bluuebunny/crossref_metadata_2025_split huggingface dataset:
Recognition by Human Monoclonal Antibodies of Free and Complexed Peptides Representing the Prefusogenic and Fusogenic Forms of Human Immunodeficiency Virus Type 1 gp41
Authors: Gorny Miroslaw K. , Zolla-Pazner Susan 
Published: July 2002

ABSTRACTHuman immunodeficiency virus type 1 (HIV-1) entry into target cells appears to be triggered when two heptad repeat regions in the ectodomain of gp41 associate, converting the prefusogenic form of gp41 to a fusogenic form. Peptides from these two heptad repeat regions, designated N51 and C43, form a coiled coil consisting of an α-helical trimer of heterodimers which approximates the core of the fusogenic form of gp41. To understand the antigenic structures of gp41 in these two configurations, and to examine the specificity of anti-gp41 antibodies produced by HIV-1-infected individuals, human anti-gp41 monoclonal antibodies (MAbs) were tested for their reactivity against N51, C43, and the complex formed by these peptides. Of 11 MAbs, 7 reacted with the complex but with neither of the parent peptides. These MAbs reacted optimally with the N51-C43 complex prepared at a 1:1 ratio and appeared to recognize the fusogenic form of gp41 in which the two heptad repeat regions are associated to form the coiled coil. The existence of antibodies from HIV-infected humans that exclusively recognize the N51-C43 complex constitutes the first proof that the coiled-coil conformation of gp41 exists in vivo and is immunogenic. Two of the 11 MAbs were specific for the hydrophilic loop region of gp41 and failed to react with either peptide alone or with the peptide complex, while the remaining 2 MAbs reacted with peptide C43. One of these two latter MAbs, 98-6, also reacted well with the equimolar N51-C43 complex, while reactivity with C43 by the other MAb, 2F5, was inhibited by even small amounts of N51, suggesting that the interaction of these peptides occludes or disrupts the epitope recognized by MAb 2F5. MAbs 98-6 and 2F5 are also unusual among the MAbs tested in their ability to neutralize multiple primary HIV isolates, although 2F5 displays more broad and potent activity. The data suggest that anti-gp41 neutralizing activity is associated with specificity for a region in C43 which participates in complex formation with N51.
Thebfl-1Gene Is Transcriptionally Upregulated by the Epstein-Barr Virus LMP1, and Its Expression Promotes the Survival of a Burkitt's Lymphoma Cell Line
Authors: D'Souza Brendan , Rowe Martin , Walls Dermot 
Published: July 2002

ABSTRACTThe recently identifiedbfl-1gene (also known asA1orGRS), a homologue ofbcl-2, encodes an antiapoptotic protein that suppresses apoptosis induced by the p53 tumor suppressor protein and exhibits proliferative and potent cooperative transforming activities. We show that elevated levels ofbfl-1mRNA are a feature of Epstein-Barr virus (EBV)-immortalized B-cell lines and Burkitt's lymphoma cell lines expressing the full spectrum of EBV latent proteins. Using an EBV-negative Burkitt's lymphoma cell line in which the expression of EBV latent membrane protein 1 (LMP1) is inducibly regulated by tetracycline, we demonstrate that LMP1 expression coincides with a dramatic increase in the level ofbfl-1mRNA. Also in this system, an increase in the level of Bcl-2 protein was seen to occur earlier than that ofbcl-2mRNA, suggesting that both transcriptional and translational mechanisms are involved in the control of Bcl-2 expression by LMP-1. We show that elevatedbfl-1mRNA stability can contribute to this effect of LMP-1, thus providing evidence of a novel mechanism of gene regulation by this EBV protein. Upregulation ofbfl-1by LMP1 was not observed in the T-cell line Jurkat or the epithelial cell line C33A. Ectopic expression of Bfl-1 in an EBV-positive cell line exhibiting a latency type I infection protects against apoptosis induced by growth factor deprivation, thereby providing a functional role for Bfl-1 in this cellular context and adding Bfl-1 to the list of antiapoptotic proteins whose expression is modulated by EBV. This is the first report of the regulation ofbfl-1expression by a viral protein, and this novel finding may thus represent an important link between the EBV oncoprotein LMP1 and its cellular growth-transforming properties.
Inhibition of Human Immunodeficiency Virus Type 1 Replication in Primary CD4+T Lymphocytes, Monocytes, and Dendritic Cells by Cytotoxic T Lymphocytes
Authors: Severino Michael E. , Sipsas Nikolaos V. , Nguyen Phuong Thi , Kalams Spyros A. , Walker Bruce D. , Johnson R. Paul , Yang Otto O. 
Published: July 2002

