bluuebunny/crossref_metadata_2025_split huggingface dataset:
Transgenic Plants Expressing Potato Virus X ORF2 Protein (p24) Are Resistant to Tobacco Mosaic Virus and Ob Tobamoviruses
Authors: Ares X. , Calamante G. , Cabral S. , Lodge J. , Hemenway P. , Beachy R. N. , Mentaberry A. 
Published: December 2019

ABSTRACT

The p24 protein, one of the three proteins implicated in local movement of potato virus X (PVX), was expressed in transgenic tobacco plants (
Nicotiana tabacum
Xanthi D8 NN). Plants with the highest level of p24 accumulation exhibited a stunted and slightly chlorotic phenotype. These transgenic plants facilitate the cell-to-cell movement of a mutant of PVX that contained a frameshift mutation in p24. Upon inoculation with tobacco mosaic virus (TMV), the size of necrotic local lesions was significantly smaller in p24+ plants than in nontransgenic, control plants. Systemic resistance to tobamoviruses was also evidenced after inoculation of p24+ plants with Ob, a virus that evades the hypersensitive response provided by the N gene. In the latter case, no systemic symptoms were observed, and virus accumulation remained low or undetectable by Western immunoblot analysis and back-inoculation assays. In contrast, no differences were observed in virus accumulation after inoculation with PVX, although more severe symptoms were evident on p24-expressing plants than on control plants. Similarly, infection assays conducted with potato virus Y showed no differences between control and transgenic plants. On the other hand, a considerable delay in virus accumulation and symptom development was observed when transgenic tobacco plants containing the movement protein (MP) of TMV were inoculated with PVX. Finally, a movement defective mutant of TMV was inoculated on p24+ plants or in mixed infections with PVX on nontransgenic plants. Both types of assays failed to produce TMV infections, implying that TMV MP is not interchangeable with the PVX MPs.
Viral E6-E7 Transcription in the Basal Layer of Organotypic Cultures without Apparent p21cip1 Protein Precedes Immortalization of Human Papillomavirus Type 16- and 18-Transfected Human Keratinocytes
Authors: Steenbergen Renske D. M. , Parker Jacqueline N. , Isern Sharon , Snijders Peter J. F. , Walboomers Jan M. M. , Meijer Chris J. L. M. , Broker Thomas R. , Chow Louise T. 
Published: December 2019

ABSTRACT
Organotypic cultures of human keratinocytes provide a useful model system to study human papillomavirus (HPV)-host cell interactions. In this study, we analyzed organotypic cultures of two HPV type 16 (HPV16) (FK16A and FK16B)- and two HPV18 (FK18A and FK18B)-transfected keratinocyte cell lines through the process of immortalization in vitro. For FK16A and FK18B cells, passages of both mortal cells in their extended life span and subsequent immortal stages were studied. Mortal cells of FK16A and FK18B showed a morphology reminiscent of mild to moderate dysplasia, whereas in their immortal descendants, severely dysplastic features were observed. Immortal FK18A cells were mildly to moderately dysplastic, while FK16B cells were severely dysplastic. The increasing degrees of dysplasia were associated with a decreasing expression of differentiation markers cytokeratin 10 and profilaggrin. All raft cultures expressed E6-E7 mRNAs in the basal layer, while the amount of viral transcripts in the suprabasal cells was in general proportional to the degree of dysplasia. In all cases, E6-E7 transcription and dysplastic features were highly correlated with cellular proliferation, as assessed by Ki-67 (MIB-1) antigen expression. Moreover, high levels of E6-E7 transcription and expression of p21cip1 protein in the basal layer seemed to be mutually exclusive. We conclude that expression of E6-E7 in the basal cells associated with increased proliferation in the absence of detectable p21cip1 protein is apparently necessary but not sufficient for immortalization, or for the loss of terminal differentiation, for which yet to be discovered additional events are required. The model system described in this study provides a valuable tool to analyze alterations in viral transcription regulation during HPV-mediated cell transformation.
An Arenavirus RING (Zinc-Binding) Protein Binds the Oncoprotein Promyelocyte Leukemia Protein (PML) and Relocates PML Nuclear Bodies to the Cytoplasm
Authors: Borden Katherine L. B. , Campbell Dwyer Elizabeth J. , Salvato Maria S. 
Published: December 2019

