An Evolutionarily Young Polar Bear (Ursus maritimus) Endogenous Retrovirus Identified from Next Generation Sequence Data.

PubMed

Tsangaras, Kyriakos; Mayer, Jens; Alquezar-Planas, David E; Greenwood, Alex D

2015-11-24

Transcriptome analysis of polar bear (Ursus maritimus) tissues identified sequences with similarity to Porcine Endogenous Retroviruses (PERV). Based on these sequences, four proviral copies and 15 solo long terminal repeats (LTRs) of a newly described endogenous retrovirus were characterized from the polar bear draft genome sequence. Closely related sequences were identified by PCR analysis of brown bear (Ursus arctos) and black bear (Ursus americanus) but were absent in non-Ursinae bear species. The virus was therefore designated UrsusERV. Two distinct groups of LTRs were observed including a recombinant ERV that contained one LTR belonging to each group indicating that genomic invasions by at least two UrsusERV variants have recently occurred. Age estimates based on proviral LTR divergence and conservation of integration sites among ursids suggest the viral group is only a few million years old. The youngest provirus was polar bear specific, had intact open reading frames (ORFs) and could potentially encode functional proteins. Phylogenetic analyses of UrsusERV consensus protein sequences suggest that it is part of a pig, gibbon and koala retrovirus clade. The young age estimates and lineage specificity of the virus suggests UrsusERV is a recent cross species transmission from an unknown reservoir and places the viral group among the youngest of ERVs identified in mammals.

An Evolutionarily Young Polar Bear (Ursus maritimus) Endogenous Retrovirus Identified from Next Generation Sequence Data

PubMed Central

Tsangaras, Kyriakos; Mayer, Jens; Alquezar-Planas, David E.; Greenwood, Alex D.

2015-01-01

Transcriptome analysis of polar bear (Ursus maritimus) tissues identified sequences with similarity to Porcine Endogenous Retroviruses (PERV). Based on these sequences, four proviral copies and 15 solo long terminal repeats (LTRs) of a newly described endogenous retrovirus were characterized from the polar bear draft genome sequence. Closely related sequences were identified by PCR analysis of brown bear (Ursus arctos) and black bear (Ursus americanus) but were absent in non-Ursinae bear species. The virus was therefore designated UrsusERV. Two distinct groups of LTRs were observed including a recombinant ERV that contained one LTR belonging to each group indicating that genomic invasions by at least two UrsusERV variants have recently occurred. Age estimates based on proviral LTR divergence and conservation of integration sites among ursids suggest the viral group is only a few million years old. The youngest provirus was polar bear specific, had intact open reading frames (ORFs) and could potentially encode functional proteins. Phylogenetic analyses of UrsusERV consensus protein sequences suggest that it is part of a pig, gibbon and koala retrovirus clade. The young age estimates and lineage specificity of the virus suggests UrsusERV is a recent cross species transmission from an unknown reservoir and places the viral group among the youngest of ERVs identified in mammals. PMID:26610552

Immunising with the transmembrane envelope proteins of different retroviruses including HIV-1

PubMed Central

Denner, Joachim

2013-01-01

The induction of neutralizing antibodies is a promising way to prevent retrovirus infections. Neutralizing antibodies are mainly directed against the envelope proteins, which consist of two molecules, the surface envelope (SU) protein and the transmembrane envelope (TM) protein. Antibodies broadly neutralizing the human immunodeficiencvy virus-1 (HIV-1) and binding to the TM protein gp41 of the virus have been isolated from infected individuals. Their epitopes are located in the membrane proximal external region (MPER). Since there are difficulties to induce such neutralizing antibodies as basis for an effective AIDS vaccine, we performed a comparative analysis immunising with the TM proteins of different viruses from the family Retroviridae. Both subfamilies, the Orthoretrovirinae and the Spumaretrovirinae were included. In this study, the TM proteins of three gammaretroviruses including (1) the porcine endogenous retrovirus (PERV), (2) the Koala retrovirus (KoRV), (3) the feline leukemia virus (FeLV), of two lentiviruses, HIV-1, HIV-2, and of two spumaviruses, the feline foamy virus (FFV) and the primate foamy virus (PFV) were used for immunisation. Whereas in all immunisation studies binding antibodies were induced, neutralizing antibodies were only found in the case of the gammaretroviruses. The induced antibodies were directed against the MPER and the fusion peptide proximal region (FPPR) of their TM proteins; however only the antibodies against the MPER were neutralizing. Most importantly, the epitopes in the MPER were localized in the same position as the epitopes of the antibodies broadly neutralizing HIV-1 in the TM protein gp41 of HIV-1, indicating that the MPER is an effective target for the neutralization of retroviruses. PMID:23249763

Ultrasound enhances retrovirus-mediated gene transfer.

PubMed

Naka, Toshio; Sakoda, Tsuyoshi; Doi, Takashi; Tsujino, Takeshi; Masuyama, Tohru; Kawashima, Seinosuke; Iwasaki, Tadaaki; Ohyanagi, Mitsumasa

2007-01-01

Viral vector systems are efficient for transfection of foreign genes into many tissues. Especially, retrovirus based vectors integrate the transgene into the genome of the target cells, which can sustain long term expression. However, it has been demonstrated that the transduction efficiency using retrovirus is relatively lower than those of other viruses. Ultrasound was recently reported to increase gene expression using plasmid DNA, with or without, a delivery vehicle. However, there are no reports, which show an ultrasound effect to retrovirus-mediated gene transfer efficiency. Retrovirus-mediated gene transfer systems were used for transfection of 293T cells, bovine aortic endothelial cells (BAECs), rat aortic smooth muscle cells (RASMCs), and rat skeletal muscle myoblasts (L6 cells) with beta-galactosidase (beta-Gal) genes. Transduction efficiency and cell viability assay were performed on 293T cells that were exposed to varying durations (5 to 30 seconds) and power levels (1.0 watts/cm(2) to 4.0 watts/cm(2)) of ultrasound after being transduced by a retrovirus. Effects of ultrasound to the retrovirus itself was evaluated by transduction efficiency of 293T cells. After exposure to varying power levels of ultrasound to a retrovirus for 5 seconds, 293T cells were transduced by a retrovirus, and transduction efficiency was evaluated. Below 1.0 watts/cm(2) and 5 seconds exposure, ultrasound showed increased transduction efficiency and no cytotoxicity to 293T cells transduced by a retrovirus. Also, ultrasound showed no toxicity to the virus itself at the same condition. Exposure of 5 seconds at the power of 1.0 watts/cm(2) of an ultrasound resulted in significant increases in retrovirus-mediated gene expression in all four cell types tested in this experiment. Transduction efficiencies by ultrasound were enhanced 6.6-fold, 4.8-fold, 2.3-fold, and 3.2-fold in 293T cells, BAECs, RASMCs, and L6 cells, respectively. Furthermore, beta-Gal activities were also increased

Expression of the pol gene of human endogenous retroviruses HERV-K and -W in leukemia patients.

