bluuebunny/crossref_metadata_2025 huggingface dataset:
A single antigenomic open reading frame of the hepatitis delta virus encodes the epitope(s) of both hepatitis delta antigen polypeptides p24 delta and p27 delta
Authors: Weiner A J , Choo Q L , Wang K S , Govindarajan S , Redeker A G , Gerin J L , Houghton M
Published: January 2020

On the basis of the complete nucleotide sequence of the single-stranded, covalently closed circular hepatitis delta virus RNA genome (K.-S. Wang, Q.-L. Choo, A. J. Weiner, J.-H. Ou, R. C. Najarian, R. M. Thayer, G. T. Mullenbach, K. J. Denniston, J. L. Gerin, and M. Houghton, Nature [London] 323:508-514, 1986 [Author's correction, 328:456, 1987]), five long open reading frames (ORFs) encoding polypeptides containing a methionine proximal to the amino terminus were expressed in bacteria. Only polypeptides encoded by the antigenomic ORF5 cross-reacted with antisera obtained from patients with hepatitis delta virus infections. Immunological analysis of viral extracts and the recombinant ORF5 polypeptides synthesized in bacteria and yeast cells revealed that ORF5 encodes the immunogenic epitope(s) shared by both hepatitis delta viral polypeptides p27 delta and p24 delta and probably represents the complete structural gene for p27 delta and p24 delta. We also present evidence that ORF5 encodes the hepatitis delta antigen, an antigen originally found in the nuclei of hepatocytes of infected individuals (M. Rizzetto, M. G. Canese, S. Arico, O. Crivelli, F. Bonino, C. G. Trepo, and G. Verme, Gut 18:997-1003, 1977). A comparison of the primary structure of the predicted hepatitis delta antigen polypeptides with that of the core antigen of the hepatitis B virus shows that these polypeptides are very dissimilar.
Role of endogenous alpha/beta interferon in the selection of virus nonproducer Friend leukemia cells after serial intraperitoneal passages in syngeneic mice
Authors: Ferrantini M , Belardelli F , Locardi C
Published: January 2020

Serial intraperitoneal passage of interferon (IFN)-sensitive Friend leukemia cells (FLCs) and L1210-S and RBL-5 tumor cells in syngeneic mice resulted in the selection of tumor cells exhibiting a marked decrease in the capacity to release reverse transcriptase (RT) activity. The virus nonproducer phenotype was a stable characteristic of clones derived from in vivo-passaged IFN-sensitive 745 FLCs. In contrast, in vivo passages of IFN-resistant 3Cl-8 FLCs and L1210-R cells did not result in any significant decrease in the capacity of these tumor cells to release in vitro RT activity. Although in vitro treatment of IFN-sensitive FLCs with mouse alpha/beta IFN (IFN-alpha/beta) for 1 or 10 passages resulted in a marked inhibition in the release of RT activity, these effects were completely reversible after removal of IFN from the culture medium. In addition, in vitro treatment of 745 FLCs with IFN resulted in a marked increase in the expression of H-2 (class I) and gp70 Friend virus antigens on the cell membrane. These effects were not observed in IFN-resistant 3Cl-8 cells. To investigate the possible role of endogenous IFN in the in vivo selection of virus nonproducer tumor cells, IFN-sensitive virus producer FLCs were serially passaged intraperitoneally in mice treated with antibodies to IFN-alpha/beta and in control mice, and the recovered tumor cells were cloned in vitro. Most (83 to 91%) of the clones derived from 745 cells recovered from control mice did not produce any detectable RT activity in the culture supernatants. In contrast, 96% of the clones (26 of 27) derived from 745 cells recovered from mice serially treated with antibodies to IFN-alpha/beta were still capable of releasing high levels of RT activity in the culture medium, indicating that endogenous IFN-alpha/beta was indeed an important host component for the in vivo selection of virus nonproducer tumor cell variants. The results reported in this article indicate that both direct effects of IFN on tumor cells and host-mediated effects are involved in this phenomenon.
Rearrangement of a common cellular DNA domain on chromosome 4 in human primary liver tumors
Authors: Pasquinelli C , Garreau F , Bougueleret L , Cariani E , Grzeschik K H , Thiers V , Croissant O , Hadchouel M , Tiollais P , Bréchot C
Published: January 2020

Hepatitis B virus (HBV) DNA integration has been shown to occur frequently in human hepatocellular carcinomas. We have investigated whether common cellular DNA domains might be rearranged, possibly by HBV integration, in human primary liver tumors. Unique cellular DNA sequences adjacent to an HBV integration site were isolated from a patient with hepatitis B surface antigen-positive hepatocellular carcinoma. These probes detected rearrangement of this cellular region of chromosomal DNA in 3 of 50 additional primary liver tumors studied. Of these three tumor samples, two contained HBV DNA, without an apparent link between the viral DNA and the rearranged allele; HBV DNA sequences were not detected in the third tumor sample. By use of a panel of somatic cell hybrids, these unique cellular DNA sequences were shown to be located on chromosome 4. Therefore, this region of chromosomal DNA might be implicated in the formation of different tumors at one step of liver cell transformation, possibly related to HBV integration.
Isolation of an integrated provirus of Moloney murine leukemia virus with long terminal repeats in inverted orientation: integration utilizing two U3 sequences
Authors: Colicelli J , Goff S P
Published: January 2020