ABSTRACTThe promyelocytic leukemia protein (PML) forms nuclear bodies which are altered in some disease conditions. We report that the cytoplasmic RNA virus lymphocytic choriomeningitis virus (LCMV) influences the distribution of PML bodies. In cells infected with LCMV, the Z protein and PML form large bodies primarily in the cytoplasm. Transient transfection studies indicate that Z alone is sufficient to redistribute PML to the cytoplasm and that PML and Z colocalize. Coimmunoprecipitation studies show specific interaction between PML and Z proteins. A similar result was observed with a Z protein from another arenavirus, Lassa virus, suggesting that this is a general feature of theArenaviridae. Genetically engineered mutations in PML were used to show that the Z protein binds the N-terminal region of PML and does not need the PML RING or the nuclear localization signal to colocalize. The Z protein acts dominantly to overcome the diffuse phenotype observed in several PML mutants. The interaction between PML and Z may influence certain unique characteristics of arenavirus infection.
Mechanism of Borna Disease Virus Entry into Cells
Authors: Gonzalez-Dunia Daniel , Cubitt Beatrice , de la Torre Juan Carlos 
Published: December 2019

ABSTRACT
We have investigated the entry pathway of Borna disease virus (BDV). Virus entry was assessed by detecting early viral replication and transcription. Lysosomotropic agents (ammonium chloride, chloroquine, and amantadine), as well as energy depletion, prevented BDV infection, indicating that BDV enters host cells by endocytosis and requires an acidic intracellular compartment to allow membrane fusion and initiate infection. Consistent with this hypothesis, we observed that BDV-infected cells form extensive syncytia upon low-pH treatment. Entry of enveloped viruses into animal cells usually requires the membrane-fusing activity of viral surface glycoproteins (GPs). BDV GP is expressed as two products of 84 and 43 kDa (GP-84 and GP-43, respectively). We show here that only GP-43 is present at the surface of BDV-infected cells and therefore is likely the viral polypeptide responsible for triggering fusion events. We also present evidence that GP-43, which corresponds to the C terminus of GP-84, is generated by cleavage of GP-84 by the cellular protease furin. Hence, we propose that BDV GP-84 is involved in attachment to the cell surface receptor whereas its furin-cleaved product, GP-43, is involved in pH-dependent fusion after internalization of the virion by endocytosis.
Characterization of Provirus Clones of Simian Foamy Virus Type 1
Authors: Mergia Ayalew , Wu Min 
Published: December 2019

ABSTRACT

We have cloned proviral DNA of simian foamy virus type 1 (SFV-1) from linear unintegrated DNA (pSFV-1). Transfection of pSFV-1 induces cytopathology in several cell lines with supernatants from the transfected cell culture containing infectious viral particles. Electron microscopy of the transfected cells revealed foamy virus particles. Deletion analysis of pSFV-1 indicated that the transcriptional transactivator (
tas
) gene located between
env
and the long terminal repeat is critical for virus replication, whereas the second open reading frame (ORF-2) in this region is dispensable. Although the
tas
and ORF-2 regions of foamy viruses have significantly diverged, the results presented here suggested that the gene products have similar functions. Recombinant pSFV-1 containing the
cat
gene was able to transduce the heterologous gene, indicating the utility of SFV-1 as a vector. An infectious clone of SFV-1 which is distantly related to the human foamy virus will provide a means to understand the biology of this unique group of viruses.
CCR5 Expression Correlates with Susceptibility of Maturing Monocytes to Human Immunodeficiency Virus Type 1 Infection
Authors: Naif Hassan M. , Li Shan , Alali Mohammed , Sloane Andrew , Wu Lijun , Kelly Mark , Lynch Garry , Lloyd Andrew , Cunningham Anthony L. 
Published: December 2019

ABSTRACTThe chemokine receptor CCR5 and to a lesser extent CCR3 and CCR2b have been shown to serve as coreceptors for human immunodeficiency virus type 1 (HIV-1) entry into blood- or tissue-derived macrophages. Therefore, we examined the expression of the chemokine receptors CCR1, CCR2b, CCR3, CCR5, and CXCR4 as RNAs or as membrane-expressed antigens in monocytes maturing into macrophages and correlated these results with the susceptibility of macrophages to HIV-1 infection, as measured by their concentrations of extracellular p24 antigen and levels of intracellular HIV DNA by quantitative PCR. There was little change in levels of CCR1, CCR2b, and CCR5 RNAs. CCR3 RNA and surface antigen were undetectable throughout maturation of adherent monocytes over 10 days. CXCR4 RNA and membrane antigen were strongly expressed in newly adherent monocytes, but their levels declined at day 7. The amounts of CCR5 RNA remained stable, but the amounts of CCR5 antigen increased from undetectable to peak levels at day 7 and then declined slightly at day 10. Levels of susceptibility to laboratory (HIV-1BaL) and clinical strains of HIV-1 showed parallel kinetics, peaking at day 7 and then decreasing at days 10 to 14. The concordance of levels of HIV DNA and p24 antigen suggested that the changes in susceptibility with monocyte maturation were at or immediately after entry and correlated well with CCR5 expression and inversely with CXCR4 expression.
Apoptotic Regulation of T Cells and Absence of Immune Deficiency in Virus-Infected Gamma Interferon Receptor Knockout Mice
Authors: Lohman Barbara L. , Welsh Raymond M. 
Published: December 2019

