bluuebunny/crossref_metadata_2025_split huggingface dataset:
Testing the Hypothesis of a Recombinant Origin of Human Immunodeficiency Virus Type 1 Subtype E
Authors: Anderson Jon P. , Rodrigo Allen G. , Learn Gerald H. , Madan Anup , Delahunty Claire , Coon Michael , Girard Marc , Osmanov Saladin , Hood Leroy , Mullins James I. 
Published: July 2002

ABSTRACT
The human immunodeficiency virus type 1 (HIV-1) epidemic in Southeast Asia has been largely due to the emergence of clade E (HIV-1E). It has been suggested that HIV-1E is derived from a recombinant lineage of subtype A (HIV-1A) and subtype E, with multiple breakpoints along the E genome. We obtained complete genome sequences of clade E viruses from Thailand (93TH057 and 93TH065) and from the Central African Republic (90CF11697 and 90CF4071), increasing the total number of HIV-1E complete genome sequences available to seven. Phylogenetic analysis of complete genomes showed that subtypes A and E are themselves monophyletic, although together they also form a larger monophyletic group. The apparent phylogenetic incongruence at different regions of the genome that was previously taken as evidence of recombination is shown to be not statistically significant. Furthermore, simulations indicate that bootscanning and pairwise distance results, previously used as evidence for recombination, can be misleading, particularly when there are differences in substitution or evolutionary rates across the genomes of different subtypes. Taken jointly, our analyses suggest that there is inadequate support for the hypothesis that subtype E variants are derived from a recombinant lineage. In contrast, many other HIV strains claimed to have a recombinant origin, including viruses for which only a single parental strain was employed for analysis, do indeed satisfy the statistical criteria we propose. Thus, while intersubtype recombinant HIV strains are indeed circulating, the criteria for assigning a recombinant origin to viral structures should include statistical testing of alternative hypotheses to avoid inappropriate assignments that would obscure the true evolutionary properties of these viruses.
The Region between Amino Acids 245 and 265 of the Bovine Leukemia Virus (BLV) Tax Protein Restricts Transactivation Not Only via the BLV Enhancer but Also via Other Retrovirus Enhancers
Authors: Tajima Shigeru , Aida Yoko 
Published: July 2002

ABSTRACT

Bovine leukemia virus (BLV) is associated with enzootic bovine leukosis and is closely related to human T-cell leukemia virus type 1 (HTLV-1). The Tax protein of BLV acts through the 5′ long terminal repeat (LTR) of BLV and activates the transcription of BLV. In this study, we amplified
tax
genes from BLV-infected cattle using PCR. We cloned the genes and monitored the transcriptional activities of the products. Seven independent mutant Tax proteins, with at least one amino acid substitution between residues 240 and 265, exhibited a markedly stronger ability to stimulate the viral LTR-directed transcription than the wild-type Tax protein. Analysis of chimeric Tax proteins derived from wild-type and mutant Tax proteins clearly demonstrated that a single substitution between residue 240 and 265 might be critical for the higher activities of the Tax mutant proteins. Furthermore, it appeared that transient expression of a Tax mutant protein was better able to increase the production of viral proteins and particles from a defective recombinant proviral clone of BLV than was wild-type Tax. Analysis of mutations within the U3 region of the LTR revealed that a cyclic AMP-responsive element in Tax-responsive element 2 might be sufficient for the enhanced activation mediated by the mutant proteins. In addition to the LTR of BLV, other viral enhancers, such as the enhancers of HTLV-1 and of mouse mammary tumor virus, which cannot be activated by wild-type BLV Tax protein, were activated by a Tax mutant protein. Our observations suggest that the transactivation activity and target sequence specificity of BLV Tax might be limited or negatively regulated by the region of the protein between amino acids 240 and 265.
Structural and Functional Analysis of the
Xestia c-nigrum
Granulovirus Matrix Metalloproteinase
Authors: Ko Rinkei , Okano Kazuhiro , Maeda Susumu 
Published: July 2002