PubMed

Bergallo, Massimiliano; Montanari, Paola; Mareschi, Katia; Merlino, Chiara; Berger, Massimo; Bini, Ilaria; Daprà, Valentina; Galliano, Ilaria; Fagioli, Franca

2017-12-01

The human endogenous retroviruses (HERVs) are a family of endogenous retroviruses that integrated into the germ cell DNA of primates over 30 million years ago. HERV expression seems impaired in several diseases, ranging from autoimmune to neoplastic disorders. The purpose of this study was to evaluate the overall endogenous retroviral transcription profile in bone marrow (BM) samples. A total of 30 paediatric high-risk leukaemia patients (lymphoid and myeloid malignancies) were tested for HERVs virus gene expression. Our findings show that HERV-K expression was significantly higher in leukaemia patients when compared to healthy donors of a similar median age. We observed a significantly high expression of HERV-K in acute lymphoblastic leukemia (ALL) patients. In this study, we also found a relative overexpression of the endogenous retrovirus HERV-K in BM cells from the majority of leukemia samples analyzed, in particular in ALL. This overexpression might be related to lymphatic leukemogenesis and it warrants further investigations.

Induction of neutralizing antibodies specific for the envelope proteins of the koala retrovirus by immunization with recombinant proteins or with DNA.

PubMed

Fiebig, Uwe; Dieckhoff, Britta; Wurzbacher, Christian; Möller, Annekathrin; Kurth, Reinhard; Denner, Joachim

2015-04-30

The koala retrovirus (KoRV) is the result of a transspecies transmission of a gammaretrovirus with fatal consequences for the new host. Like many retroviruses, KoRV induces lymphoma, leukemia and an immunodeficiency that is associated with opportunistic infections in the virus-infected animals. We recently reported the induction of neutralizing antibodies by immunization with the recombinant ectodomain of the transmembrane envelope protein p15E of KoRV. Since the neutralization titers of the p15E-specific sera were only moderate, we investigated the use of the surface envelope protein gp70 to induce neutralizing antibodies. We immunized rats and goats with the recombinant gp70 protein of the KoRV, an unglycosylated protein of 52kD (rgp70/p52) or with the corresponding DNA. In parallel we immunized with recombinant rp15E or with a combination of rp15E and rgp70/p52. In all cases binding and neutralizing antibodies were induced. The gp70-specific sera had titers of neutralizing antibodies that were 15-fold higher than the p15E-specific sera. Combining rp15E and rgp70/p52 did not significantly increase neutralizing titers compared to rgp70/p52 alone. High titers of neutralizing antibodies specific for gp70 were also induced by immunization with DNA. Since KoRV and PERV are closely related, we investigated cross-neutralization of the antisera. The antisera against p15E and gp70 of PERV and KoRV inhibited infection by both viruses. The envelope proteins of the KoRV may therefore form the basis of an effective preventive vaccine to protect uninfected koalas from infection and possibly an immunotherapeutic treatment for those already infected.

An effective method for the quantitative detection of porcine endogenous retrovirus in pig tissues.

PubMed

Zhang, Peng; Yu, Ping; Wang, Wei; Zhang, Li; Li, Shengfu; Bu, Hong

2010-05-01

Xenotransplantation shows great promise for providing a virtually limitless supply of cells, tissues, and organs for a variety of therapeutical procedures. However, the potential of porcine endogenous retrovirus (PERV) as a human-tropic pathogen, particularly as a public health risk, is a major concern for xenotransplantation. This study focus on the detection of copy number in various tissues and organs in Banna Minipig Inbreed (BMI) from 2006 to 2007 in West China Hospital, Sichuan University. Real-time quantitative polymerase chain reaction (SYBR Green I) was performed in this study. The results showed that the pol gene had the most copy number in tissues compared with gag, envA, and envB. Our experiment will offer a rapid and accurate method for the detection of the copy number in various tissues and was especially suitable for the selection of tissues or organs in future clinical xenotransplantation.

Porcine endogenous retroviral nucleic acid in peripheral tissues is associated with migration of porcine cells post islet transplant.

PubMed

Binette, Tanya M; Seeberger, Karen L; Lyon, James G; Rajotte, Ray V; Korbutt, Gregory S

2004-07-01

Porcine islets represent an alternative source of insulin-producing tissue, however, porcine endogenous retrovirus (PERV) remains a concern. In this study, SCID mice were transplanted with nonencapsulated (non-EC), microencapsulated (EC) or macroencapsulated (in a TheraCyte trade mark device) neonatal porcine islets (NPIs), and peripheral tissues were screened for presence of viral DNA and mRNA. To understand the role of an intact immune system in PERV incidence, mice with established NPI grafts were reconstituted with splenocytes. Peripheral tissues were screened for PERV and porcine DNA using PCR. Tissues with positive DNA were analyzed for PERV mRNA using RT-PCR. No significant difference was observed between non-EC and EC transplants regarding presence of PERV or porcine-specific DNA or mRNA. In reconstituted animals, little PERV or porcine DNA, and no PERV mRNA was detected. No PERV or porcine-specific DNA was observed in animals implanted with a TheraCyte trade mark device. In conclusion, an intact immune system significantly lowered the presence of PERV. Microencapsulation of islets did not alter PERV presence, however, macroencapsulation in the TheraCyte device did. Lower PERV incidence coincided with lower porcine DNA in peripheral tissues, linking the presence of PERV to migration of porcine cells.

Susceptibility of human liver cells to porcine endogenous retrovirus.

PubMed

Lin, Xinzi; Qi, Lin; Li, Zhiguo; Chi, Hao; Lin, Wanjun; Wang, Yan; Jiang, Zesheng; Pan, Mingxin; Gao, Yi

2013-12-01

The risk of porcine endogenous retrovirus infection is a major barrier for pig-to-human xenotransplant. Porcine endogenous retrovirus, present in porcine cells, can infect many human and nonhuman primate cells in vitro, but there is no evidence available about in vitro infection of human liver cells. We investigated the susceptibility of different human liver cells to porcine endogenous retrovirus. The supernatant from a porcine kidney cell line was added to human liver cells, including a normal hepatocyte cell line (HL-7702 cells), primary hepatocytes (Phh cells), and a liver stellate cell line (Lx-2 cells), and to human embryonic kidney cells as a reference control. Expression of the porcine endogenous retrovirus antigen p15E in the human cells was evaluated with polymerase chain reaction, reverse transcription-polymerase chain reaction, and Western blot. The porcine endogenous retrovirus antigen p15E was not expressed in any human liver cells (HL-7702, Phh, or Lx-2 cells) that had been exposed to supernatants from porcine kidney cell lines. Porcine endogenous retrovirus-specific fragments were amplified in human kidney cells. Human liver cells tested were not susceptible to infection by porcine endogenous retrovirus. Therefore, not all human cells are susceptible to porcine endogenous retrovirus.

Computational Evaluation of the Strict Master and Random Template Models of Endogenous Retrovirus Evolution

PubMed Central

Nascimento, Fabrícia F.; Rodrigo, Allen G.

