIV-1) expression but competed efficiently for encapsidation, inhibiting the infectivity and spread of HIV-1 in culture and leading to mobilization and recombination of the vector. These effects were abrogated with a self-inactivating vector.
Unique Properties of the Large Antigen of Hepatitis Delta Virus
Authors: Moraleda Gloria , Seeholzer Steven , Bichko Vadim , Dunbrack Roland , Otto James , Taylor John 
Published: December 2019

ABSTRACT

The large form of the hepatitis delta virus (HDV) protein (L) can be isoprenylated near its C terminus, and this modification is considered essential for particle assembly. Using gel electrophoresis, we separated L into two species of similar mobilities. The slower species could be labeled by the incorporation of [
14
C]mevalonolactone and is interpreted to be isoprenylated L (L
i
). In serum particles, infected liver, transfected cells, and assembled particles, 25 to 85% of L was isoprenylated. Isoprenylation was also demonstrated by
14
C incorporation in vitro with a rabbit reticulocyte coupled transcription-translation system. However, the species obtained migrated even slower than that detected by labeling in vivo. Next, in studies of HDV particle assembly in the presence of the surface proteins of human hepatitis B virus, we observed the following. (i) Relative to L, L
i
was preferentially assembled into virus-like particles. (ii) L
i
could coassemble the unmodified L and the small delta protein, S. (iii) In contrast, a form of L with a deletion in the dimerization domain was both isoprenylated and assembled, but it could not support the coassembly of S. Finally, to test the expectation that the isoprenylation of L would increase its hydrophobicity, we applied a phase separation strategy based on micelle formation with the nonionic detergent Triton X-114. We showed the following. (i) The unique C-terminal 19 amino acids present on L relative to S caused a significant increase in the hydrophobicity. (ii) This increase was independent of isoprenylation. (iii) In contrast, other, artificial modifications at either the N or C terminus of S did not increase the hydrophobicity. (iv) The increased hydrophobicity was not sufficient for particle assembly; nevertheless, we speculate that it might facilitate virion assembly.
Effect of Retreatment with Interferon Alone or Interferon plus Ribavirin on Hepatitis C Virus Quasispecies Diversification in Nonresponder Patients with Chronic Hepatitis C
Authors: Gerotto Martina , Sullivan Daniel G. , Polyak Stephen J. , Chemello Liliana , Cavalletto Luisa , Pontisso Patrizia , Alberti Alfredo , Gretch David R. 
Published: December 2019

ABSTRACT

Alpha interferon (IFN-α) treatment is effective on a long-term basis in only 15 to 25% of patients with chronic hepatitis C. The results of recent trials indicate that response rates can be significantly increased when IFN-α is given in combination with ribavirin. However, a large number of patients do not respond even to combination therapy. Nonresponsiveness to IFN is characterized by evolution of the hepatitis C virus (HCV) quasispecies. Little is known about the changes occurring within the HCV genomes when nonresponder patients are retreated with IFN or with IFN plus ribavirin. In the present study we have examined the genetic divergence of HCV quasispecies during unsuccessful retreatment with IFN or IFN plus ribavirin. Fifteen nonresponder patients with HCV-1 (4 patients with HCV-1a and 11 patients with HCV-1b) infection were studied while being retreated for 2 months (phase 1) with IFN-α (6 MU given three times a week), followed by IFN plus ribavirin or IFN alone for an additional 6 months (phase 2). HCV quasispecies diversification in the E2 hypervariable region-1 (HVR1) and in the putative NS5A IFN sensitivity determining region (ISDR) were analyzed for phase 1 and phase 2 by using the heteroduplex tracking assay and clonal frequency analysis techniques. A major finding of this study was the relatively rapid evolution of the HCV quasispecies observed in both treatment groups during the early phase 1 compared to the late phase 2 of treatment. The rate of quasispecies diversification in HVR1 was significantly higher during phase 1 versus phase 2 both in patients who received IFN plus ribavirin (
P
= 0.017) and in patients who received IFN alone (
P
= 0.05). A trend toward higher rates of quasispecies evolution in the ISDR was also observed during phase 1 in both groups, although the results did not reach statistical significance. However, the NS5A quasispecies appeared to be rather homogeneous and stable in most nonresponder patients, suggesting the presence of a single well-fit major variant, resistant to antiviral treatment, in agreement with published data which have identified an IFN sensitivity determinant region within the NS5A. During the entire 8 months of retreatment, there was no difference in the rate of fixation of mutation between patients who received combination therapy and patients who were treated with IFN alone, suggesting that ribavirin had no major effects on the evolution of the HCV quasispecies after the initial 2 months of IFN therapy.
Tyrosine Phosphorylation of A17 during Vaccinia Virus Infection: Involvement of the H1 Phosphatase and the F10 Kinase
Authors: Derrien M. , Punjabi A. , Khanna M. , Grubisha O. , Traktman P. 
Published: December 2019