ABSTRACT

Sequence analysis of the
Xestia c-nigrum
granulovirus (XcGV) genome identified an open reading frame encoding a 469-amino-acid (54-kDa) protein with over 30% amino acid sequence identity to a region of about 150 amino acids that includes the catalytic domains of human stromelysin 1 (Str1)/matrix metalloproteinase 3 (MMP-3) (EC
3.4.24.17
) and sea urchin hatching enzyme (HE). Stromelysin homologs have not been reported from baculoviruses or other viruses. Unlike human Str1 and sea urchin HE, the putative XcGV-MMP does not have a signal peptide and lacks the peptide motif involved in the cysteine switch that maintains other MMPs in an inactive form. The putative XcGV-MMP, however, possesses a conserved zinc-binding motif in its putative catalytic domain. The XcGV-MMP homolog was cloned, and a recombinant
Bombyx mori
nucleopolyhedrovirus (BmNPV) that expresses XcGV-MMP under the polyhedrin promoter was constructed. A distinct pattern of melanization was observed in
B. mori
larvae infected with MMP-expressing BmNPV. Fat body extracts from larvae overexpressing the 54-kDa recombinant MMP digested dye-impregnated collagen (Azocoll). The enzymatic activity was inhibited by two metalloproteinase inhibitors, EDTA and 1,10-phenanthroline. These results suggest that the XcGV MMP-3 gene homolog encodes a functional metalloproteinase.
Interaction between Herpes Simplex Virus Type 1 IE63 Protein and Cellular Protein p32
Authors: Bryant Helen E. , Matthews David A. , Wadd Sarah , Scott James E. , Kean Joy , Graham Susan , Russell William C. , Clements J. Barklie 
Published: July 2002

ABSTRACT
The herpes simplex virus type 1 (HSV-1) immediate-early gene IE63 (ICP27), the only HSV-1 regulatory gene with a homologue in every mammalian and avian herpesvirus sequenced so far, is a multifunctional protein which regulates transcriptional and posttranscriptional processes. One of its posttranscriptional effects is the inhibition of splicing of viral and cellular transcripts. We previously identified heterogeneous nuclear ribonucleoprotein (hnRNP) K and casein kinase 2 (CK2) as two protein partners of IE63 (H. Bryant et al., J. Biol. Chem. 274:28991–28998, 1999). Here, using a yeast two-hybrid assay, we identify another partner of IE63, the cellular protein p32. Confirmation of this interaction was provided by coimmunoprecipitation from virus-infected cells and recombinant p32 binding assays. A p32-hnRNP K-CK2 complex, which required IE63 to form, was isolated from HSV-1-infected cells, and coimmunoprecipitating p32 was phosphorylated by CK2. Expression of IE63 altered the cytoplasmic distribution of p32, with some now colocalizing with IE63 in the nuclei of infected and transfected cells. As p32 copurifies with splicing factors and can inhibit splicing, we propose that IE63 together with p32, possibly with other IE63 partner proteins, acts to disrupt or regulate pre-mRNA splicing. As well as contributing to host cell shutoff, this effect could facilitate splicing-independent nuclear export of viral transcripts.
Interaction between Hepatitis B Virus Core Protein and Reverse Transcriptase
Authors: Lott Lisa , Beames Burton , Notvall Lena , Lanford Robert E. 
Published: July 2002

ABSTRACT
Previous mutagenesis studies with hepatitis B virus (HBV) suggest that continued interactions with core are required for several steps in genomic replication. To examine core-polymerase (Pol) interactions, insect cells were coinfected with baculovirus constructs that independently expressed core and Pol. The results demonstrated several features with implications that core plays an interactive role with HBV Pol: (i) core coprecipitated with constructs expressing full-length Pol as well as the terminal protein (TP), reverse transcriptase (RT) and RNase H domains of Pol, independently; (ii) coprecipitation of core was not dependent on the presence of an epsilon stem-loop sequence; and (iii) core-Pol complexes migrated as intact capsid particles, as detected by sucrose gradient analysis. To analyze the structural and sequence requirements of core in recognition of Pol, a series of core mutants with two- to four-amino-acid insertions or carboxy-terminal deletions were assessed for Pol interaction. The results indicated that capsid formation is required but not sufficient for interaction with Pol and that the TP and RT domains of Pol have different requirements for interaction with core. To map the core binding sites on Pol, a panel of amino- and carboxy-terminal deletion mutants of the TP and RT domains of Pol were analyzed for interaction with core. At least three separate core binding sites on Pol were detected. This analysis begins to define basic requirements for core-Pol interactions, but further study is necessary to delineate the effects of these interactions on encapsidation and genome replication.
Genetic Manipulation of Arterivirus Alternative mRNA Leader-Body Junction Sites Reveals Tight Regulation of Structural Protein Expression
Authors: Pasternak Alexander O. , Gultyaev Alexander P. , Spaan Willy J. M. , Snijder Eric J. 
Published: July 2002