2016-01-01

Transposable elements (TEs) are DNA sequences that are able to replicate and move within and between host genomes. Their mechanism of replication is also shared with endogenous retroviruses (ERVs), which are also a type of TE that represent an ancient retroviral infection within animal genomes. Two models have been proposed to explain TE proliferation in host genomes: the strict master model (SMM), and the random template (or transposon) model (TM). In SMM only a single copy of a given TE lineage is able to replicate, and all other genomic copies of TEs are derived from that master copy. In TM, any element of a given family is able to replicate in the host genome. In this paper, we simulated ERV phylogenetic trees under variations of SMM and TM. To test whether current phylogenetic programs can recover the simulated ERV phylogenies, DNA sequence alignments were simulated and maximum likelihood trees were reconstructed and compared to the simulated phylogenies. Results indicate that visual inspection of phylogenetic trees alone can be misleading. However, if a set of statistical summaries is calculated, we are able to distinguish between models with high accuracy by using a data mining algorithm that we introduce here. We also demonstrate the use of our data mining algorithm with empirical data for the porcine endogenous retrovirus (PERV), an ERV that is able to replicate in human and pig cells in vitro. PMID:27649303

Design of retrovirus vectors for transfer and expression of the human. beta. -globin gene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miller, A.D.; Bender, M.A.; Harris, E.A.S.

1988-11-01

Regulated expression of the human ..beta..-globin gene has been demonstrated in cultured murine erythroleukemia cells and in mice after retrovirus-mediated gene transfer. However, the low titer of recombinant viruses described to date results in relatively inefficient gene transfer, which limits their usefulness for animal studies and for potential gene therapy in humans for diseases involving defective ..beta..-globin genes. The authors found regions that interfered with virus production within intron 2 of the ..beta..-globin gene and on both sides of the gene. The flanking regions could be removed, but intron 2 was required for ..beta..-globin expression. Inclusion of ..beta..-globin introns necessitatesmore » an antisense orientation of the gene within the retrovirus vector. However, they found no effect of the antisense ..beta..-globin transcription on virus production. A region downstream of the ..beta..-globin gene that stimulates expression of the gene in transgenic mice was included in the viruses without detrimental effects on virus titer. Virus titers of over 10/sup 6/ CFU/ml were obtained with the final vector design, which retained the ability to direct regulated expression of human ..beta..-globin in murine erythroleukemia cells. The vector also allowed transfer and expression of the human ..beta..-globin gene in hematopoietic cells (CFU-S cells) in mice.« less

The potential roles of endogenous retroviruses in autoimmunity.

PubMed

Nakagawa, K; Harrison, L C

1996-08-01

Endogenous retroviruses (ERVs) are estimated to comprise up to 1% of human DNA. While the genome of many ERVs is interrupted by termination codons, deletions or frame shift mutations, some ERVs are transcriptionally active and recent studies reveal protein expression or particle formation by human ERVs. ERVs have been implicated as aetiological agents of autoimmune disease, because of their structural and sequence similarities to exogenous retroviruses associated with immune dysregulation and their tissue-specific or differentiation-dependent expression. In fact, retrovirus-like particles distinct from those of known exogenous retroviruses and immune responses to ERV proteins have been observed in autoimmune disease. Quantitatively or structurally aberrant expression of normally cryptic ERVs, induced by environmental or endogenous factors, could initiate autoimmunity through direct or indirect mechanisms. ERVs may lead to immune dysregulation as insertional mutagens or cis-regulatory elements of cellular genes involved in immune function. ERVs may also encode elements like tax in human T-lymphotrophic virus type I (HTLV-I) or tat in human immunodeficiency virus-I (HIV-I) that are capable of transactivating cellular genes. More directly, human ERV gene products themselves may be immunologically active, by analogy with the superantigen activity in the long terminal repeat (LTR) of mouse mammary tumour viruses (MMTV) and the non-specific immunosuppressive activity in mammalian type C retrovirus env protein. Alternatively, increased expression of an ERV protein, or expression of a novel ERV protein not expressed in the thymus during acquisition of immune tolerance, may lead to its perception as a neoantigen. Paraneoplastic syndromes raise the possibility that novel ERV-encoded epitopes expressed by a tumour elicit immunity to cross-reactive epitopes in normal tissues. Recombination events between different but related ERVs, to whose products the host is immunologically

Prolonged expression of MHC class I - peptide expression in bone marrow derived retrovirus transfected matured dendritic cells by continuous centrifugation in the presence of IL-4.

PubMed

Hettihewa, L M

2011-11-01

Dendritic cells (DCs) are potent antigen presenting cells which proceed from immature to a mature stage during their differentiation. There are several methods of obtaining long lasting mature antigen expressing DCs and different methods show different levels of antigen expressions. We investigated bone marrow derived DCs for the degree of maturation and genetically engineered antigen presentation in the presence of interleukin-4 (IL-4) as a maturity enhancer. DCs and transfected retrovirus were cultured together in the presence of granulocyte-macrophage colony stimulating factor (GMCSF)-IL4, GMCSF +IL4, lipopolysaccharide (LPS). B 7.1, B7.2 and CD11c were measured by the degree of immune fluorescence using enhanced green fluorescent protein (EGFP) shuttled retrovirus transfected antigen. Degree of MHC class I molecule with antigen presentation of antigen was also evaluated by fluorescence activated cell sorting. The antigen presenting capacity of transfected DCs was investigated. Bone marrow DCs were generated in the presence of GMCSF and IL-4 in vitro. Dividing bone marrow cells were infected with EGFP shuttled retrovirus expressing SSP2 by prolonged centrifugation for three consecutive days from day 5, 6 and 7 and continued to culture in the presence of GMSCF and IL-4 until day 8. IL-4 as a cytokine increased the maturation of retrovirus transfected DCs by high expression of B 7-1 and B 7-2. Also, IL-4 induced DC enhanced by the prolonged centrifugation and it was shown by increased antigen presentation of these dendric cells as antigen presenting cell (APC). Cytolytic effects were significantly higher in cytotoxic T cell response (CTLs) mixed with transfected DCs than CTLs mixed with pulsed DCs. There was an enhanced antigen presentation by prolonged expression of antigen loaded MHC class I receptors in DCs in the presence of IL-4 by prolonged centrifugation.

PD-L1 Expression on Retrovirus-Infected Cells Mediates Immune Escape from CD8+ T Cell Killing.

PubMed

Akhmetzyanova, Ilseyar; Drabczyk, Malgorzata; Neff, C Preston; Gibbert, Kathrin; Dietze, Kirsten K; Werner, Tanja; Liu, Jia; Chen, Lieping; Lang, Karl S; Palmer, Brent E; Dittmer, Ulf; Zelinskyy, Gennadiy

2015-10-01

Cytotoxic CD8+ T Lymphocytes (CTL) efficiently control acute virus infections but can become exhausted when a chronic infection develops. Signaling of the inhibitory receptor PD-1 is an important mechanism for the development of virus-specific CD8+ T cell dysfunction. However, it has recently been shown that during the initial phase of infection virus-specific CD8+ T cells express high levels of PD-1, but are fully competent in producing cytokines and killing virus-infected target cells. To better understand the role of the PD-1 signaling pathway in CD8+ T cell cytotoxicity during acute viral infections we analyzed the expression of the ligand on retrovirus-infected cells targeted by CTLs. We observed increased levels of PD-L1 expression after infection of cells with the murine Friend retrovirus (FV) or with HIV. In FV infected mice, virus-specific CTLs efficiently eliminated infected target cells that expressed low levels of PD-L1 or that were deficient for PD-L1 but the population of PD-L1high cells escaped elimination and formed a reservoir for chronic FV replication. Infected cells with high PD-L1 expression mediated a negative feedback on CD8+ T cells and inhibited their expansion and cytotoxic functions. These findings provide evidence for a novel immune escape mechanism during acute retroviral infection based on PD-L1 expression levels on virus infected target cells.