We have previously described the construction of a mutant of Moloney murine leukemia virus, in594-2, which carries a 2-base-pair insertion in the U5 region of the genome and is partially defective in forming the integrated proviral DNA. We have now recovered a cloned copy of an unusual provirus from rat cells infected with this mutant. The viral genome is flanked by long terminal repeats in inverted orientation, with U3 sequences joined to cellular DNA at both of the outer edges. In addition, the provirus is a recombinant, containing a segment of a VL30 element in inverted orientation in place of the Moloney murine leukemia virus env region. The recovery of this provirus indicates that two U3 regions can be used for viral integration and suggests that there may be no absolute requirement in the reaction for those U5 sequences outside the 13-base-pair inverted repeats.
Tissue preferential expression of the hepatitis B virus (HBV) surface antigen gene in two lines of HBV transgenic mice
Authors: Burk R D , DeLoia J A , elAwady M K , Gearhart J D
Published: January 2020

Two transgenic mice were produced by microinjection of the entire hepatitis B virus (HBV) genome as a 3.2-kilobase EcoRI DNA fragment into one-cell embryos. Each animal contained a single, unique locus of HBV sequence. One founder animal, G7, contained a partially deleted HBV genome lacking both putative HBV surface antigen (HBsAg) promoters. The other animal, G26, contained greater-than-genome-length HBV sequences organized as a partial head-to-tail dimer. Both transgenic animals transmitted the HBV sequences in a Mendelian fashion, and all subsequent transgenic animals had detectable HBsAg in the serum. Expression of HBV sequences in tissues from G7- and G26-derived mice showed preferential expression of the 2.1-kilobase HBsAg RNA transcript in liver and kidney tissues by Northern (RNA) blot analysis. These data are consistent with the notion that HBV DNA contains cis-acting regulatory sequences which are responsible for the predominant expression of HBsAg transcripts in the liver and kidney of transgenic mice.
Characterization of vaccinia virus growth factor biosynthetic pathway with an antipeptide antiserum
Authors: Chang W , Lim J G , Hellström I , Gentry L E
Published: January 2020

A synthetic peptide derived from vaccinia virus growth factor (VGF) was used as an immunogen to prepare antiserum able to immunoprecipitate native VGF from both vaccinia virus-infected cell lysate and cell-free medium. Pulse-chase, tunicamycin treatment, and carbohydrate trimming experiments revealed that VGF is synthesized as a 19-kilodalton (kDa) precursor which is rapidly modified to a high-mannose-type 22-kDa protein. This cell-associated form is further processed into a 25-kDa polypeptide which, after proteolytic cleavage, releases the mature VGF into the medium as a 22-kDa glycoprotein.
Expression of the Hantaan virus M genome segment by using a vaccinia virus recombinant
Authors: Pensiero M N , Jennings G B , Schmaljohn C S , Hay J
Published: January 2020

A cDNA containing the complete open reading frame of the Hantaan virus (HTN) M genome segment has been cloned into vaccinia virus. This recombinant virus expresses two glycoproteins which are similar to the HTN structural glycoproteins, G1 and G2, in molecular weight, cleavage pattern, and cellular distribution. Both HTN and recombinant vaccinia virus glycoproteins are exclusively associated with the Golgi apparatus of the cell. Despite this intracellular restriction, mice inoculated with the recombinant vaccinia virus raised neutralizing antibodies against HTN. The specificity of virus neutralization appears to reside in the HTN glycoproteins, since a vaccinia virus recombinant expressing the HTN nucleocapsid protein was unable to elicit a neutralizing antibody response.
Latent herpes simplex virus type 1 transcripts in peripheral and central nervous system tissues of mice map to similar regions of the viral genome
Authors: Deatly A M , Spivack J G , Lavi E , O'Boyle D R , Fraser N W
Published: January 2020

Herpes simplex virus type 1 (HSV-1) DNA and RNA have been detected in peripheral nervous system (PNS) and central nervous system (CNS) tissues of latently infected mice. However, explant methods are successful in reactivating HSV-1 only from latently infected PNS tissues. In this report, latent herpesvirus infections in mouse PNS and CNS tissues were compared by in situ hybridization to determine whether the difference in reactivation was at the level of the virus or the host tissue. It was demonstrated that the HSV-1 transcripts present during latency in the mouse PNS and CNS originated from the same region of the genome, the repeats which bracket the long unique sequence. Therefore, the difference in reactivation with PNS and CNS tissues cannot be accounted for by differences in the extent of the HSV-1 genome transcribed during herpesvirus latency. Latent HSV-1 RNA was detected in the trigeminal ganglia (PNS) and the trigeminal system in the CNS from the mesencephalon to the spinal cord as well as other regions of the CNS not noted previously. Latent HSV-1 RNA was found predominantly in neurons but also in a small number of cells which could not be identified as neuronal cells. It is suggested that host differences in CNS and PNS tissues, rather than differences in latent virus transcription, may be important determinants in the HSV-1 reactivation process in explanted tissues.
Conservation of gene organization in the lymphotropic herpesviruses herpesvirus Saimiri and Epstein-Barr virus
Authors: Gompels U A , Craxton M A , Honess R W
Published: January 2020