ABSTRACTAcute viral infections often induce a transient period of immune deficiency in which the host’s T cells fail to proliferate in response to T-cell mitogens and fail to make an antigen-specific memory recall response. This has been associated with the enhanced sensitivity of these highly activated T cells to undergo apoptosis, or activation-induced cell death (AICD), upon T-cell receptor ligation. Here we show that gamma interferon receptor-deficient (IFN-γ R−/−) mice mount a T-cell response to lymphocytic choriomeningitis virus (LCMV) infection but fail to undergo the transient immune deficiency. Instead, their T cells were hyperproliferative and relatively, but not completely, resistant to AICD. The immune response returned to homeostasis, but with delayed kinetics, in parallel with delayed clearance of the virus. Wild-type mice receiving high doses of disseminating LCMV Clone 13 are known to undergo clonal exhaustion of their virus-specific cytotoxic T lymphocytes (CTL). To determine whether this process was mediated by AICD associated with IFN-γ or with Fas-Fas ligand interactions, LCMV-specific precursor CTL frequencies were examined in LCMV Clone 13-infected IFN-γ R−/−orlpr(Fas-deficient) mice. In both instances, viral persistence was established and CTL precursors were greatly eliminated. This finding indicates that clonal exhaustion of CTL does not require IFN-γ or Fas, even though both molecules influence AICD and the transient immune deficiency seen in the LCMV infection.
An Envelope Modification That Renders a Primary, Neutralization-Resistant Clade B Human Immunodeficiency Virus Type 1 Isolate Highly Susceptible to Neutralization by Sera from Other Clades
Authors: Stamatatos Leonidas , Cheng-Mayer Cecilia 
Published: December 2019

ABSTRACTSF162 is a primary (PR), non-syncytium-inducing, macrophagetropic human immunodeficiency virus type 1 (HIV-1) clade B isolate which is resistant to antibody-mediated neutralization. Deletion of the first or second hypervariable envelope gp120 region (V1 or V2 loop, respectively) of this virus does not abrogate its ability to replicate in peripheral blood mononuclear cells and primary macrophages, nor does it alter its coreceptor usage profile. The mutant virus with the V1 loop deletion, SF162ΔV1, remains as resistant to antibody-mediated neutralization as the wild-type virus SF162. In contrast, the mutant virus with the V2 loop deletion, SF162ΔV2, exhibits enhanced susceptibility to neutralization by certain monoclonal antibodies whose epitopes are located within the CD4-binding site and conserved regions of gp120. More importantly, SF162ΔV2 is now up to 170-fold more susceptible to neutralization than SF162 by sera collected from patients infected with clade B HIV-1 isolates. In addition, it becomes susceptible to neutralization by sera collected from patients infected with clade A, C, D, E, and F HIV-1 isolates. These findings suggest that the V2, but not the V1, loop of SF162 shields an as yet unidentified region of the HIV envelope rich in neutralization epitopes and that the overall structure of this region appears to be conserved among clade B, C, D, E, and F HIV-1 PR isolates.
Env-Independent Protection Induced by Live, Attenuated Simian Immunodeficiency Virus Vaccines
Authors: Gundlach Björn R. , Reiprich Stefan , Sopper Sieghart , Means Robert E. , Dittmer Ulf , Mätz-Rensing Kerstin , Stahl-Hennig Christiane , Überla Klaus 
Published: December 2019