Exploiting position effects and the gypsy retrovirus insulator to engineer precisely expressed transgenes.

PubMed

Markstein, Michele; Pitsouli, Chrysoula; Villalta, Christians; Celniker, Susan E; Perrimon, Norbert

2008-04-01

A major obstacle to creating precisely expressed transgenes lies in the epigenetic effects of the host chromatin that surrounds them. Here we present a strategy to overcome this problem, employing a Gal4-inducible luciferase assay to systematically quantify position effects of host chromatin and the ability of insulators to counteract these effects at phiC31 integration loci randomly distributed throughout the Drosophila genome. We identify loci that can be exploited to deliver precise doses of transgene expression to specific tissues. Moreover, we uncover a previously unrecognized property of the gypsy retrovirus insulator to boost gene expression to levels severalfold greater than at most or possibly all un-insulated loci, in every tissue tested. These findings provide the first opportunity to create a battery of transgenes that can be reliably expressed at high levels in virtually any tissue by integration at a single locus, and conversely, to engineer a controlled phenotypic allelic series by exploiting several loci. The generality of our approach makes it adaptable to other model systems to identify and modify loci for optimal transgene expression.

Significant differences in genotoxicity induced by retrovirus integration in human T cells and induced pluripotent stem cells.

PubMed

Zheng, Weiyan; Wang, Yingjia; Chang, Tammy; Huang, He; Yee, Jiing-Kuan

2013-04-25

Retrovirus is frequently used in the genetic modification of mammalian cells and the establishment of induced pluripotent stem cells (iPSCs) via cell reprogramming. Vector-induced genotoxicity could induce profound effect on the physiology and function of these stem cells and their differentiated progeny. We analyzed retrovirus-induced genotoxicity in somatic cell Jurkat and two iPSC lines. In Jurkat cells, retrovirus frequently activated host gene expression and gene activation was not dependent on the distance between the integration site and the transcription start site of the host gene. In contrast, retrovirus frequently down-regulated host gene expression in iPSCs, possibly due to the action of chromatin silencing that spreads from the provirus to the nearby host gene promoter. Our data raises the issue that some of the phenotypic variability observed among iPSC clones derived from the same parental cell line may be caused by retrovirus-induced gene expression changes rather than by the reprogramming process itself. It also underscores the importance of characterizing retrovirus integration and carrying out risk assessment of iPSCs before they can be applied in basic research and clinics. Copyright © 2013 Elsevier B.V. All rights reserved.

Linking human beta retrovirus infection with primary biliary cirrhosis.

PubMed

Mason, A L; Zhang, G

2010-01-01

Several environmental agents have been linked with primary biliary cirrhosis (PBC) that include bacteria, xenobiotics and viruses. A human beta retrovirus (HBRV) related to mouse mammary tumor virus has been cloned and characterized from patients with PBC. This agent can be detected in the majority of patients' perihepatic lymph nodes by immunochemistry and RT-PCR. The HBRV has recently been isolated in culture and integration sites have been identified in the genome of patients to provide convincing evidence of beta retrovirus infection in patients. Three lines of evidence support a role for the virus in PBC. First, the beta retrovirus is linked with aberrant expression of mitochondrial protein(s) on the biliary epithelium cell (BEC) surface, a disease specific phenotype. Second, the related agent, mouse mammary tumor virus has been linked with autoimmune biliary disease in the NOD.c3c4 mouse model for PBC. In this mouse model, the virus is localized to diseased biliary epithelium that also display aberrant expression of the mitochondrial autoantigens. In translational studies, both patients with PBC and NOD.c3c4 mice demonstrate significant improvement in biliary disease with combination antiviral therapy. An overview of the biological relevance of the beta retrovirus infection in PBC will be discussed in this review. Copyright 2010 Elsevier Masson SAS. All rights reserved.

Expression and regulation of human endogenous retrovirus W elements.

PubMed

Li, Fang; Karlsson, Håkan

2016-01-01

Human endogenous retroviruses (HERV) comprise 8% of the human genome and can be classified into at least 31 families. A typical HERV provirus consists of internal gag, pol and env genes, flanked by two long terminal repeats (LTRs). No single provirus is capable of engendering infectious particles. HERV are by nature repetitive and have with few notable exceptions lost their protein-coding capacity. Therefore, HERV have consistently been excluded from array-based expression studies and hence little is known of their expression, regulation, and potential functional significance. An increasing number of studies have, however, observed expression of the W family of HERV in various human tissues and cells, predominantly in placenta. HERV-W LTRs act as promoters in directing transcription of HERV-W members, contribute to their tissue-specific and highly diversified expression pattern. Furthermore, leaky transcription originating from adjacent genes plays a role in the transcription initiation of HERV-W psudoelements. It has been reported that HERV-W elements, including ERVWE1 (the so far only known HERV-W locus harboring a gene (env) functionally adopted by the human host to critically participate in placenta biogenesis), can become transactivated in a range of human non-placental cell-lines during exogenous virus infections. Aberrant expression of HERV-W has been associated with human diseases, such as cancer, multiple sclerosis, and schizophrenia. Based on published reports, transcriptional activities of HERV-W appear to be influenced by several mechanisms; binding of transcription factors to LTR promoters and enhancers outside of LTRs, genetic variation and alteration in DNA methylation and histone modification. Emerging mechanistic studies support the notion that HERV-W represents a potential marker or mediator of environmental exposures (e.g., virus infection) in the development of chronic complex diseases. © 2016 APMIS. Published by John Wiley & Sons Ltd.

Retroviruses: Gaining an Understanding.

ERIC Educational Resources Information Center

DiSpezio, Michael A.

1990-01-01

Contrasted are DNA viruses, RNA viruses, and RNA retroviruses. The structure, genome, and replication of retroviruses are discussed. The discovery, structure, and action of the HIV virus are described. A list of 17 references is included. (CW)

Endogenous Retroviruses in the Genomics Era.

PubMed

Johnson, Welkin E

2015-11-01

Endogenous retroviruses comprise millions of discrete genetic loci distributed within the genomes of extant vertebrates. These sequences, which are clearly related to exogenous retroviruses, represent retroviral infections of the deep past, and their abundance suggests that retroviruses were a near-constant presence throughout the evolutionary history of modern vertebrates. Endogenous retroviruses contribute in myriad ways to the evolution of host genomes, as mutagens and as sources of genetic novelty (both coding and regulatory) to be acted upon by the twin engines of random genetic drift and natural selection. Importantly, the richness and complexity of endogenous retrovirus data can be used to understand how viruses spread and adapt on evolutionary timescales by combining population genetics and evolutionary theory with a detailed understanding of retrovirus biology (gleaned from the study of extant retroviruses). In addition to revealing the impact of viruses on organismal evolution, such studies can help us better understand, by looking back in time, how life-history traits, as well as ecological and geological events, influence the movement of viruses within and between populations.