ABSTRACTVaccinia virus encodes two protein kinases (B1 and F10) and a dual-specificity phosphatase (VH1), suggesting that phosphorylation and dephosphorylation of substrates on serine/threonine and tyrosine residues are important in regulating diverse aspects of the viral life cycle. Using a recombinant in which expression of the H1 phosphatase can be regulated experimentally (vindH1), we have previously demonstrated that repression of H1 leads to the maturation of noninfectious virions that contain several hyperphosphorylated substrates (K. Liu et al., J. Virol. 69:7823–7834). In this report, we demonstrate that among these is a 25-kDa protein that is phosphorylated on tyrosine residues in H1-deficient virions and can be dephosphorylated by recombinant H1. We demonstrate that the 25-kDa phosphoprotein represents the product of the A17 gene and that A17 is phosphorylated on serine, threonine, and tyrosine residues during infection. Detection of phosphotyrosine within A17 is abrogated when Tyr203(but not Tyr3, Tyr6, or Tyr7) is mutated to phenylalanine, suggesting strongly that this amino acid is the site of tyrosine phosphorylation. Phosphorylation of A17 fails to occur during nonpermissive infections performed with temperature-sensitive mutants defective in the F10 kinase. Our data suggest that this enzyme, which was initially characterized as a serine/threonine kinase, might in fact have dual specificity. This hypothesis is strengthened by the observation thatEscherichia coliinduced to express F10 contain multiple proteins which are recognized by antiphosphotyrosine antiserum. This study presents the first evidence for phosphotyrosine signaling during vaccinia virus infection and implicates the F10 kinase and the H1 phosphatase as the dual-specificity enzymes that direct this cycle of reversible phosphorylation.
Characterization of the
oriI
and
oriII
Origins of Replication in Phage-Plasmid P4
Authors: Tocchetti Arianna , Galimberti Gloria , Dehò Gianni , Ghisotti Daniela 
Published: December 2019

ABSTRACT

In the
Escherichia coli
phage-plasmid P4, two partially overlapping replicons with bipartite
ori
sites coexist. The essential components of the
oriI
replicon are the α and
cnr
genes and the
ori1
and
crr
sites; the
oriII
replicon is composed of the α gene, with the internal
ori2
site, and the
crr
region. The P4 α protein has primase and helicase activities and specifically binds type I iterons, present in
ori1
and
crr
. Using a complementation test for plasmid replication, we demonstrated that the two replicons depend on both the primase and helicase activities of the α protein. Moreover, neither replicon requires the host DnaA, DnaG, and Rep functions. The bipartite origins of the two replicons share the
crr
site and differ for
ori1
and
ori2
, respectively. By deletion mapping, we defined the minimal
ori1
and
ori2
regions sufficient for replication. The
ori1
site was limited to a 123-bp region, which contains six type I iterons spaced regularly close to the helical periodicity, and a 35-bp AT-rich region. Deletion of one or more type I iterons inactivated
oriI
. Moreover, insertion of 6 or 10 bp within the
ori1
region also abolished replication ability, suggesting that the relative arrangement of the iterons is relevant. The
ori2
site was limited to a 36-bp P4 region that does not contain type I iterons. In vitro, the α protein did not bind
ori2
. Thus, the α protein appears to act differently at the two origins of replication.
Inhibitory Activity of Constitutive Nitric Oxide on the Expression of Alpha/Beta Interferon Genes in Murine Peritoneal Macrophages
Authors: Guillemard Eric , Varano Barbara , Belardelli Filippo , Quero A. M. , Gessani Sandra 
Published: December 2019