ABSTRACTWe demonstrate that human immunodeficiency virus type 1 (HIV-1)-specific CD8+cytotoxic T lymphocytes (CTL) suppress HIV-1 replication in primary lymphocytes, monocytes, and dendritic cells individually. Viral inhibition is significantly diminished in lymphocyte-dendritic cell clusters, suggesting that these clusters in vivo could be sites where viral replication is more difficult to control by CTL.
CCR8 on Human Thymocytes Functions as a Human Immunodeficiency Virus Type 1 Coreceptor
Authors: Lee Shirley , Tiffany H. Lee , King Lisa , Murphy Philip M. , Golding Hana , Zaitseva Marina B. 
Published: July 2002

ABSTRACTTo determine whether human immunodeficiency virus type 1 (HIV-1) coreceptors besides CXCR4 and CCR5 are involved in HIV-1 infection of the thymus, we focused on CCR8, a receptor for the chemokine I-309, because of its high expression in the thymus. Similar levels of CCR8 mRNA were detected in immature and mature primary human thymocytes. Consistent with this, [125I]I-309 was shown to bind specifically and with similar affinity to the surface of immature and mature human thymocytes. Fusion of human thymocytes with cells expressing HIV-1 X4 or X4R5 envelope glycoprotein was inhibited by I-309 in a dose-dependent manner. In addition, I-309 partially inhibited productive infection of human thymocytes by X4, R5, and X4R5 HIV-1 strains. Our data provide the first evidence that CCR8 functions as an HIV-1 coreceptor on primary human cells and suggest that CCR8 may contribute to HIV-1-induced thymic pathogenesis.
Overexpression of p21
waf1
in Human T-Cell Lymphotropic Virus Type 1-Infected Cells and Its Association with Cyclin A/cdk2
Authors: de la Fuente Cynthia , Santiago Francisco , Chong Siew Yen , Deng Longwen , Mayhood Todd , Fu Peng , Stein Dana , Denny Thomas , Coffman Frederick , Azimi Nazli , Mahieux Renaud , Kashanchi Fatah 
Published: July 2002