Single-Step Conversion of Cells to Retrovirus Vector Producers with Herpes Simplex Virus–Epstein-Barr Virus Hybrid Amplicons

PubMed Central

Sena-Esteves, Miguel; Saeki, Yoshinaga; Camp, Sara M.; Chiocca, E. Antonio; Breakefield, Xandra O.

1999-01-01

We report here on the development and characterization of a novel herpes simplex virus type 1 (HSV-1) amplicon-based vector system which takes advantage of the host range and retention properties of HSV–Epstein-Barr virus (EBV) hybrid amplicons to efficiently convert cells to retrovirus vector producer cells after single-step transduction. The retrovirus genes gag-pol and env (GPE) and retroviral vector sequences were modified to minimize sequence overlap and cloned into an HSV-EBV hybrid amplicon. Retrovirus expression cassettes were used to generate the HSV-EBV-retrovirus hybrid vectors, HERE and HERA, which code for the ecotropic and the amphotropic envelopes, respectively. Retrovirus vector sequences encoding lacZ were cloned downstream from the GPE expression unit. Transfection of 293T/17 cells with amplicon plasmids yielded retrovirus titers between 106 and 107 transducing units/ml, while infection of the same cells with amplicon vectors generated maximum titers 1 order of magnitude lower. Retrovirus titers were dependent on the extent of transduction by amplicon vectors for the same cell line, but different cell lines displayed varying capacities to produce retrovirus vectors even at the same transduction efficiencies. Infection of human and dog primary gliomas with this system resulted in the production of retrovirus vectors for more than 1 week and the long-term retention and increase in transgene activity over time in these cell populations. Although the efficiency of this system still has to be determined in vivo, many applications are foreseeable for this approach to gene delivery. PMID:10559361

Endogenous Retrovirus ev21 Dose Not Recombine with ALV-J and Induces the Expression of ISGs in the Host.

PubMed

Feng, Min; Tan, Yan; Dai, Manman; Li, Yuanfang; Xie, Tingting; Li, Hongmei; Shi, Meiqing; Zhang, Xiquan

2016-01-01

Avian leukosis virus subgroup J (ALV-J) infection can cause tumors and immunosuppression. Endogenous viruses integrate into host genomes and can recombine with exogenous avian leukosis virus (ALV). In this study, we analyzed the interaction of endogenous retrovirus 21 ( ev21 ) with the ALV-J in late-feathering Chinese yellow chicken. Two ALV-J strains M180 and K243 were isolated from late-feathering and fast-feathering Chinese yellow chicken flocks, respectively. The env gene of the two strains showed 94.2-94.8% nucleotide identity with reference ALV-J strains. Compared with the env gene and the LTR of ev21 and M180, the nucleotide identity of LTR was 69.7% and env gene was 58.4%, respectively, especially the amino acid identity of env gene as low as 14.2%. Phylogenetic analysis of the nucleotide sequence of the env gene and the 3'LTR showed that M180 was closely related to ALV-J, and was located in a distinct group with ev21 in the phylogenetic tree. Using co-immunoprecipitation (co-IP), we next demonstrate that the envelope protein of ev21 does not interact with the M180 envelope protein. We further show that the envelope protein of ev21 cannot activate ALV-J LTR promoter activity using luciferase-reporter assays. qPCR and western blot analysis revealed that envelope protein of endogenous ev21 can facilitate the expression of PKR at 6h post ALV-J infection (hpi) and facilitate the expression of ISG12 and CH25H at 24 hpi. However, the expression of the env gene of M180 strain was not significantly at 6 and 24 hpi. We conclude that there is no evidence of recombination between endogenous retrovirus ev21 and ALV-J strain M180 in late-feathering Chinese yellow chicken, and envelope protein of ev21 can affect the expression of host ISGs, but appears not to influence the replication of ALV-J strain M180. This is the first report of interaction among the endogenous retrovirus ev21, ALV-J and the late-feathering chicken.

Oncogenes in retroviruses and cells

NASA Astrophysics Data System (ADS)

Kurth, Reinhard

1983-09-01

Oncogenes are genes that cause cancer. Retroviruses contain oncogenes and cause cancer in animals and, perhaps, in man. The viruses have appropriated their oncogenes from normal cellular DNA by genetic recombination. Correspondingly, uninfected vertebrate cells contain a family of evolutionary conserved cellular oncogenes. Retrovirus infection, introducing additional viral oncogenes into the cells, as well as carcinogen-mediated activation of cellular oncogenes may both lead to increased synthesis of oncogene encoded transforming proteins which convert normal cells to tumor cells. Unique retroviruses of human origin have recently been identified. They may, on occasion, directly cause tumors in man. However, the general significance of retroviruses may better be illustrated by their remarkable genetic composition which allows them to promote tumor growth by a variety of genetic mechanisms.

A natural allele of Nxf1/TAP supresses retrovirus insertional mutations

PubMed Central

Floyd, Jennifer A.; Gold, David A.; Concepcion, Dorothy; Poon, Tiffany H.; Wang, Xiaobo; Keithley, Elizabeth; Chen, Dan; Ward, Erica J.; Chinn, Steven B.; Friedman, Rick A.; Yu, Hon-Tsen; Moriwaki, Kazuo; Shiroishi, Toshihiko; Hamilton, Bruce A.

2009-01-01

Endogenous retroviruses have shaped the evolution of mammalian genomes. Host genes that control the effects of retrovirus insertions are therefore of great interest. The Modifier-of-vibrator-1 locus controls level of correctly processed mRNA from genes mutated by endogenous retrovirus insertions into introns, including the pitpnvb tremor mutation and the Eya1BOR model of human branchiootorenal syndrome. Positional complementation cloning identifies Mvb1 as the nuclear export factor Nxf1, providing an unexpected link between mRNA export receptor and pre-mRNA processing. Population structure of the suppressing allele in wild M. m. castaneus suggests selective advantage. A congenic Mvb1CAST allele is a useful tool for modifying gene expression from existing mutations and could be used to manipulate engineered mutations containing retroviral elements. PMID:14517553

Pinpointing retrovirus entry sites in cells expressing alternatively spliced receptor isoforms by single virus imaging.