ABSTRACT

To express its structural proteins, the arterivirus
Equine arteritis virus
(EAV) produces a nested set of six subgenomic (sg) RNA species. These RNA molecules are generated by a mechanism of discontinuous transcription, during which a common leader sequence, representing the 5′ end of the genomic RNA, is attached to the bodies of the sg RNAs. The connection between the leader and body parts of an mRNA is formed by a short, conserved sequence element termed the transcription-regulating sequence (TRS), which is present at the 3′ end of the leader as well as upstream of each of the structural protein genes. With the exception of RNA3, only one body TRS was previously assumed to be used to join the leader and body of each EAV sg RNA. Here we show that for the synthesis of two other sg RNAs, RNA4 and RNA5, alternative leader-body junction sites that differ substantially in transcriptional activity are used. By site-directed mutagenesis of an EAV infectious cDNA clone, the alternative TRSs used to generate RNA3, -4, and -5 were inactivated, which strongly influenced the corresponding RNA levels and the production of infectious progeny virus. The relative amounts of RNA produced from alternative TRSs differed significantly and corresponded to the relative infectivities of the virus mutants. This strongly suggested that the structural proteins that are expressed from these RNAs are limiting factors during the viral life cycle and that the discontinuous step in sg RNA synthesis is crucial for the regulation of their expression. On the basis of a theoretical analysis of the predicted RNA structure of the 3′ end of the EAV genome, we propose that the local secondary RNA structure of the body TRS regions is an important factor in the regulation of the discontinuous step in EAV sg mRNA synthesis.
Role of the 3′ tRNA-Like Structure in Tobacco Mosaic Virus Minus-Strand RNA Synthesis by the Viral RNA-Dependent RNA Polymerase In Vitro
Authors: Osman T. A. M. , Hemenway C. L. , Buck K. W. 
Published: July 2002

ABSTRACTA template-dependent RNA polymerase has been used to determine the sequence elements in the 3′ untranslated region of tobacco mosaic virus RNA that are required for promotion of minus-strand RNA synthesis and binding to the RNA polymerase in vitro. Regions which were important for minus-strand synthesis were domain D1, which is equivalent to a tRNA acceptor arm; domain D2, which is similar to a tRNA anticodon arm; an upstream domain, D3; and a central core, C, which connects domains D1, D2, and D3 and determines their relative orientations. Mutational analysis of the 3′-terminal 4 nucleotides of domain D1 indicated the importance of the 3′-terminal CA sequence for minus-strand synthesis, with the sequence CCCA or GGCA giving the highest transcriptional efficiency. Several double-helical regions, but not their sequences, which are essential for forming pseudoknot and/or stem-loop structures in domains D1, D2, and D3 and the central core, C, were shown to be required for high template efficiency. Also important were a bulge sequence in the D2 stem-loop and, to a lesser extent, a loop sequence in a hairpin structure in domain D1. The sequence of the 3′ untranslated region upstream of domain D3 was not required for minus-strand synthesis. Template-RNA polymerase binding competition experiments showed that the highest-affinity RNA polymerase binding element region lay within a region comprising domain D2 and the central core, C, but domains D1 and D3 also bound to the RNA polymerase with lower affinity.
Functional Role of pX Open Reading Frame II of Human T-Lymphotropic Virus Type 1 in Maintenance of Viral Loads In Vivo
Authors: Bartoe Joshua T. , Albrecht Björn , Collins Nathaniel D. , Robek Michael D. , Ratner Lee , Green Patrick L. , Lairmore Michael D. 
Published: July 2002