ABSTRACT

Live attenuated simian immunodeficiency viruses (SIV), such as
nef
deletion mutants, are the most effective vaccines tested in the SIV-macaque model so far. To modulate the antiviral immune response induced by live attenuated SIV vaccines, we had previously infected rhesus monkeys with a
nef
deletion mutant of SIV expressing interleukin 2 (SIV-IL2) (B. R. Gundlach, H. Linhart, U. Dittmer, S. Sopper, S. Reiprich, D. Fuchs, B. Fleckenstein, G. Hunsmann, S. Stahl-Hennig, and K. Überla, J. Virol. 71:2225–2232, 1997). In the present study, SIV-IL2-infected macaques and macaques infected with the
nef
deletion mutant SIVΔNU were challenged with pathogenic SIV 9 to 11 months postvaccination. In contrast to the results with naive control monkeys, no challenge virus could be isolated from the SIV-IL2- and SIVΔNU-infected macaques. However, challenge virus sequences could be detected by nested PCR in some of the vaccinated macaques. To determine the role of immune responses directed against Env of SIV, four vaccinated macaques were rechallenged with an SIV-murine leukemia virus (MLV) hybrid in which the
env
gene of SIV had been functionally replaced by the
env
gene of amphotropic MLV. All vaccinated macaques were protected from productive infection with the SIV-MLV hybrid in the absence of measurable neutralizing antibodies, while two naive control monkeys were readily infected. Since the SIV-MLV hybrid uses the MLV Env receptor Pit2 and not CD4 and a coreceptor for virus entry, chemokine inhibition and receptor interference phenomena were not involved in protection. These results indicate that the protective responses induced by live attenuated SIV vaccines can be independent of host immune reactions directed against Env.
Role of the TRAF Binding Site and NF-κB Activation in Epstein-Barr Virus Latent Membrane Protein 1-Induced Cell Gene Expression
Authors: Devergne Odile , McFarland Ellen Cahir , Mosialos George , Izumi Kenneth M. , Ware Carl F. , Kieff Elliott 
Published: December 2019

ABSTRACTIn this study, we investigated the induction of cellular gene expression by the Epstein-Barr Virus (EBV) latent membrane protein 1 (LMP1). Previously, LMP1 was shown to induce the expression of ICAM-1, LFA-3, CD40, and EBI3 in EBV-negative Burkitt lymphoma (BL) cells and of the epidermal growth factor receptor (EGF-R) in epithelial cells. We now show that LMP1 expression also increased Fas and tumor necrosis factor receptor-associated factor 1 (TRAF1) in BL cells. LMP1 mediates NF-κB activation via two independent domains located in its C-terminal cytoplasmic tail, a TRAF-interacting site that associates with TRAF1, -2, -3, and -5 through a PXQXT/S core motif and a TRADD-interacting site. In EBV-transformed B cells or transiently transfected BL cells, significant amounts of TRAF1, -2, -3, and -5 are associated with LMP1. In epithelial cells, very little TRAF1 is expressed, and only TRAF2, -3, and -5, are significantly complexed with LMP1. The importance of TRAF binding to the PXQXT/S motif in LMP1-mediated gene induction was studied by using an LMP1 mutant that contains alanine point mutations in this motif and fails to associate with TRAFs. This mutant, LMP1(P204A/Q206A), induced 60% of wild-type LMP1 NF-κB activation and had approximately 60% of wild-type LMP1 effect on Fas, ICAM-1, CD40, and LFA-3 induction. In contrast, LMP1(P204A/Q206A) was substantially more impaired in TRAF1, EBI3, and EGF-R induction. Thus, TRAF binding to the PXQXT/S motif has a nonessential role in up-regulating Fas, ICAM-1, CD40, and LFA-3 expression and a critical role in up-regulating TRAF1, EBI3, and EGF-R expression. Further, D1 LMP1, an LMP1 mutant that does not aggregate failed to induce TRAF1, EBI3, Fas, ICAM-1, CD40, and LFA-3 expression confirming the essential role for aggregation in LMP1 signaling. Overexpression of a dominant form of IκBα blocked LMP1-mediated TRAF1, EBI3, Fas, ICAM-1, CD40, and LFA-3 up-regulation, indicating that NF-κB is an important component of LMP1-mediated gene induction from both the TRAF- and TRADD-interacting sites.
The Coxsackievirus-Adenovirus Receptor Protein Can Function as a Cellular Attachment Protein for Adenovirus Serotypes from Subgroups A, C, D, E, and F
Authors: Roelvink Peter W. , Lizonova Alena , Lee Jennifer G. M. , Li Yuan , Bergelson Jeffrey M. , Finberg Robert W. , Brough Douglas E. , Kovesdi Imre , Wickham Thomas J. 
Published: December 2019