ABSTRACT

Human T-cell lymphotropic virus type 1 (HTLV-1) is associated with adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). T-cell transformation is mainly due to the actions of the viral phosphoprotein Tax. Tax interacts with multiple transcriptional factors, aiding the transcription of many cellular genes. Here, we report that the cyclin-dependent kinase inhibitor p21/waf1 is overexpressed in all HTLV-1-infected cell lines tested as well as in ATL and HAM/TSP patient samples. Tax was found to be able to transactivate the endogenous p21/waf1 promoter, as detected by RNase protection, as well as activate a series of wild-type and 5′-deletion constructs linked to a luciferase reporter cassette. Wild-type but not a mutant form of Tax (M47) transactivated the p21/waf1 promoter in a p53-independent manner and utilized a minimal promoter that contained E2A and TATA box sequences. The p21/waf1 protein was reproducibly observed to be complexed with cyclin A/cdk2 and not with any other known G
1
, S, or G
2
/M cyclins. Functionally, the association of p21/cyclin A/cdk2 decreased histone H1 phosphorylation in vitro, as observed in immunoprecipitations followed by kinase assays, and affected other substrates, such as the C terminus of Rb protein involved in c-Abl and histone deacetylase-1 (HDAC1) regulation. Interestingly, upon the use of a stress signal, such as gamma-irradiation, we found that the p21/cyclin A/cdk2 complex was able to block all known phosphorylation sites on the Rb molecule. Finally, using elutriated cell cycle fractions and a stress signal, we observed that the HTLV-1-infected T cells containing wild-type Tax, which had been in early or mid-G
1
phase prior to gamma-irradiation, arrested in G
1
and did not undergo apoptosis. This may be an important mechanism for an oncogenic virus such as HTLV-1 to stop the host at the G
1
/S boundary and to repair the damaged DNA upon injury, prior to S-phase entry.
Identification of Domains of the Human Papillomavirus Type 11 E1 Helicase Involved in Oligomerization and Binding to the Viral Origin
Authors: Titolo Steve , Pelletier Alex , Pulichino Anne-Marie , Brault Karine , Wardrop Elizabeth , White Peter W. , Cordingley Michael G. , Archambault Jacques 
Published: July 2002

ABSTRACT
The E1 helicase of papillomavirus is required, in addition to host cell DNA replication factors, during the initiation and elongation phases of viral episome replication. During initiation, the viral E2 protein promotes the assembly of enzymatically active multimeric E1 complexes at the viral origin of DNA replication. In this study we used the two-hybrid system and chemical cross-linking to demonstrate that human papillomavirus type 11 (HPV11) E1 can self-associate in yeast and form hexamers in vitro in a reaction stimulated by single-stranded DNA. Self-association in yeast was most readily detected using constructs spanning the E1 C-terminal domain (amino acids 353 to 649) and was dependent on a minimal E1-E1 interaction region located between amino acids 353 and 431. The E1 C-terminal domain was also able to oligomerize in vitro but, in contrast to wild-type E1, did so efficiently in the absence of single-stranded DNA. Sequences located between amino acids 191 and 353 were necessary for single-stranded DNA to modulate oligomerization of E1 and were also required, together with the rest of the C terminus, for binding of E1 to the origin. Two regions within the C-terminal domain were identified as important for oligomerization: the ATP-binding domain and region A, which is located within the minimal E1-E1 interaction domain and is one of four regions of E1 that is highly conserved with the large T antigens of simian virus 40 and polyomavirus. Amino acid substitutions of highly conserved residues within the ATP-binding domain and region A were identified that reduced the ability of E1 to oligomerize and bind to the origin in vitro and to support transient DNA replication in vivo. These results support the notion that oligomerization of E1 occurs primarily through the C-terminal domain of the protein and is allosterically regulated by DNA and ATP. The bipartite organization of the E1 C-terminal domain is reminiscent of that found in other hexameric proteins and suggests that these proteins may oligomerize by a similar mechanism.
Construction and Characterization of Murine Cytomegaloviruses That Contain Transposon Insertions at Open Reading Frames m09 and M83
Authors: Zhan Xiaoyan , Lee Manfred , Xiao Jianqiao , Liu Fenyong 
Published: July 2002