By analyses of short DNA sequences, we have deduced the overall arrangement of genes in the (A + T)-rich coding sequences of herpesvirus saimiri (HVS) relative to the arrangements of homologous genes in the (G + C)-rich coding sequences of the Epstein-Barr virus (EBV) genome and the (A + T)-rich sequences of the varicella-zoster virus (VZV) genome. Fragments of HVS DNA from 13 separate sites within the 111 kilobase pairs of the light DNA coding sequences of the genome were subcloned into M13 vectors, and sequences of up to 350 bases were determined from each of these sites. Amino acid sequences predicted for fragments of open reading frames defined by these sequences were compared with a library of the protein sequences of major open reading frames predicted from the complete DNA sequences of VZV and EBV. Of the 13 short amino acid sequences obtained from HVS, only 3 were recognizably homologous to proteins encoded by VZV, but all 13 HVS sequences were unambiguously homologous to gene products encoded by EBV. The HVS reading frames identified by this method included homologs of the major capsid polypeptides, glycoprotein H, the major nonstructural DNA-binding protein, thymidine kinase, and the homolog of the regulatory gene product of the BMLF1 reading frame of EBV. Locally as well as globally, the order and relative orientation of these genes resembled that of their homologs on the EBV genome. Despite the major differences in their nucleotide compositions and in the nature and arrangements of reiterated DNA sequences, the genomes of the lymphotropic herpesviruses HVS and EBV encode closely related proteins, and they share a common organization of these coding sequences which differs from that of the neurotropic herpesviruses, VZV and herpes simplex virus.
Unusual DNA structure in the regulatory region of the human papovavirus JC virus
Authors: Amirhaeri S , Wohlrab F , Major E O , Wells R D
Published: January 2020

The human papovavirus JC virus (JCV) was analyzed for the presence of unusual DNA conformations. Recombinant plasmids containing 60% of the JCV prototype Mad-1 strain DNA were constructed and analyzed with both enzymatic and chemical probes. Fine-mapping studies revealed that the most prominent S1 nuclease-sensitive and bromoacetaldehyde-modified sites were located within the TATA boxes of each 98-base-pair tandem repeat. Further studies revealed that the S1 nuclease-sensitive site in the first TATA box (proximal to the origin) was approximately 50-fold stronger than the site in the second TATA box (distal from the origin). Deletion of the first TATA box drastically reduced the extent of bromoacetaldehyde modification in the second TATA box, whereas deletion of the second TATA box had little or no effect on the reactivity at the first TATA box. Hence, the biological and conformational role of the second TATA box remains unclear. No supercoil-induced relaxation was found, and reactions with the probes were not pH dependent. Also, fragments containing this regulatory region did not appear to be bent, although the A+T-rich segment contained a tract of eight consecutive A's. We conclude that the regulatory region of JCV contains non-B, but right-handed, DNA conformations which account for this behavior.
Differentiation and DNA contact points of host proteins binding at the cis site for virion-mediated induction of alpha genes of herpes simplex virus 1
Authors: Kristie T M , Roizman B
Published: January 2020

Transcriptional trans-activation of the five herpes simplex virus 1 alpha genes by the alpha trans-inducing factor requires a cis-acting site (alpha TIC; with the consensus 5'-GyATGnTAATGArATTCyTTGnGGG-3') located in the promoter-regulatory domains of the alpha genes. In DNA band shift assays with nuclear extracts from either mock-infected or infected cells, the DNA fragments containing an alpha TIC sequence from the alpha 0, alpha 4, and alpha 27 genes formed several cellular protein-DNA complexes designated alpha H1, alpha H2, and alpha H3. The host proteins that formed the alpha H2 and alpha H3 complexes were differentiated from those that formed the alpha H1 complex but not from each other by chromatography and specificity of the DNA-binding sites. The alpha H1 proteins protected the alpha TIC sequence of all three genes from DNase I digestion. Methylation of the purines in the sequence 5'-GyATGnTAAT-3' located at the 5' terminus of the alpha TIC sites precluded the binding of alpha H1. The binding site of the alpha H2-alpha H3 proteins in the alpha 27 gene alpha TIC overlapped, in part, with the alpha H1-binding site. The binding of these proteins was precluded by methylation of the purine residues in the sequence 5'-GCCACGTG-3' located at the 3' terminus of the DNase I footprint. The maximum apparent molecular weight of alpha H1 was 110,000, whereas that of alpha H2-alpha H3 was 64,000. A protein designated alpha H2', resembling alpha H2-alpha H3 with respect to molecular weight and chromatographic properties but differing in sequence specificity, bound to a site adjacent to the alpha H1 site in the fragment carrying an alpha TIC sequence of the alpha 4 gene. alpha H1 and alpha H2-alpha H3 or alpha H2' bound concurrently, notwithstanding the apparent overlap in the DNase I footprints.
Drosophila S virus, a hereditary reolike virus, probable agent of the morphological S character in Drosophila simulans
Authors: Louis C , Lopez-Ferber M , Comendador M , Plus N , Kuhl G , Baker S
Published: January 2020

Isometric reolike virions were found in all the examined Drosophila simulans flies from two strains (SimES-st and Israel-st) presenting the S phenotype, a maternally inherited morphological trait (abnormalities of bristles). Normal flies of both strains appeared virus-free. Virions were found in the cytoplasm of male and female gonads and epidermal cells, including the bristle-forming cells, which appeared disorganized. Steps of virogenesis were described. A positive correlation was demonstrated between expressivity of the S phenotype and degree of viral infection. This hereditary reolike virus seems to be responsible for the S character of D. simulans and was named DSV (Drosophila S virus).
Human T-cell leukemia virus types I and II exhibit different DNase I protection patterns
Authors: Altman R , Harrich D , Garcia J A , Gaynor R B
Published: January 2020