PubMed

Padilla-Parra, Sergi; Marin, Mariana; Kondo, Naoyuki; Melikyan, Gregory B

2014-06-16

The majority of viruses enter host cells via endocytosis. Current knowledge of viral entry pathways is largely based upon infectivity measurements following genetic and/or pharmacological interventions that disrupt vesicular trafficking and maturation. Imaging of single virus entry in living cells provides a powerful means to delineate viral trafficking pathways and entry sites under physiological conditions. Here, we visualized single avian retrovirus co-trafficking with markers for early (Rab5) and late (Rab7) endosomes, acidification of endosomal lumen and the resulting viral fusion measured by the viral content release into the cytoplasm. Virus-carrying vesicles either merged with the existing Rab5-positive early endosomes or slowly accumulated Rab5. The Rab5 recruitment to virus-carrying endosomes correlated with acidification of their lumen. Viral fusion occurred either in early (Rab5-positive) or intermediate (Rab5- and Rab7-positive) compartments. Interestingly, different isoforms of the cognate receptor directed virus entry from distinct endosomes. In cells expressing the transmembrane receptor, viruses preferentially entered and fused with slowly maturing early endosomes prior to accumulation of Rab7. By comparison, in cells expressing the GPI-anchored receptor, viruses entered both slowly and quickly maturing endosomes and fused with early (Rab5-positive) and intermediate (Rab5- and Rab7-positive) compartments. Since the rate of low pH-triggered fusion was independent of the receptor isoform, we concluded that the sites of virus entry are determined by the kinetic competition between endosome maturation and viral fusion. Our findings demonstrate the ability of this retrovirus to enter cells via alternative endocytic pathways and establish infection by releasing its content from distinct endosomal compartments.

Anti-CHMP5 single chain variable fragment antibody retrovirus infection induces programmed cell death of AML leukemic cells in vitro.

PubMed

Wang, Hai-rong; Xiao, Zhen-yu; Chen, Miao; Wang, Fei-long; Liu, Jia; Zhong, Hua; Zhong, Ji-hua; Ou-Yang, Ren-rong; Shen, Yan-lin; Pan, Shu-ming

2012-06-01

Over-expressed CHMP5 was found to act as oncogene that probably participated in leukemogenesis. In this study, we constructed the CHMP5 single chain variable fragment antibody (CHMP5-scFv) retrovirus and studied the changes of programmed cell death (PCD) of AML leukemic cells after infection by the retrovirus. The anti-CHMP5 KC14 hybridoma cell line was constructed to generate monoclonal antibody of CHMP5. The protein expression of CHMP5 was studied using immunofluorescence analysis. pMIG-CHMP5 scFv antibody expressible retroviral vector was constructed to prepare CHMP5-scFv retrovirus. AML leukemic U937 cells were infected with the retrovirus, and programmed cell death was studied using confocal microscope, FCM and Western blot. We obtained a monoclonal antibody of CHMP5, and found the expression of CHMP5 was up-regulated in the leukemic cells. After U937 cells were infected with CHMP5-scFv retrovirus, CHMP5 protein was neutralized. Moreover, the infection resulted in a significant increase in apoptosis and necrosis of U937 cells. In U937 cells infected with CHMP5-scFv retrovirus, apoptosis-inducing factor (AIF)-mediated caspase-independent necrotic PCD was activated, but autophagic programmed cell death was not observed. Neither the intrinsic nor extrinsic apoptotic PCD pathway was activated. The granzyme B/perforin-mediated caspase-dependent apoptotic PCD pathway was not activated. CHMP5-scFv retrovirus can neutralize the abnormally high levels of the CHMP5 protein in the cytosol of AML leukemic U937 cells, thereby inducing the programmed cell death of the leukemic cells via AIF-mediated caspase-independent necrosis and apoptosis.

Insufficient natural killer cell responses against retroviruses: how to improve NK cell killing of retrovirus-infected cells.

PubMed

Littwitz-Salomon, Elisabeth; Dittmer, Ulf; Sutter, Kathrin

2016-11-08

Natural killer (NK) cells belong to the innate immune system and protect against cancers and a variety of viruses including retroviruses by killing transformed or infected cells. They express activating and inhibitory receptors on their cell surface and often become activated after recognizing virus-infected cells. They have diverse antiviral effector functions like the release of cytotoxic granules, cytokine production and antibody dependent cellular cytotoxicity. The importance of NK cell activity in retroviral infections became evident due to the discovery of several viral strategies to escape recognition and elimination by NK cells. Mutational sequence polymorphisms as well as modulation of surface receptors and their ligands are mechanisms of the human immunodeficiency virus-1 to evade NK cell-mediated immune pressure. In Friend retrovirus infected mice the virus can manipulate molecular or cellular immune factors that in turn suppress the NK cell response. In this model NK cells lack cytokines for optimal activation and can be functionally suppressed by regulatory T cells. However, these inhibitory pathways can be overcome therapeutically to achieve full activation of NK cell responses and ultimately control dissemination of retroviral infection. One effective approach is to modulate the crosstalk between NK cells and dendritic cells, which produce NK cell-stimulating cytokines like type I interferons (IFN), IL-12, IL-15, and IL-18 upon retrovirus sensing or infection. Therapeutic administration of IFNα directly increases NK cell killing of retrovirus-infected cells. In addition, IL-2/anti-IL-2 complexes that direct IL-2 to NK cells have been shown to significantly improve control of retroviral infection by NK cells in vivo. In this review, we describe novel approaches to improve NK cell effector functions in retroviral infections. Immunotherapies that target NK cells of patients suffering from viral infections might be a promising treatment option for the

A novel recombinant retrovirus in the genomes of modern birds combines features of avian and mammalian retroviruses.

PubMed

Henzy, Jamie E; Gifford, Robert J; Johnson, Welkin E; Coffin, John M

2014-03-01

Endogenous retroviruses (ERVs) represent ancestral sequences of modern retroviruses or their extinct relatives. The majority of ERVs cluster alongside exogenous retroviruses into two main groups based on phylogenetic analyses of the reverse transcriptase (RT) enzyme. Class I includes gammaretroviruses, and class II includes lentiviruses and alpha-, beta-, and deltaretroviruses. However, analyses of the transmembrane subunit (TM) of the envelope glycoprotein (env) gene result in a different topology for some retroviruses, suggesting recombination events in which heterologous env sequences have been acquired. We previously demonstrated that the TM sequences of five of the six genera of orthoretroviruses can be divided into three types, each of which infects a distinct set of vertebrate classes. Moreover, these classes do not always overlap the host range of the associated RT classes. Thus, recombination resulting in acquisition of a heterologous env gene could in theory facilitate cross-species transmissions across vertebrate classes, for example, from mammals to reptiles. Here we characterized a family of class II avian ERVs, "TgERV-F," that acquired a mammalian gammaretroviral env sequence. Although TgERV-F clusters near a sister clade to alpharetroviruses, its genome also has some features of betaretroviruses. We offer evidence that this unusual recombinant has circulated among several avian orders and may still have infectious members. In addition to documenting the infection of a nongalliform avian species by a mammalian retrovirus, TgERV-F also underscores the importance of env sequences in reconstructing phylogenies and supports a possible role for env swapping in allowing cross-species transmissions across wide taxonomic distances. Retroviruses can sometimes acquire an envelope gene (env) from a distantly related retrovirus. Since env is a key determinant of host range, such an event affects the host range of the recombinant virus and can lead to the creation

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

PubMed Central

Palmarini, Massimo; Hallwirth, Claus; York, Denis; Murgia, Claudio; de Oliveira, Tulio; Spencer, Thomas; Fan, Hung

2000-01-01

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 enJRSVs) 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 enJSRVs 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 JSRV21. 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 enJSRVs 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. PMID:10933716

Recent Amplification of the Kangaroo Endogenous Retrovirus, KERV, Limited to the Centromere▿

PubMed Central

Ferreri, Gianni C.; Brown, Judith D.; Obergfell, Craig; Jue, Nathaniel; Finn, Caitlin E.; O'Neill, Michael J.; O'Neill, Rachel J.