ABSTRACT

Attachment of an adenovirus (Ad) to a cell is mediated by the capsid fiber protein. To date, only the cellular fiber receptor for subgroup C serotypes 2 and 5, the so-called coxsackievirus-adenovirus receptor (CAR) protein, has been identified and cloned. Previous data suggested that the fiber of the subgroup D serotype Ad9 also recognizes CAR, since Ad9 and Ad2 fiber knobs cross-blocked each other’s cellular binding. Recombinant fiber knobs and
3
H-labeled Ad virions from serotypes representing all six subgroups (A to F) were used to determine whether the knobs cross-blocked the binding of virions from different subgroups. With the exception of subgroup B, all subgroup representatives cross-competed, suggesting that they use CAR as a cellular fiber receptor as well. This result was confirmed by showing that CAR, produced in a soluble recombinant form (sCAR), bound to nitrocellulose-immobilized virions from the different subgroups except subgroup B. Similar results were found for blotted fiber knob proteins. The subgroup F virus Ad41 has both short and long fibers, but only the long fiber bound sCAR. The sCAR protein blocked the attachment of all virus serotypes that bound CAR. Moreover, CHO cells expressing human CAR, in contrast to untransformed CHO cells, all specifically bound the sCAR-binding serotypes. We conclude therefore that Ad serotypes from subgroups A, C, D, E, and F all use CAR as a cellular fiber receptor.
Inactivation of Human Immunodeficiency Virus Type 1 Infectivity with Preservation of Conformational and Functional Integrity of Virion Surface Proteins
Authors: Rossio J. L. , Esser M. T. , Suryanarayana K. , Schneider D. K. , Bess J. W. , Vasquez G. M. , Wiltrout T. A. , Chertova E. , Grimes M. K. , Sattentau Q. , Arthur L. O. , Henderson L. E. , Lifson J. D. 
Published: December 2019

ABSTRACTWhole inactivated viral particles have been successfully used as vaccines for some viruses, but procedures historically used for inactivation can denature virion proteins. Results have been inconsistent, with enhancement of disease rather than protection seen in some notable instances following vaccination. We used the compound 2,2′-dithiodipyridine (aldrithiol-2; AT-2) to covalently modify the essential zinc fingers in the nucleocapsid (NC) protein of human immunodeficiency virus type 1 (HIV-1) or simian immunodeficiency virus (SIV) virions, thereby inactivating infectivity. The inactivated virus was not detectably infectious in vitro (up to 5 log units of inactivation). However, in contrast to virions inactivated by conventional methods such as heat or formalin treatment, viral and host cell-derived proteins on virion surfaces retained conformational and functional integrity. Thus, immunoprecipitation of AT-2-treated virions was comparable to precipitation of matched untreated virus, even when using antibodies to conformational determinants on gp120. AT-2 inactivated virions bound to CD4+target cells and mediated virus-induced, CD4-dependent “fusion from without” comparably to native virions. However, viral entry assays demonstrated that the viral life cycle of AT-2-treated virions was arrested before initiation of reverse transcription. The major histocompatibility complex (MHC) class II molecules on the surface of AT-2-treated virions produced from MHC class II-expressing cells retained the ability to support class II-dependent, superantigen-triggered proliferative responses by resting T lymphocytes. These findings indicate that inactivation via this method results in elimination of infectivity with preservation of conformational and functional integrity of virion surface proteins, including both virally encoded determinants and proteins derived from the host cells in which the virus was produced. Such inactivated virions should provide a promising candidate vaccine antigen and a useful reagent for experimentally probing the postulated involvement of virion surface proteins in indirect mechanisms of HIV-1 pathogenesis.
Processing of Proteinase Precursors and Their Effect on Hepatitis A Virus Particle Formation
Authors: Probst Christian , Jecht Monika , Gauss-Müller Verena 
Published: December 2019

ABSTRACT

Proteolytic processing of the picornaviral polyprotein mediated by the differential action of virus-encoded proteinase(s) is pivotal to both RNA genome replication and capsid formation. Possibly to enlarge the array of viral proteins, picornaviral polyprotein processing results in intermediate and mature products which apparently have distinct functions within the viral life cycle. For hepatitis A virus (HAV), we report here on the autoproteolysis of precursor polypeptides comprising the only viral proteinase, 3C
pro
, and on their role in viral particle formation. Following transient expression of a nested set of 3C
pro
-containing proteins (P3, 3ABC, 3BCD, 3CD, 3BC, and 3C) in eukaryotic cells, the extent of processing was determined by analyzing the cleavage products. The 3C/3D site was more efficiently cleaved than those at the 3A/3B and 3B/3C sites, leading to the accumulation of the intermediate product 3ABC. In the absence of 3A from the precursor, cleavage at the 3B/3C site was further reduced and a switch to an alternative 3C/3D site was observed. Coexpression of various parts of P3 with the precursor of the viral structural proteins P1-2A showed that all 3C-containing intermediates cleaved P1-2A with almost equal efficiency; however, viral particles carrying the neutralizing epitope form much more readily in the presence of the complete P3 domain than with parts of it. These data support the notion that efficient liberation of structural proteins from P1-2A is necessary but not sufficient for productive HAV capsid formation and suggest that the polypeptides flanking 3C
pro
promote the assembly of viral particles.