Human T-cell leukemia virus types I (HTLV-I) and II (HTLV-II) are human retroviruses which normally infect T-lymphoid cells. HTLV-I infection is associated with adult T-cell leukemia-lymphoma, and HTLV-II is associated with an indolent form of hairy-cell leukemia. To identify potential transcriptional regulatory elements of these two related human retroviruses, we performed DNase I footprinting of both the HTLV-I and HTLV-II long terminal repeats (LTRs) by using extracts prepared from uninfected T cells, HTLV-I and HTLV-II transformed T cells, and HeLa cells. Five regions of the HTLV-I LTR and three regions of the HTLV-II LTR showed protection by DNase I footprinting. All three of the 21-base-pair repeats previously shown to be important in HTLV transcriptional regulation were protected in the HTLV-I LTR, whereas only one of these repeats was protected in the HTLV-II LTR. Several regions exhibited altered protection in extracts prepared from lymphoid cells as compared with HeLa cells, but there were minimal differences in the protection patterns between HTLV-infected and uninfected lymphoid extracts. A number of HTLV-I and HTLV-II LTR fragments which contained regions showing protection in DNase I footprinting were able to function as inducible enhancer elements in transient CAT gene expression assays in the presence of the HTLV-II tat protein. The alterations in the pattern of the cellular proteins which bind to the HTLV-I and HTLV-II LTRs may in part be responsible for differences in the transcriptional regulation of these two related viruses.
Location, transcript analysis, and partial nucleotide sequence of the cytomegalovirus gene encoding an early DNA-binding protein with similarities to ICP8 of herpes simplex virus type 1
Authors: Anders D G , Gibson W
Published: January 2020

The results presented here locate the gene encoding an early, nonvirion, single-stranded DNA-binding protein of human and simian strains of cytomegalovirus (CMV) [HCMV(Towne) DB140 and SCMV(Colburn) DB129, respectively] and provide additional evidence that this protein is the CMV homolog of the herpes simplex virus type 1 (HSV-1) major DNA-binding protein (ICP8), as proposed earlier (D. G. Anders, A. Irmiere, and W. Gibson, J. Virol. 58:253-262). The ICP8 gene was used as a probe in Southern analyses done at moderate stringency as an approach to locating similar sequences in the CMV genome. The BamHI K and EcoRI V fragments from the center of the long unique segment of HCMV(Towne) hybridized with the ICP8 probe and were in turn used to identify corresponding sequences in the EcoRI D fragment of SCMV(Colburn). RNA prepared from SCMV(Colburn)-infected cells directed the in vitro synthesis of DB129. If the RNA was first hybridized with the cloned 12.5-kilobase EcoRI D fragment, in vitro synthesis of DB129 was specifically inhibited. Additional hybrid-arrested in vitro translation experiments with subclones spanning the EcoRI D fragment demonstrated that the DB129 gene is located in the left half of that fragment, approximately bisected by a SalI site. RNA analyses identified 3.9-, 8.9-, and 10.0-kilobase RNA species expressed from this region. A partial nucleotide sequence of the Colburn region mapping within the boundaries of the 3.9-kilobase transcript, suspected to be the primary coding species, showed significant sequence similarity to the major DNA-binding protein gene homolog identified in B95-8 Epstein-Barr virus.
Localization of a herpes simplex virus neurovirulence gene dissociated from high-titer virus replication in the brain
Authors: Javier R T , Izumi K M , Stevens J G
Published: January 2020

Previous studies with the herpes simplex virus type 1 X type 2 intertypic recombinant RS6 suggested that the genomic region from 0.11 to 0.14 map units is involved in neurovirulence (R. T. Javier, R. L. Thompson, and J. G. Stevens, J. Virol. 61:1978-1984, 1987). To study this further, we isolated an RS6-derived herpes simplex virus intertypic recombinant (R13-1) which has a genetic defect within this area. After inoculation into mouse brains, R13-1 was found to be approximately 10,000-fold less neurovirulent than either the wild-type type 1 or type 2 parental virus. However, R13-1 replicated in the mouse brain to titers resembling those of the wild-type parents. Further comparisons with wild-type counterparts indicated that R13-1 expressed equivalent levels of the enzyme thymidine kinase and replicated to intermediate levels in primary mouse embryo fibroblasts maintained at the normal body temperature for mice. Using marker rescue techniques combined with in vivo selection, we found that recombination between unit-length R13-1 DNA and a cloned type 1 DNA fragment spanning the region from 0.11 to 0.14 map units (EcoRI-d, 0.079 to 0.192 map units) generated viruses with a wild-type neurovirulence phenotype. To further refine the genomic region of interest, we performed marker rescue experiments using two EcoRI-d subclones, EcoRI/BamHI dc (0.079 to 0.143 map units) and BamHI/EcoRI and (0.143 to 0.192 map units), representing the left and right halves of the EcoRI d fragment, respectively. In these experiments the EcoRI/BamHI dc clone, but not the BamHI/EcoRI ad clone, yielded recombinant viruses exhibiting wild-type neurovirulence. These results show that at least one herpes simplex virus gene function associated with neurovirulence is located within a 9.1-kilobase region at 0.079 to 0.143 map units of the viral genome. Perhaps more significantly, the results indicate that this neurovirulence property functions independently of high-titer virus replication in the brain.
Analysis of adenovirus type 2 L1 RNA 3'-end formation in vivo and in vitro
Authors: Hales K H , Birk J M , Imperiale M J
Published: January 2020