ABSTRACT

A transposon derived from
Escherichia coli
Tn
3
was introduced into the genome of murine cytomegalovirus (MCMV) to generate a pool of viral mutants, including two recombinant viruses that contained the transposon sequence within open reading frames m09 and M83. Our studies provide the first direct evidence to suggest that m09 is not essential for viral replication in mouse NIH 3T3 cells. Studies in cultured cells and in both BALB/c-Byj and CB17 severe combined immunodeficient (SCID) mice indicated that the transposon insertion is stable during viral propagation both in vitro and in vivo. Moreover, the virus that contained the insertion mutation in m09 exhibited a titer similar to that of the wild-type virus in the salivary glands, lungs, livers, spleens, and kidneys of both the BALB/c and SCID mice and was as virulent as the wild-type virus in killing the SCID mice when these animals were intraperitoneally infected with these viruses. These results suggest that m09 is dispensable for viral growth in these organs and that the presence of the transposon sequence in the viral genome does not significantly affect viral replication in vivo. In contrast, the virus that contained the insertion mutation in M83 exhibited a titer of at least 60-fold lower than that of the wild-type virus in the organs of the SCID mice and was attenuated in killing the SCID mice. These results demonstrate the utility of using the Tn
3
-based system as a mutagenesis approach for studying the function of MCMV genes in both immunocompetent and immunodeficient animals.
Antibodies to CD9, a Tetraspan Transmembrane Protein, Inhibit Canine Distemper Virus-Induced Cell-Cell Fusion but Not Virus-Cell Fusion
Authors: Schmid Erik , Zurbriggen Andreas , Gassen Uta , Rima Bert , ter Meulen Volker , Schneider-Schaulies Jürgen 
Published: July 2002

ABSTRACTCanine distemper virus (CDV) causes a life-threatening disease in several carnivores including domestic dogs. Recently, we identified a molecule, CD9, a member of the tetraspan transmembrane protein family, which facilitates, and antibodies to which inhibit, the infection of tissue culture cells with CDV (strain Onderstepoort). Here we describe that an anti-CD9 monoclonal antibody (MAb K41) did not interfere with binding of CDV to cells and uptake of virus. In addition, in single-step growth experiments, MAb K41 did not induce differences in the levels of viral mRNA and proteins. However, the virus release of syncytium-forming strains of CDV, the virus-induced cell-cell fusion in lytically infected cultures, and the cell-cell fusion of uninfected with persistently CDV-infected HeLa cells were strongly inhibited by MAb K41. These data indicate that anti-CD9 antibodies selectively block virus-induced cell-cell fusion, whereas virus-cell fusion is not affected.
Fast Screening Procedures for Random Transposon Libraries of Cloned Herpesvirus Genomes: Mutational Analysis of Human Cytomegalovirus Envelope Glycoprotein Genes
Authors: Hobom Urs , Brune Wolfram , Messerle Martin , Hahn Gabriele , Koszinowski Ulrich H. 
Published: July 2002

ABSTRACT

We have cloned the human cytomegalovirus (HCMV) genome as an infectious bacterial artificial chromosome (BAC) in
Escherichia coli
. Here, we have subjected the HCMV BAC to random transposon (Tn) mutagenesis using a Tn
1721
-derived insertion sequence and have provided the conditions for excision of the BAC cassette. We report on a fast and efficient screening procedure for a Tn insertion library. Bacterial clones containing randomly mutated full-length HCMV genomes were transferred into 96-well microtiter plates. A PCR screening method based on two Tn primers and one primer specific for the desired genomic position of the Tn insertion was established. Within three consecutive rounds of PCR a Tn insertion of interest can be assigned to a specific bacterial clone. We applied this method to retrieve mutants of HCMV envelope glycoprotein genes. To determine the infectivities of the mutant HCMV genomes, the DNA of the identified BACs was transfected into permissive fibroblasts. In contrast to BACs with mutations in the genes coding for gB, gH, gL, and gM, which did not yield infectious virus, BACs with disruptions of open reading frame
UL4
(gp48) or
UL74
(gO) were viable, although gO-deficient viruses showed a severe growth deficit. Thus, gO (
UL74
), a component of the glycoprotein complex III, is dispensable for viral growth. We conclude that our approach of PCR screening for Tn insertions will greatly facilitate the functional analysis of herpesvirus genomes.
Molecular Cloning and Functional Analysis of Three Type D Endogenous Retroviruses of Sheep Reveal a Different Cell Tropism from That of the Highly Related Exogenous Jaagsiekte Sheep Retrovirus
Authors: Palmarini Massimo , Hallwirth Claus , York Denis , Murgia Claudio , de Oliveira Tulio , Spencer Thomas , Fan Hung 
Published: July 2002