ABSTRACT

Human T-lymphotropic virus type 1
(HTLV-1) causes adult T-cell leukemia/lymphoma and is associated with a variety of immune-mediated disorders. The role of four open reading frames (ORFs), located between
env
and the 3′ long terminal repeat of HTLV-1, in mediating disease is not entirely clear. By differential splicing, ORF II encodes two proteins, p13
II
and p30
II
, both of which have not been functionally defined. p13
II
localizes to mitochondria and may alter the configuration of the tubular network of this cellular organelle. p30
II
localizes to the nucleolus and shares homology with the transcription factors Oct-1 and -2, Pit-1, and POU-M1. Both p13
II
and p30
II
are dispensable for infection and immortalization of primary human and rabbit lymphocytes in vitro. To test the role of ORF II gene products in vivo, we inoculated rabbits with lethally irradiated cell lines expressing the wild-type molecular clone of HTLV-1 (ACH.1) or a clone containing selected mutations in ORF II (ACH.30/13.1). ACH.1-inoculated animals maintained higher HTLV-1-specific antibody titers than animals inoculated with ACH.30/13.1. Viral p19 antigen was transiently detected in ex vivo cultures of peripheral blood mononuclear cells (PBMC) from only two ACH.30/13.1-inoculated rabbits, while PBMC cultures from all ACH.1-inoculated rabbits routinely produced p19 antigen. In only three of six animals exposed to the ACH.p30
II
/p13
II
clone could provirus be consistently PCR amplified from extracted PBMC DNA and quantitative competitive PCR showed the proviral loads in PBMC from ACH.p30
II
/p13
II
-infected rabbits to be dramatically lower than the proviral loads in rabbits exposed to ACH. Our data indicate selected mutations in pX ORF II diminish the ability of HTLV-1 to maintain high viral loads in vivo and suggest an important function for p13
II
and p30
II
in viral pathogenesis.
Biochemical and Structural Analysis of Isolated Mature Cores of Human Immunodeficiency Virus Type 1
Authors: Welker Reinhold , Hohenberg Heinrich , Tessmer Uwe , Huckhagel Carola , Kräusslich Hans-Georg 
Published: July 2002

ABSTRACT
Mature human immunodeficiency virus type 1 (HIV-1) particles contain a cone-shaped core structure consisting of the internal ribonucleoprotein complex encased in a proteinaceous shell derived from the viral capsid protein. Because of their very low stability after membrane removal, HIV-1 cores have not been purified in quantities sufficient for structural and biochemical analysis. Based on our in vitro assembly experiments, we have developed a novel method for isolation of intact mature HIV-1 cores. Concentrated virus suspensions were briefly treated with nonionic detergent and immediately centrifuged in a microcentrifuge for short periods of time. The resuspended pellet was subsequently analyzed by negative-stain and thin-section electron microscopy and by immunoelectron microscopy. Abundant cone-shaped cores as well as tubular and aberrant structures were observed. Stereo images showed that core structures preserved their three-dimensional architecture and exhibited a regular substructure. Detailed analysis of 155 cores revealed an average length of ca. 103 nm, an average diameter at the base of ca. 52 nm, and an average angle of 21.3°. There was significant variability in all parameters, indicating that HIV cores are not homogeneous. Immunoblot analysis of core preparations allowed semiquantitative estimation of the relative amounts of viral and cellular proteins inside the HIV-1 core, yielding a model for the topology of various proteins inside the virion.
Two Widely Spaced Initiator Binding Sites Create an HMG1-Dependent Parvovirus Rolling-Hairpin Replication Origin
Authors: Cotmore Susan F. , Christensen Jesper , Tattersall Peter 
Published: July 2002

ABSTRACT
Minute virus of mice (MVM) replicates via a linearized form of rolling-circle replication in which the viral nickase, NS1, initiates DNA synthesis by introducing a site-specific nick into either of two distinct origin sequences. In vitro nicking and replication assays with substrates that had deletions or mutations were used to explore the sequences and structural elements essential for activity of one of these origins, located in the right-end (5′) viral telomere. This structure contains 248 nucleotides, most-favorably arranged as a simple hairpin with six unpaired bases. However, a pair of opposing NS1 binding sites, located near its outboard end, create a 33-bp palindrome that could potentially assume an alternate cruciform configuration and hence directly bind HMG1, the essential cofactor for this origin. The palindromic nature of this sequence, and thus its ability to fold into a cruciform, was dispensable for origin function, as was the NS1 binding site occupying the inboard arm of the palindrome. In contrast, the NS1 site in the outboard arm was essential for initiation, even though positioned 120 bp from the nick site. The specific sequence of the nick site and an additional NS1 binding site which directly orients NS1 over the initiation site were also essential and delimited the inboard border of the minimal right-end origin. DNase I and hydroxyl radical footprints defined sequences protected by NS1 and suggest that HMG1 allows the NS1 molecules positioned at each end of the origin to interact, creating a distortion characteristic of a double helical loop.
Productive Replication of Human Adenoviruses in Mouse Epidermal Cells
Authors: Ganly Ian , Mautner Vivien , Balmain Allan 
Published: July 2002

ABSTRACT
In contrast to most cells of mouse origin, cell lines derived from mouse epidermis are permissive for replication of human adenovirus type 5. The extent of epidermal cell differentiation correlated with the level of E1A expression and virus replication. Mouse epidermal cells may provide useful models for cancer therapy using replication-competent human adenoviruses.
Characterization of the R572T Point Mutant of a Putative Cleavage Site in Human Foamy Virus Env
Authors: Bansal Anju , Shaw Kit L. , Edwards Bradley H. , Goepfert Paul A. , Mulligan Mark J. 
Published: July 2002