Downstream sequence requirements for efficient cleavage and polyadenylation at the adenovirus type 2 L1 poly(A) site were determined in vivo in 293 cells and in vitro by using RNA precursors in HeLa cell nuclear extracts. The two cleavage sites used were found to differ in sensitivity to 3'-end deletion in vivo and in vitro.
Study of the E2 gene product of the cottontail rabbit papillomavirus reveals a common mechanism of transactivation among papillomaviruses
Authors: Giri I , Yaniv M
Published: January 2020

The long control region (LCR) of the cottontail rabbit papillomavirus (CRPV) harbors a transcriptional promoter which can be transactivated, as reflected by cat gene expression, by cotransfection with plasmids which express the intact E2 open reading frame of CRPV, human papillomavirus type 18 (HPV18), and bovine papillomavirus type 1 (BPV1). The E2 protein of CRPV can also transactivate the LCRs of BPV1, HPV1, and HPV18 inserted in front of the cat gene in enhancer or promoter configuration. Competition experiments in vivo and binding studies with CRPV E2 protein synthesized in vitro suggest that the different E2 proteins transactivate transcription by a common mechanism involving binding to the same ACCG-CGGT target sequence. The C-terminal part of the protein is necessary for its DNA-binding function. Analysis of the transactivation data and of the LCR sequences of these four viruses suggests that the two cutaneous viruses (CRPV and BPV1) present a similar pattern of promoter regulation but that the activity of the promoters of genital human viruses is less dependent on E2 regulation and is at least partially regulated by cellular factors.
DNA rearrangements in organ-specific variants of polyomavirus JC strain GS
Authors: Loeber G , Dörries K
Published: January 2020

Variants of JC virus (JCV) strain GS were isolated directly from the central nervous system (variant GS/B) and the kidney (variant GS/K) of a patient with progressive multifocal leukoencephalopathy and were cloned and sequenced. The genomes of the isolates were shown to be nearly identical in the nucleotide sequences of their protein-coding regions, suggesting that both had originated from a single infecting JCV genome. In contrast, the arrangement of the putative elements of transcriptional control revealed considerable differences. The tandemly repeated elements found twice within the enhancer region of JCV GS/B variant were not present in the GS/K variant. The missing elements were replaced by DNA segments containing simian virus 40 and adenovirus E1A core enhancer elements. These differences in the organ-specific GS variants suggest that rearrangements within elements of transcriptional control might be involved in altering the virus-cell interaction in the course of a JCV infection.
Identification of the gene coding for the precursor of adenovirus core protein X
Authors: Weber J M , Anderson C W
Published: January 2020

Nucleoprotein complexes extracted from infected cells (J. Weber and L. Philipson, Virology 136:321-327, 1984) or ts1 virions grown at 39 degrees C (J. M. Weber, G. Khitto, and A. R. Bhatti, Can. J. Microbiol. 29:235-241, 1983) contain, in addition to known virion proteins, a polypeptide with a molecular weight of 11,000 (11K polypeptide). We identified the gene for this polypeptide by sequence analysis of the radioactively labeled 11K polypeptide isolated from ts1 virions and comparison of this sequence with the nucleotide sequence of the adenovirus type 2 genome. The 11K polypeptide was encoded by an open reading frame of 80 residues located between nucleotides 17,676 and 17,915 in late transcription region 2 of adenovirus type 2; the initiating methionine residue was removed leaving a 79-residue product. The late transcription region 2 that encoded the 79-residue polypeptide (11K) was arginine rich (21%) and had a predicted molecular weight of 8,715. It was cleaved by the viral endoprotease to give two products which comigrated on sodium dodecyl sulfate-polyacrylamide gels as virion polypeptide X. Our data suggest that additional cleavage of the carboxy-terminal 48-residue fragment generates a 19-amino-acid fragment with the amino acid composition of mu (K. Hosokawa and M. T. Sung, J. Virol. 17:924-934, 1976).
Three-dimensional structure of poliovirus serotype 1 neutralizing determinants
Authors: Page G S , Mosser A G , Hogle J M , Filman D J , Rueckert R R , Chow M
Published: January 2020

Antigenic mutants of poliovirus (Sabin strain, serotype 1) were isolated by the resistance of the virus to anti-Sabin neutralizing monoclonal antibodies. The amino acid replacements within the capsid protein sequence causing the altered antigenicity were identified for each of 63 isolates. The mutations cluster into distinct nonoverlapping peptide segments that group into three general immunological phenotypes on the basis of cross-neutralization analyses with 15 neutralizing anti-Sabin monoclonal antibodies. Location of the mutated amino acid residues within the three-dimensional structure of the virion indicates that the majority of these amino acid residues are highly exposed and located within prominent structural features of the viral surface. Those mutated amino acid residues that are less accessible to antibody interaction are often involved in hydrogen bonds or salt bridges that would stabilize the local tertiary structure of the antigenic site. The interactions of the peptide segments that form these neutralizing sites suggest specific models for the generation of neutralization-resistant variants and for the interaction between the viral surface and antibody.
Prediction of human rotavirus serotype by nucleotide sequence analysis of the VP7 protein gene
Authors: Green K Y , Sears J F , Taniguchi K , Midthun K , Hoshino Y , Gorziglia M , Nishikawa K , Urasawa S , Kapikian A Z , Chanock R M
Published: January 2020