ABSTRACT

Integrated into the sheep genome are 15 to 20 copies of type D endogenous loci that are highly related to two exogenous oncogenic viruses, jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus (ENTV). The exogenous viruses cause infectious neoplasms of the respiratory tract in small ruminants. In this study, we molecularly cloned three intact type D endogenous retroviruses of sheep (enJS56A1, enJS5F16, and enJS59A1; collectively called
enJRSV
s) and analyzed their genomic structures, their phylogenies with respect to their exogenous counterparts, their capacity to form viral particles, and the expression specificities of their long terminal repeats (LTRs). In addition, the pattern of expression of
enJSRV
s in vivo was studied by in situ hybridization. All of the three
enJSRV
proviruses had open reading frames for at least one of the structural genes. In particular, enJS56A1 had open reading frames for all structural genes, but it could not assemble viral particles when highly expressed in human 293T cells. We localized the defect for viral assembly in the first two-thirds of the
gag
gene by making a series of chimeras between enJS56A1 and the exogenous infectious molecular clone JSRV
21
. Phylogenetic analysis distinguished five ovine type D retroviruses:
enJSRV
groups A and B, ENTV, and two exogenous JSRV groups (African versus United Kingdom/North America isolates). Transient transfection assays indicated that the LTRs of the three
enJSRV
s were not preferentially active in differentiated lung epithelial cells. This suggests that the pulmonary tropic JSRV developed from a type D retrovirus that did not have lung specificity. Consistent with this, in situ hybridization of a panel of normal ovine tissues revealed high expression of
enJSRV
mRNA in the luminal epithelium and glandular epithelium of the uterus; lower expression was localized in the lamina propria of the gut and in the bronchiolar epithelium of the lungs.
The Collagen Repeat Sequence Is a Determinant of the Degree of Herpesvirus Saimiri STP Transforming Activity
Authors: Choi Joong-Kook , Ishido Satoshi , Jung Jae U. 
Published: July 2002

ABSTRACTHerpesvirus saimiri (HVS) is divided into three subgroups, A, B, and C, based on sequence divergence at the left end of genomic DNA in which the saimiri transforming protein (STP) resides. Subgroup A and C strains transform primary common marmoset lymphocytes to interleukin-2-independent growth, whereas subgroup B strains do not. To investigate the nononcogenic phenotype of the subgroup B viruses, STP genes from seven subgroup B virus isolates were cloned and sequenced. Consistent with the lack of oncogenic activity of HVS subgroup B viruses, STP-B was deficient for transforming activity in rodent fibroblast cells. Sequence comparison reveals that STP-B lacks the signal-transducing modules found in STP proteins of the other subgroups, collagen repeats and an authentic SH2 binding motif. Substitution mutations demonstrated that the lack of collagen repeats but not an SH2 binding motif contributed to the nontransforming phenotype of STP-B. Introduction of the collagen repeat sequence induced oligomerization of STP-B, resulting in activation of NF-κB activity and deregulation of cell growth control. These results demonstrate that the collagen repeat sequence is a determinant of the degree of HVS STP transforming activity.
Transfer of Specificity for Human Immunodeficiency Virus Type 1 into Primary Human T Lymphocytes by Introduction of T-Cell Receptor Genes
Authors: Cooper Laurence J. N. , Kalos Michael , Lewinsohn Deborah A. , Riddell Stanley R. , Greenberg Philip D. 
Published: July 2002