2011-01-01

Mammalian retrotransposons, transposable elements that are processed through an RNA intermediate, are categorized as short interspersed elements (SINEs), long interspersed elements (LINEs), and long terminal repeat (LTR) retroelements, which include endogenous retroviruses. The ability of transposable elements to autonomously amplify led to their initial characterization as selfish or junk DNA; however, it is now known that they may acquire specific cellular functions in a genome and are implicated in host defense mechanisms as well as in genome evolution. Interactions between classes of transposable elements may exert a markedly different and potentially more significant effect on a genome than interactions between members of a single class of transposable elements. We examined the genomic structure and evolution of the kangaroo endogenous retrovirus (KERV) in the marsupial genus Macropus. The complete proviral structure of the kangaroo endogenous retrovirus, phylogenetic relationship among relative retroviruses, and expression of this virus in both Macropus rufogriseus and M. eugenii are presented for the first time. In addition, we show the relative copy number and distribution of the kangaroo endogenous retrovirus in the Macropus genus. Our data indicate that amplification of the kangaroo endogenous retrovirus occurred in a lineage-specific fashion, is restricted to the centromeres, and is not correlated with LINE depletion. Finally, analysis of KERV long terminal repeat sequences using massively parallel sequencing indicates that the recent amplification in M. rufogriseus is likely due to duplications and concerted evolution rather than a high number of independent insertion events. PMID:21389136

Single-Particle Discrimination of Retroviruses from Extracellular Vesicles by Nanoscale Flow Cytometry.

PubMed

Tang, Vera A; Renner, Tyler M; Fritzsche, Anna K; Burger, Dylan; Langlois, Marc-André

2017-12-19

Retroviruses and small EVs overlap in size, buoyant densities, refractive indices and share many cell-derived surface markers making them virtually indistinguishable by standard biochemical methods. This poses a significant challenge when purifying retroviruses for downstream analyses or for phenotypic characterization studies of markers on individual virions given that EVs are a major contaminant of retroviral preparations. Nanoscale flow cytometry (NFC), also called flow virometry, is an adaptation of flow cytometry technology for the analysis of individual nanoparticles such as extracellular vesicles (EVs) and retroviruses. In this study we systematically optimized NFC parameters for the detection of retroviral particles in the range of 115-130 nm, including viral production, sample labeling, laser power and voltage settings. By using the retroviral envelope glycoprotein as a selection marker, and evaluating a number of fluorescent dyes and labeling methods, we demonstrate that it is possible to confidently distinguish retroviruses from small EVs by NFC. Our findings make it now possible to individually phenotype genetically modified retroviral particles that express a fluorescent envelope glycoprotein without removing EV contaminants from the sample.

Neuropeptide Y Negatively Influences Monocyte Recruitment to the Central Nervous System during Retrovirus Infection.

PubMed

Woods, Tyson A; Du, Min; Carmody, Aaron; Peterson, Karin E

2015-12-30

Monocyte infiltration into the CNS is a hallmark of several viral infections of the central nervous system (CNS), including retrovirus infection. Understanding the factors that mediate monocyte migration in the CNS is essential for the development of therapeutics that can alter the disease process. In the current study, we found that neuropeptide Y (NPY) suppressed monocyte recruitment to the CNS in a mouse model of polytropic retrovirus infection. NPY(-/-) mice had increased incidence and kinetics of retrovirus-induced neurological disease, which correlated with a significant increase in monocytes in the CNS compared to wild-type mice. Both Ly6C(hi) inflammatory and Ly6C(lo) alternatively activated monocytes were increased in the CNS of NPY(-/-) mice following virus infection, suggesting that NPY suppresses the infiltration of both cell types. Ex vivo analysis of myeloid cells from brain tissue demonstrated that infiltrating monocytes expressed high levels of the NPY receptor Y2R. Correlating with the expression of Y2R on monocytes, treatment of NPY(-/-) mice with a truncated, Y2R-specific NPY peptide suppressed the incidence of retrovirus-induced neurological disease. These data demonstrate a clear role for NPY as a negative regulator of monocyte recruitment into the CNS and provide a new mechanism for suppression of retrovirus-induced neurological disease. Monocyte recruitment to the brain is associated with multiple neurological diseases. However, the factors that influence the recruitment of these cells to the brain are still not well understood. In the current study, we found that neuropeptide Y, a protein produced by neurons, affected monocyte recruitment to the brain during retrovirus infection. We show that mice deficient in NPY have increased influx of monocytes into the brain and that this increase in monocytes correlates with neurological-disease development. These studies provide a mechanism by which the nervous system, through the production of NPY

Negative regulation of retrovirus expression in embryonal carcinoma cells mediated by an intragenic domain.

PubMed

Loh, T P; Sievert, L L; Scott, R W

1988-11-01

An intragenic region spanning the tRNA primer binding site of a Moloney murine leukemia virus recombinant retrovirus was found to restrict expression specifically in embryonal carcinoma (EC) cells. When the inhibitory domain was present, the levels of steady-state RNA synthesized from integrated recombinant templates in stable cotransformation assays were reduced 20-fold in EC cells but not in C2 myoblast cells. Transient-cotransfection assays showed that repression of a template containing the EC-specific inhibitory component was relieved by an excess of specific competitor DNA. In addition, repression mediated by the inhibitory component was orientation independent. This evidence demonstrates the presence of a saturable, trans-acting negative regulatory factor(s) in EC cells and suggests that the interaction of the factor(s) with the intragenic inhibitory component occurs at the DNA level.

Retroviruses.

ERIC Educational Resources Information Center

Varmus, Harold

1988-01-01

Discusses the growth, development, and unusual parasitic nature of the retrovirus community. Reviews these infectious cancer-causing agents as models for the study of fundamental biological problems, tools for genetic manipulations, and problems posed by their pathogenic potential in humans and animal hosts where they cause diseases such as…

HIV-1 Infection of Primary CD4+ T Cells Regulates the Expression of Specific Human Endogenous Retrovirus HERV-K (HML-2) Elements.