ABSTRACTWe investigated the role of the constitutive nitric oxide (NO) in the expression of interferon (IFN) genes in mouse peritoneal macrophages (PM). The treatment of PM withl-arginine-NG-amine (AA), a potent inhibitor of NO-producing enzymes, resulted in a marked accumulation of IFN-α4 mRNA and, to a minor extent, of IFN-β mRNA. In contrast, the expression of IFN-γ mRNA, as well as tumor necrosis factor alpha and interleukin-6 mRNA, was not affected. Furthermore, a remarkable increase in the expression of the IFN regulating factor 1 (IRF-1), but not of IRF-2, mRNA was detected in AA-treated PM. To investigate whether the AA-induced activation of the IFN system correlates with the production and antiviral activity of IFN, the extent of encephalomyocarditis virus (EMCV) replication was monitored in AA-treated PM with respect to control cultures. AA treatment strongly inhibited, in a dose-dependent manner, EMCV yields in PM. Likewise, similar results were obtained by the addition of the NO-scavenger carboxyphenyl-tetramethylimidazoline-oxyl-oxide. In addition, inhibition of NO synthesis byNG-mono-methyl-l-arginine in PM strongly decreased virus replication in coculture of PM and EMCV-infected L929 cells, whereas no antiviral effect was observed in L929 cells alone. Moreover, the AA-mediated antiviral activity was abrogated in the presence of antibody to IFN-α/β, whereas antibody to IFN-γ was completely ineffective. Taken together, these results indicate that low levels of NO, constitutively released by resting PM, negatively regulate the expression and activity of IFN-α/β in PM. We suggest that NO acts as a homeostatic agent in the regulation of IFN pathway expression in macrophages.
Point Mutations in the Avian Sarcoma/Leukosis Virus 3′ Untranslated Region Result in a Packaging Defect
Authors: Aschoff Jonathan M. , Foster Douglas , Coffin John M. 
Published: December 2019

ABSTRACT

The 3′ untranslated region (3′ UTR) between the 3′ end of
env
and the long terminal repeat is well conserved among avian retroviruses and is essential for efficient replication. Deletion of the dr1 element within the 3′ UTR has been reported to have various effects, including reduced levels of unspliced RNA in the cytoplasm, decreased stability of unspliced RNA, decreased particle production, and decreased genomic RNA packaging. To probe the role of specific sequences within dr1 in virus replication, site-directed mutagenesis was utilized to perturb parts of the predicted secondary structure of dr1. Seven of thirteen mutations had no significant effect; the others resulted in an approximately 10- to 20-fold reduction in replication. These mutants were further characterized and found to impair cytoplasmic accumulation of unspliced RNA only slightly. Furthermore, no decreases were observed in the stability of the unspliced RNA or in the production of virus particles. Genomic RNA packaging, however, was reduced by about 10-fold. Similar amounts of particles were produced by cells containing the mutant and wild-type DNA, and all particles contained similar levels of reverse transcriptase activity. The results suggest that the region of the dr1 disrupted by the mutations plays a role in genomic RNA packaging, although that packaging may not be the only role for dr1.
An Inducible Human Immunodeficiency Virus Type 1 (HIV-1) Vector Which Effectively Suppresses HIV-1 Replication
Authors: An Dong Sung , Morizono Kouki , Li Qi-Xiang , Mao Si Hua , Lu Stephanie , Chen Irvin S. Y. 
Published: December 2019

ABSTRACT

Recently, gene therapy vectors based upon the human immunodeficiency virus type 1 (HIV-1) genome have been developed. Here, we create an HIV-1 vector which is defective for all HIV-1 genes, but which maintains
cis
-acting elements required for efficient packaging, infection, and expression. In T cells transduced by this vector, vector expression is low but efficiently induced following HIV-1 infection. Remarkably, although the HIV-1 vector does not contain specific anti-HIV-1 therapeutic genes, the presence of the vector alone is sufficient to inhibit the spread of HIV-1 infection. The mechanism of inhibition is likely to be at the level of competition for limiting substrates required for either efficient packaging or reverse transcription, thereby selecting against propagation of wild-type HIV-1. These results provide proof of a concept for potential application of a novel HIV-1 vector in HIV-1 disease.
Evolution and Horizontal Transfer of dUTPase-Encoding Genes in Viruses and Their Hosts
Authors: Baldo Angela M. , McClure Marcella A. 
Published: December 2019

ABSTRACT

dUTPase is a ubiquitous and essential enzyme responsible for regulating cellular levels of dUTP. The
dut
gene exists as single, tandemly duplicated, and tandemly triplicated copies. Crystallized single-copy dUTPases have been shown to assemble as homotrimers. dUTPase is encoded as an auxiliary gene in a number of virus genomes. The origin of viral
dut
genes has remained unresolved since their initial discovery. A comprehensive analysis of dUTPase amino acid sequence relationships was performed to explore the evolutionary dynamics of
dut
in viruses and their hosts. Our data set, comprised of 24 host and 51 viral sequences, includes representative sequences from available eukaryotes, archaea, eubacteria cells, and viruses, including herpesviruses. These amino acid sequences were aligned by using a hidden Markov model approach developed to align divergent data. Known secondary structures from single-copy crystals were mapped onto the aligned duplicate and triplicate sequences. We show how duplicated dUTPases might fold into a monomer, and we hypothesize that triplicated dUTPases also assemble as monomers. Phylogenetic analysis revealed at least five viral dUTPase sequence lineages in well-supported monophyletic clusters with eukaryotic, eubacterial, and archaeal hosts. We have identified all five as strong examples of horizontal transfer as well as additional potential transfer of
dut
genes among eubacteria, between eubacteria and viruses, and between retroviruses. The evidence for horizontal transfers is particularly interesting since eukaryotic
dut
genes have introns, while DNA virus
dut
genes do not. This implies that an intermediary retroid agent facilitated the horizontal transfer process between host mRNA and DNA viruses.
Three-Dimensional Structure of Herpes Simplex Virus Type 1 Glycoprotein D at 2.4-Nanometer Resolution
Authors: Pilling Andrew , Rosenberg Mark F. , Willis Sharon H. , Jäger Joachim , Cohen Gary H. , Eisenberg Roselyn J. , Meredith David M. , Holzenburg Andreas 
Published: December 2019