ABSTRACTThe introduction of genes encoding T-cell receptor (TCR) chains specific for human immunodeficiency virus into T cells of infected patients represents a means to quantitatively and qualitatively improve immunity to the virus. Our results demonstrate that the high level of TCR expression required for physiologic functioning can be reproducibly achieved with retroviral vectors encoding full-length unmodified TCR chains under the control of a strong internal constitutive phosphoglycerate kinase promoter.
Identification of Dominant Optimal HLA-B60- and HLA-B61-Restricted Cytotoxic T-Lymphocyte (CTL) Epitopes: Rapid Characterization of CTL Responses by Enzyme-Linked Immunospot Assay
Authors: Altfeld Marcus A. , Trocha Alicja , Eldridge Robert L. , Rosenberg Eric S. , Phillips Mary N. , Addo Marylyn M. , Sekaly Rafick P. , Kalams Spyros A. , Burchett Sandra A. , McIntosh Kenneth , Walker Bruce D. , Goulder Philip J. R. 
Published: July 2002

ABSTRACTHuman immunodeficiency virus type 1 (HIV-1)-specific cytotoxic T-lymphocyte (CTL) responses play a major role in the antiviral immune response, but the relative contribution of CTL responses restricted by different HLA class I molecules is less well defined. HLA-B60 or the related allele B61 is expressed in 10 to 20% of Caucasoid populations and is even more highly prevalent in Asian populations, but yet no CTL epitopes restricted by these alleles have been defined. Here we report the definition of five novel HLA-B60-restricted HIV-1-specific CTL epitopes, using peripheral blood mononuclear cells in enzyme-linked immunospot (Elispot) assays and using CTL clones and lines in cytolytic assays. The dominant HLA-B60-restricted epitope, Nef peptide KEKGGLEGL, was targeted by all eight subjects with B60 and also by both subjects with B61 studied. This study additionally establishes the utility of the Elispot assay as a more rapid and efficient method of defining novel CTL epitopes. This approach will help to define new CTL epitopes that may play an important role in the immune control of HIV-1.
Remodeling the Endoplasmic Reticulum by Poliovirus Infection and by Individual Viral Proteins: an Autophagy-Like Origin for Virus-Induced Vesicles
Authors: Suhy David A. , Giddings Thomas H. , Kirkegaard Karla 
Published: July 2002

ABSTRACT
All positive-strand RNA viruses of eukaryotes studied assemble RNA replication complexes on the surfaces of cytoplasmic membranes. Infection of mammalian cells with poliovirus and other picornaviruses results in the accumulation of dramatically rearranged and vesiculated membranes. Poliovirus-induced membranes did not cofractionate with endoplasmic reticulum (ER), lysosomes, mitochondria, or the majority of Golgi-derived or endosomal membranes in buoyant density gradients, although changes in ionic strength affected ER and virus-induced vesicles, but not other cellular organelles, similarly. When expressed in isolation, two viral proteins of the poliovirus RNA replication complex, 3A and 2C, cofractionated with ER membranes. However, in cells that expressed 2BC, a proteolytic precursor of the 2B and 2C proteins, membranes identical in buoyant density to those observed during poliovirus infection were formed. When coexpressed with 2BC, viral protein 3A was quantitatively incorporated into these fractions, and the membranes formed were ultrastructurally similar to those in poliovirus-infected cells. These data argue that poliovirus-induced vesicles derive from the ER by the action of viral proteins 2BC and 3A by a mechanism that excludes resident host proteins. The double-membraned morphology, cytosolic content, and apparent ER origin of poliovirus-induced membranes are all consistent with an autophagic origin for these membranes.
Granzyme A, a Noncytolytic Component of CD8+Cell Granules, Restricts the Spread of Herpes Simplex Virus in the Peripheral Nervous Systems of Experimentally Infected Mice
Authors: Pereira Rosemarie A. , Simon Markus M. , Simmons Anthony 
Published: July 2002

ABSTRACTControl of ganglionic herpes simplex virus (HSV) infection depends on CD8+cells but not on the death of infected neurons. Primarily, perforin and granzyme B mediate CD8+cell cytotoxicity, whereas the in vivo functions of granzyme A, a third granule protein, are unknown. Here, it is shown that granzyme A restricts the interneuronal spread of HSV and significantly influences ganglionic virus load.