Human rotavirus field isolates were characterized by direct sequence analysis of the gene encoding the serotype-specific major neutralization protein (VP7). Single-stranded RNA transcripts were prepared from virus particles obtained directly from stool specimens or after two or three passages in MA-104 cells. Two regions of the gene (nucleotides 307 through 351 and 670 through 711) which had previously been shown to contain regions of sequence divergence among rotavirus serotypes were sequenced by the dideoxynucleotide method with two different synthetic oligonucleotide primers. The resulting nucleotide sequences were compared with the corresponding sequences from rotaviruses of known serotype (serotype 1, 2, 3, or 4). A total of 25 field isolates and 10 laboratory strains examined by this method exhibited marked sequence identity in both areas of the gene with the corresponding regions of 1 of the 4 reference strains. In addition, the predicted serotype from the sequence analysis correlated in each case with the serotype determined when the rotaviruses were examined by plaque reduction neutralization or reactivity with serotype-specific monoclonal antibodies. These data suggest that as a result of the high degree of sequence conservation observed among rotaviruses of the same serotype, it is possible to predict the serotype of a rotavirus isolate by direct sequence analysis of its VP7 gene.
Sequence and transcriptional analysis of the vaccinia virus HindIII I fragment
Authors: Schmitt J F , Stunnenberg H G
Published: January 2020

The complete sequence of the vaccinia virus HindIII I fragment, which is composed of 6,498 base pairs, encodes six complete and two incomplete open reading frames (ORFs). Computer analysis revealed an amino acid sequence homology between ORF I 4 and the large subunit of the ribonucleotide reductase complex. The two small polypeptides derived from ORFs I 2 and I 5, with molecular weights of 8,500 and 8,700, respectively, have a very high hydrophobic amino acid sequence composition. S1 analysis revealed that ORF I 4 is expressed at early stages of infection, ORFs I 1, I 2, I 5, and I 7 are expressed in the late phase of infection, and ORF I 3 is constitutively expressed. Screening a vaccinia virus genomic library revealed a large vaccinia virus insert overlapping the HindIII I and O fragments which contains a previously undetected HindIII P fragment of approximately 300 base pairs. S1 analysis revealed an early (O1) and a late (O2) start site of transcription initiation located within the HindIII O fragment.
Adeno-associated virus general transduction vectors: analysis of proviral structures
Authors: McLaughlin S K , Collis P , Hermonat P L , Muzyczka N
Published: January 2020

We used two kinds of adeno-associated virus (AAV) vectors to transduce the neomycin resistance gene into human cells. The first of these (dl52-91) retains the AAV rep genes; the second (dl3-94) retains only the AAV terminal repeats and the AAV polyadenylation signal (428 base pairs). Both vectors could be packaged into AAV virions and produced proviral structures that were essentially the same. Thus, the AAV sequences that are required in cis for packaging (pac), integration (int), rescue (res), and replication (ori) of viral DNA are located within a 284-base-pair sequence that includes the terminal repeat. Most of the G418r cell lines (73%) contained proviruses which could be rescued (Res+) when the cells were superinfected with the appropriate helper viruses. Some produced high yields of viral DNA; other rescued at a 50-fold lower level. Most of the lines that were Res+ (79%) contained a tandem repeat of the AAV genome (2 to 20 copies) which was integrated randomly with respect to cellular DNA. Junctions between two consecutive AAV copies in a tandem array contained either one or two copies of the AAV terminal palindrome. Junctions between AAV and cellular sequences occurred predominantly at or within the AAV terminal repeat, but in some cases at internal AAV sequences. Two lines were seen that contained free episomal copies of AAV DNA. Res+ clones contained deleted proviruses or tandem repeats of a deleted genome. Occasionally, flanking cellular DNA was also amplified. There was no superinfection inhibition of AAV DNA integration. Our results suggest that AAV sequences are amplified by DNA replication either before or after integration and that the mechanism of replication is different from the one used during AAV lytic infections. In addition, we have described a new AAV general transduction vector, dl3-94, which provides the maximum amount of room for insertion of foreign DNA and integrates at a high frequency (80%).
Structural and functional domains of the myb oncogene: requirements for nuclear transport, myeloid transformation, and colony formation
Authors: Ibanez C E , Lipsick J S
Published: January 2020

The v-myb oncogene of avian myeloblastosis virus causes acute myelomonocytic leukemia in vivo and transforms only myeloid cells in vitro. Its product, p48v-myb, is a nuclear protein of unknown function. To determine structure-function relationships for this protein, we constructed a series of deletion mutants of v-myb, expressed them in retroviral vectors, and studied their biochemical and biological properties. We used these mutants to identify two separate domains of p48v-myb which had distinct roles in its accumulation in the cell nucleus. We showed that the viral sequences which normally encode both termini of p48v-myb were dispensible for transformation. In contrast, both copies of the highly conserved v-myb amino-terminal repeat were required for transformation. We also identified a carboxyl-terminal domain of p48v-myb which was required for the growth of v-myb-transformed myeloblasts in soft agar but not for morphological transformation.
Rapid selection of genetic and antigenic variants of foot-and-mouth disease virus during persistence in cattle
Authors: Gebauer F , de la Torre J C , Gomes I , Mateu M G , Barahona H , Tiraboschi B , Bergmann I , de Mello P A , Domingo E
Published: January 2020