ABSTRACT
Herpes simplex virus type 1 glycoprotein D (gD) is essential for virus infectivity and is responsible for binding to cellular membrane proteins and subsequently promoting fusion between the virus envelope and the cell. No structural data are available for gD or for any other herpesvirus envelope protein. Here we present a three-dimensional model for the baculovirus-expressed truncated protein gD1(306t) based on electron microscopic data. We demonstrate that gD1(306t) appears as a homotetramer containing a pronounced pocket in the center of the molecule. Monoclonal antibody binding demonstrates that the molecule is oriented such that the pocket protrudes away from the virus envelope.
Middle T Antigen Activation of Signal Transduction Pathways Does Not Overcome p53-Mediated Growth Arrest
Authors: Doherty Joanne , Freund Robert 
Published: December 2019

ABSTRACT

Polyomavirus middle T antigen does not overcome p53-mediated G
1
arrest in mouse embryo fibroblasts. Middle T antigen still associates with the signaling molecules phosphatidylinositol 3-kinase and SHC and activates the transcriptional activity of c-Myc and AP1 in p53-arrested cells. Examination of cell cycle regulatory proteins indicated that p53 does not interfere with these mitogenic signals but acts later in the G
1
phase of the cell cycle.
Creating and Maintaining the Gastrointestinal Ecosystem: What We Know and Need To Know from Gnotobiology
Authors: Falk Per G. , Hooper Lora V. , Midtvedt Tore , Gordon Jeffrey I. 
Published: December 2019

SUMMARY
Studying the cross talk between nonpathogenic organisms and their mammalian hosts represents an experimental challenge because these interactions are typically subtle and the microbial societies that associate with mammalian hosts are very complex and dynamic. A large, functionally stable, climax community of microbes is maintained in the murine and human gastrointestinal tracts. This open ecosystem exhibits not only regional differences in the composition of its microbiota but also regional differences in the differentiation programs of its epithelial cells and in the spatial distribution of its component immune cells. A key experimental strategy for determining whether “nonpathogenic” microorganisms actively create their own regional habitats in this ecosystem is to define cellular function in germ-free animals and then evaluate the effects of adding single or several microbial species. This review focuses on how gnotobiotics—the study of germ-free animals—has been and needs to be used to examine how the gastrointestinal ecosystem is created and maintained. Areas discussed include the generation of simplified ecosystems by using genetically manipulatable microbes and hosts to determine whether components of the microbiota actively regulate epithelial differentiation to create niches for themselves and for other organisms; the ways in which gnotobiology can help reveal collaborative interactions among the microbiota, epithelium, and mucosal immune system; and the ways in which gnotobiology is and will be useful for identifying host and microbial factors that define the continuum between nonpathogenic and pathogenic. A series of tests of microbial contributions to several pathologic states, using germ-free and ex-germ-free mice, are proposed.
Retroviral DNA Integration
Authors: Hindmarsh Patrick , Leis Jonathan 
Published: December 2019

SUMMARY
DNA integration is a unique enzymatic process shared by all retroviruses and retrotransposons. During integration, double-stranded linear viral DNA is inserted into the host genome in a process catalyzed by the virus-encoded integrase (IN). The mechanism involves a series of nucleophillic attacks, the first of which removes the terminal 2 bases from the 3′ ends of the long terminal repeats and of the second which inserts the viral DNA into the host genome. IN specifically recognizes the DNA sequences at the termini of the viral DNA, juxtaposing both ends in an enzyme complex that inserts the viral DNA into a single site in a concerted manner. Small duplications of the host DNA, characteristic of the viral IN, are found at the sites of insertion. At least two host proteins, HMG-I(Y) and BAF, have been shown to increase the efficiency of the integration reaction.