ABSTRACTA putative cleavage site of the human foamy virus (HFV) envelope glycoprotein (Env) was altered. Transientenvexpression revealed that the R572T mutant Env was normally expressed and modified by asparagine-linked oligosaccharide chains. However, this single-amino-acid substitution was sufficient to abolish all detectable cleavage of the gp130 precursor polyprotein. Cell surface biotinylation demonstrated that the uncleaved mutant gp130 was transported to the plasma membrane. The uncleaved mutant protein was incapable of syncytium formation. Glycoprotein-driven virion budding, a unique aspect of HFV assembly, occurred despite the absence of Env cleavage. We then substituted the R572T mutantenvinto a replication-competent HFV molecular clone. Transfection of the mutant viral DNA into BHK-21 cells followed by viral titration with the FAB (foamy virus-activated β-galactosidase expression) assay revealed that proteolysis of the HFV Env was essential for viral infectivity. Wild-type HFV Env partially complemented the defective virus phenotype. Taken together, these experimental results established the location of the HFV Env proteolytic site; the effects of cleavage on Env transport, processing, and function; and the importance of Env proteolysis for virus maturation and infectivity.
Assembly and Processing of Human Immunodeficiency Virus Gag Mutants Containing a Partial Replacement of the Matrix Domain by the Viral Protease Domain
Authors: Wang Chin-Tien , Chou Yen-Chiou , Chiang Chien-Cheng 
Published: July 2002

ABSTRACT

We constructed human immunodeficiency virus (HIV) mutants by replacing the matrix domain with sequences encoding the viral protease or p6* and protease. The chimeras retaining matrix myristylation and processing signals underwent efficient autoprocessing with severely defective particle budding. The budding defects of the chimeras were rescued by suppressing the chimera protease activity either through addition of an HIV protease inhibitor or through inactivating the chimera protease via a substitution mutation of the catalytic aspartic acid residue. This resulted in the release of chimeric virus-like particles with the density of a wild-type retrovirus particle. In addition, the assembly-competent but processing-defective chimeras produced proteolytically processed particles with significant reverse transcriptase activity when a downstream native
pol
gene was present. These results suggest that HIV has the potential to adapt heterologous sequences in place of the matrix sequence without major effects on virus-like particle budding. In addition, the positions of the protease and substrate accessibility may contribute significantly toward avoiding a premature Gag or Gag-Pol process, which leads to severe defects in both particle budding and incorporation.
Mutations in the 2C Region of Poliovirus Responsible for Altered Sensitivity to Benzimidazole Derivatives
Authors: Shimizu Hiroyuki , Agoh Masanobu , Agoh Yumi , Yoshida Hiromu , Yoshii Kumiko , Yoneyama Tetsuo , Hagiwara Akio , Miyamura Tatsuo 
Published: July 2002

ABSTRACT

MRL-1237, [1-(4-fluorophenyl)-2-(4-imino-1,4-dihydropyridin-1-yl) methylbenzimidazole hydrochloride], is a potent and selective inhibitor of the replication of enteroviruses. To reveal the target molecule of MRL-1237 in viral replication, we selected spontaneous MRL-1237-resistant poliovirus mutants. Of 15 MRL-1237-resistant mutants obtained, 14 were cross-resistant to guanidine hydrochloride (
mrgr
), while 1 was susceptible (
mrgs
). Sequence analysis of the 2C region revealed that the 14
mrgr
mutants contained a single nucleotide substitution that altered an amino acid residue from Phe-164 to Tyr. The
mrgs
mutant, on the other hand, contained a substitution of Ile-120 to Val. Through the construction of a cDNA-derived mutant, we confirmed that the single mutation at Phe-164 was really responsible for the reduced susceptibility to MRL-1237. MRL-1237 inhibited poliovirus-specific RNA synthesis in HeLa cells infected with a wild strain but not with an F164Y mutant. We furthermore examined the effect of mutations of the 2C region on the drug sensitivity of cDNA-derived guanidine-resistant and -dependent mutants. Two guanidine-resistant mutants were cross-resistant to MRL-1237 but remained susceptible to another benzimidazole, enviroxime. Either MRL-1237 or guanidine stimulated the viral replication of two guanidine-dependent mutants, but enviroxime did not. These results indicate that MRL-1237, like guanidine, targets the 2C protein of poliovirus for its antiviral effect.