Rapid evolution of foot-and-mouth disease virus (FMDV) is documented during persistent infections of cattle. The carrier state was established experimentally with plaque-purified FMDV of serotype C3. Virus was recovered from the esophageal pharyngeal area of the animals up to 539 days postinfection. Analysis of capsid proteins by electrofocusing and by electrophoretic mobility of the genomic poly(C)-rich tract suggested heterogeneity in several isolates and sequential dominance of viral subpopulations. Nucleotide sequences of the VP1-coding region of the parental FMDV C3 clones and of seven isolates from the carrier cattle showed point mutations that represented rates of fixation of mutations of 0.9 X 10(-2) to 7.4 X 10(-2) substitutions per nucleotide per year; 59% of the base changes led to amino acid substitutions, some of which were located within residues 135 to 151, a region involved in neutralization of FMDV. In the esophageal pharyngeal fluid samples, FMDV C3-neutralizing activity was present. Antigenic variation was demonstrated with monoclonal antibodies raised against FMDV C3. Two isolates from carrier cattle differed from the parental virus by 10(2)- or 10(3)-fold decreased reactivity with neutralizing monoclonal antibodies. We suggest that persistent, inapparent infections of ruminants, in addition to being a reservoir of virus, may promote the rapid selection of antigenically variant FMDVs.
The adenovirus tripartite leader may eliminate the requirement for cap-binding protein complex during translation initiation
Authors: Dolph P J , Racaniello V , Villamarin A , Palladino F , Schneider R J
Published: January 2020

The adenovirus tripartite leader is a 200-nucleotide 5' noncoding region that is found on all late viral mRNAs. This segment is required for preferential translation of viral mRNAs at late times during infection. Most tripartite leader-containing mRNAs appear to exhibit little if any requirement for intact cap-binding protein complex, a property previously established only for uncapped poliovirus mRNAs and capped mRNAs with minimal secondary structure. The tripartite leader also permits the translation of mRNAs in poliovirus-infected cells in the apparent absence of active cap-binding protein complex and does not require any adenovirus gene products for this activity. The preferential translation of viral late mRNAs may involve this unusual property.
Inoculation of baboons and macaques with simian immunodeficiency virus/Mne, a primate lentivirus closely related to human immunodeficiency virus type 2
Authors: Benveniste R E , Morton W R , Clark E A , Tsai C C , Ochs H D , Ward J M , Kuller L , Knott W B , Hill R W , Gale M J
Published: January 2020

A primate lymphotropic lentivirus was isolated on the human T-cell line HuT 78 after cocultivation of a lymph node from a pig-tailed macaque (Macaca nemestrina) that had died with malignant lymphoma. This isolate, originally designated M. nemestrina immunodeficiency virus (MnIV) and now classified as simian immunodeficiency virus (SIV/Mne), was inoculated intravenously into three juvenile rhesus monkeys (Macaca mulatta), three juvenile pig-tailed macaques (M. nemestrina), and two juvenile baboons (Papio cynocephalus). All six macaques became viremic by 3 weeks after inoculation, whereas neither of the baboons developed viremia. One pig-tailed macaque died at 15 weeks with suppurative peritonitis secondary to ulcerative, necrotizing colitis. Immunologic abnormalities included a marked decrease in CD4+ peripheral blood lymphocytes. Although five macaques mounted an antibody response to SIV/Mne, the animal that died at 15 weeks remained antibody negative. Three other macaques (two rhesus and one pig-tailed) died 66 to 87 weeks after inoculation after exhibiting progressive weight loss, anemia, and diarrhea. Histopathologic findings at necropsy included various manifestations of immune deficiency, nephropathy, subacute encephalitis, pancreatitis, adenocarcinoma, and lymphoid atrophy. SIV/Mne could be readily isolated from the spleens and lymph nodes of all necropsied macaques, and from the cerebrospinal fluid, brains, bone marrow, livers, and pancreas of some of the animals. SIV antigens were localized by avidin-biotin immunohistochemistry to pancreatic islet cells and to bone marrow endothelial cells. The data suggest that African baboons may be resistant to infection by SIV/Mne, whereas Asian macaques are susceptible to infection with this pathogenic primate lentivirus.
Reduced yield of infectious pseudorabies virus and herpes simplex virus from cell lines producing viral glycoprotein gp50
Authors: Petrovskis E A , Meyer A L , Post L E
Published: January 2020

Pseudorabies virus (PRV) glycoprotein gp50 is the homolog of herpes simplex virus (HSV) glycoprotein D. Several cell lines that constitutively synthesize gp50 were constructed. Vero cells, HeLa cells, and pig kidney (MVPK) cells that produce gp50 all gave reduced yields of PRV and HSV progeny viruses when compared with the parent cell line or the same cell line transfected to produce a different protein. The reduction in virus yield was greatest at low multiplicities of infection. The Vero and HeLa cells that produce gp50 showed an even greater reduction in HSV yield than in PRV yield. This phenomenon may be an example in a herpesvirus of the interference observed in retroviruses or cross-protection in plant virus systems.
Passive immunity to bovine rotavirus infection associated with transfer of serum antibody into the intestinal lumen
Authors: Besser T E , Gay C C , McGuire T C , Evermann J F
Published: January 2020

The effect of circulating passive antibody on immunity to bovine rotavirus infections in neonatal calves was investigated. In the first experiment, rotavirus antibody titers in the small intestinal lumina of 5- and 10-day-old calves with a wide range of serum rotavirus antibody titers were determined. Neutralizing antibody was present in the small intestinal lumina in titers that correlated with the calves' serum titers (r = +0.84, P less than 0.01). Immunoglobulin G1 was the predominant isotype of intestinal luminal rotavirus antibody. Calves not fed colostrum during the absorptive period lacked rotavirus antibody in circulation and in the intestinal lumen at 7 days of age, even when they were fed large volumes of colostrum with a high rotavirus antibody titer at 48 h after birth. Therefore, rotavirus antibody is not retained in the intestinal lumen for 5 days following a colostrum meal, and the luminal antibody in the 5- and 10-day-old seropositive calves were probably derived from circulating antibody. In a second experiment, calves were passively immunized by subcutaneous injection of colostral whey with a high immunoglobulin G1 rotavirus antibody titer and challenged with virulent bovine rotavirus 48 h later. The passively immunized calves were protected from rotavirus infection and diarrhea compared with calves with comparable serum immunoglobulin concentrations but with lower serum rotavirus with lower serum rotavirus antibody titers. The results of these experiments indicate that circulating immunoglobulin G1 antibody appears in the gastrointestinal tract of neonatal calves and that circulating rotavirus antibody can prevent infection and diarrhea after rotavirus challenge.
Influenza virus RNA replication in vitro: synthesis of viral template RNAs and virion RNAs in the absence of an added primer
Authors: Shapiro G I , Krug R M
Published: January 2020

The two steps in influenza virus RNA replication are (i) the synthesis of template RNAs, i.e., full-length copies of the virion RNAs, and (ii) the copying of these template RNAs into new virion RNAs. We prepared nuclear extracts from infected HeLa cells that catalyzed both template RNA and virion RNA synthesis in vitro in the absence of an added primer. Antibody depletion experiments implicated nucleocapsid protein molecules not associated with nucleocapsids in template RNA synthesis for antitermination at the polyadenylation site used during viral mRNA synthesis. Experiments with the WSN influenza virus temperature-sensitive mutant ts56 containing a defect in the nucleocapsid protein proved that the nucleocapsid protein was indeed required for template RNA synthesis both in vivo and in vitro. Nuclear extracts prepared from mutant virus-infected cells synthesized template RNA at the permissive temperature but not at the nonpermissive temperature, whereas the synthesis of mRNA-size transcripts was not decreased at the nonpermissive temperature. Antibody depletion experiments showed that nucleocapsid protein molecules not associated with nucleocapsids were also required for the copying of template RNA into virion RNA. In contrast to the situation with the synthesis of transcripts complementary to virion RNA, no discrete termination product(s) were made during virion RNA synthesis in vitro in the absence of nucleocapsid protein molecules.
Mechanism of c-erbB transduction: newly released transducing viruses retain poly(A) tracts of erbB transcripts and encode C-terminally intact erbB proteins
Authors: Raines M A , Maihle N J , Moscovici C , Crittenden L , Kung H J
Published: January 2020

We have previously shown that avian leukosis virus (ALV) induces erythroblastosis by insertional activation of the c-erbB gene. In 25% of the ALV-induced leukemic samples we have analyzed, acute retroviruses that have captured the activated erbB oncogene were released. The unusually high frequency at which erbB transduction occurs makes this an ideal system for studying the mechanism of oncogene transduction. In addition, these leukemic samples provide a rich source for the isolation of novel erbB-transducing viruses. We report here our characterization of several new erbB-transducing proviruses. The 5' recombination points of all these viruses mapped to the same intron in which proviral insertions cluster, supporting the hypothesis that transduction begins with proviral insertion near the oncogene. The 3' recombination points usually occurred within the 3' untranslated region downstream from the termination codon of the c-erbB gene. Three of the erbB-containing proviruses were molecularly cloned and analyzed in detail. Two of them were capable of releasing acute viruses, and interestingly, both retained poly(A) tracts of erbB messages in their genomes. A stretch of six adenosine residues in the ALV env gene appeared to mediate the 3' recombination events required for the generation of these viruses. These data provide further insight into the mechanism by which oncogenes are transduced into retroviral genomes.
Cytoplasmic assembly and accumulation of human immunodeficiency virus types 1 and 2 in recombinant human colony-stimulating factor-1-treated human monocytes: an ultrastructural study
Authors: Orenstein J M , Meltzer M S , Phipps T , Gendelman H E
Published: January 2020

Recombinant human colony-stimulating factor-1-treated human peripheral blood-derived monocytes-macrophages are efficient host cells for recovery of the human immunodeficiency virus (HIV) from blood leukocytes of patients with acquired immunodeficiency syndrome. These cells can be maintained as viable monolayers for intervals exceeding 3 months. Infection with HIV resulted in virus-induced cytopathic effects, accompanied by relatively high levels of released progeny virus, followed by a prolonged low-level release of virus from morphologically normal cells. In both acutely and chronically infected monocytes, viral particles were seen budding into and accumulating within cytoplasmic vacuoles. The number of intravacuolar virions far exceeded those associated with the plasma membrane, especially in the chronic phase, and were concentrated in the perinuclear Golgi zone. In many instances, the vacuoles were identified as Golgi elements. Fusion of virus-laden vacuoles with primary lysosomes were rare. The pattern of cytoplasmic assembly of virus was observed with both HIV types 1 and 2 and in brain macrophages of an individual with acquired immunodeficiency syndrome encephalopathy. Immunoglobulin-coated gold beads added to acutely infected cultures were segregated from the vacuoles containing virus; relatively few beads and viral particles colocalized. The assembly of HIV virions within vacuoles of macrophages is in contrast to the exclusive surface assembly of HIV by T lymphocytes. Intracytoplasmic virus hidden from immune surveillance in monocytes-macrophages may explain, in part, the persistence of HIV in the infected human host.