PubMed

Young, George R; Terry, Sandra N; Manganaro, Lara; Cuesta-Dominguez, Alvaro; Deikus, Gintaras; Bernal-Rubio, Dabeiba; Campisi, Laura; Fernandez-Sesma, Ana; Sebra, Robert; Simon, Viviana; Mulder, Lubbertus C F

2018-01-01

Endogenous retroviruses (ERVs) occupy extensive regions of the human genome. Although many of these retroviral elements have lost their ability to replicate, those whose insertion took place more recently, such as the HML-2 group of HERV-K elements, still retain intact open reading frames and the capacity to produce certain viral RNA and/or proteins. Transcription of these ERVs is, however, tightly regulated by dedicated epigenetic control mechanisms. Nonetheless, it has been reported that some pathological states, such as viral infections and certain cancers, coincide with ERV expression, suggesting that transcriptional reawakening is possible. HML-2 elements are reportedly induced during HIV-1 infection, but the conserved nature of these elements has, until recently, rendered their expression profiling problematic. Here, we provide comprehensive HERV-K HML-2 expression profiles specific for productively HIV-1-infected primary human CD4 + T cells. We combined enrichment of HIV-1 infected cells using a reporter virus expressing a surface reporter for gentle and efficient purification with long-read single-molecule real-time sequencing. We show that three HML-2 proviruses-6q25.1, 8q24.3, and 19q13.42-are upregulated on average between 3- and 5-fold in HIV-1-infected CD4 + T cells. One provirus, HML-2 12q24.33, in contrast, was repressed in the presence of active HIV replication. In conclusion, this report identifies the HERV-K HML-2 loci whose expression profiles differ upon HIV-1 infection in primary human CD4 + T cells. These data will help pave the way for further studies on the influence of endogenous retroviruses on HIV-1 replication. IMPORTANCE Endogenous retroviruses inhabit big portions of our genome. Moreover, although they are mainly inert, some of the evolutionarily younger members maintain the ability to express both RNA and proteins. We have developed an approach using long-read single-molecule real-time (SMRT) sequencing that produces long reads that

Morphology and ultrastructure of retrovirus particles

PubMed Central

Zhang, Wei; Cao, Sheng; Martin, Jessica L.; Mueller, Joachim D.; Mansky, Louis M.

2015-01-01

Retrovirus morphogenesis entails assembly of Gag proteins and the viral genome on the host plasma membrane, acquisition of the viral membrane and envelope proteins through budding, and formation of the core through the maturation process. Although in both immature and mature retroviruses, Gag and capsid proteins are organized as paracrystalline structures, the curvatures of these protein arrays are evidently not uniform within one or among all virus particles. The heterogeneity of retroviruses poses significant challenges to studying the protein contacts within the Gag and capsid lattices. This review focuses on current understanding of the molecular organization of retroviruses derived from the sub-nanometer structures of immature virus particles, helical capsid protein assemblies and soluble envelope protein complexes. These studies provide insight into the molecular elements that maintain the stability, flexibility and infectivity of virus particles. Also reviewed are morphological studies of retrovirus budding, maturation, infection and cell-cell transmission, which inform the structural transformation of the viruses and the cells during infection and viral transmission, and lead to better understanding of the interplay between the functioning viral proteins and the host cell. PMID:26448965

Microbiological characterization of a newly established pig breed, Aachen Minipigs.

PubMed

Plotzki, Elena; Heinrichs, Gerd; Kubícková, Barbara; Ulrich, Rainer G; Denner, Joachim

2016-03-01

To alleviate the shortage of human donor organs or tissues for the treatment of organ and tissue failure including diabetes, pigs are considered suitable donor animals. As organs from conventional pigs are usually too large, those from minipigs may be better suited. We recently characterized the Göttingen Minipigs, a breed well characterized concerning the presence of zoonotic microorganisms and found hepatitis E virus (HEV) and porcine cytomegalovirus (PCMV) in some animals. Here, we characterize another minipig, the Aachen Minipig (AaMP), a pig breed recently established close to the town Aachen in Germany. The animals were tested for the prevalence and expression of porcine endogenous retroviruses (PERVs) and the presence of some selected microorganisms, among them HEV, PCMV, and porcine lymphotropic herpesviruses (PLHVs) using highly sensitive and specific PCR and RT-PCR methods. In addition, we screened for antibodies against HEV and PLHV. PERV-A, PERV-B, and PERV-C sequences were found in the genome of all Aachen Minipigs. HEV RNA was found by real-time RT-PCR in most, and DNA of PCMV, PLHV-2, and PLHV-3 was found by PCR in some animals. The animals were free of eight other microorganisms tested, but some were seropositive for porcine circovirus 2 (PCV2), porcine reproductive and respiratory syndrome virus (PRRSV), and porcine epidemic diarrhea virus (PEDV). Based on medical examinations by veterinarians, the AaMP are in a good health status and seem to harbor only few microorganisms. To improve their status for use as donor pigs in xenotransplantation, the viruses detected might be eliminated by selection of negative animals, Cesarean section, and vaccination. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Changes in expression of cellular oncogenes and endogenous retrovirus-like sequences during hepatocarcinogenesis induced by a peroxisome proliferator.

PubMed Central

Hsieh, L. L.; Shinozuka, H.; Weinstein, I. B.

1991-01-01

Previous studies have demonstrated that BR-931, a hepatic peroxisome proliferator, can induce liver tumours in mice and rats. Since alterations in gene expression may play a critical role in multistage hepatocarcinogenesis, the present studies examined the expression of the c-myc, c-H-ras, epidermal growth factor (EGF) receptor and ODC (ornithine decarboxylase) genes, as well as endogenous retrovirus-like sequences, in F344 rat liver during the first 8 weeks of feeding a 0.16% Br931 diet and in liver tumours induced by chronic feeding of this diet. Northern blot analysis of poly A + liver RNA samples showed an increase in the level of RNAs homologous to rat leukaemia virus (RaLV) but no significant change in the level of 30S-retrovirus related RNAs in the liver RNA samples obtained from rats during the first 8 weeks of feeding the diet containing BR931. An increase in the levels of c-myc, c-H-ras and ODC transcripts was also seen in the liver RNA samples from the treated rats. Of particular interest was a decrease in the abundance of EGF receptor transcripts in the liver RNA samples from rats fed the BR931 diet. Increased levels of RaLV, c-myc, and ODC RNAs were also seen in the tumours induced by BR931, but this was not the case for 30S and c-H-ras. The liver tumour samples also showed a decrease in EGF receptor RNA. These changes in cellular levels of specific RNAs resemble, in several respect, those we previously described in rodent liver during regeneration and tumour promotion, and also those seen in rodent hepatomas induced by other agents. Therefore, they may reflect a common profile of gene expression relevant to liver proliferation and carcinogenesis. Images Figure 1 Figure 2 PMID:1931600

Pan-vertebrate comparative genomics unmasks retrovirus macroevolution.

PubMed

Hayward, Alexander; Cornwallis, Charlie K; Jern, Patric

2015-01-13

Although extensive research has demonstrated host-retrovirus microevolutionary dynamics, it has been difficult to gain a deeper understanding of the macroevolutionary patterns of host-retrovirus interactions. Here we use recent technological advances to infer broad patterns in retroviral diversity, evolution, and host-virus relationships by using a large-scale phylogenomic approach using endogenous retroviruses (ERVs). Retroviruses insert a proviral DNA copy into the host cell genome to produce new viruses. ERVs are provirus insertions in germline cells that are inherited down the host lineage and consequently present a record of past host-viral associations. By mining ERVs from 65 host genomes sampled across vertebrate diversity, we uncover a great diversity of ERVs, indicating that retroviral sequences are much more prevalent and widespread across vertebrates than previously appreciated. The majority of ERV clades that we recover do not contain known retroviruses, implying either that retroviral lineages are highly transient over evolutionary time or that a considerable number of retroviruses remain to be identified. By characterizing the distribution of ERVs, we show that no major vertebrate lineage has escaped retroviral activity and that retroviruses are extreme host generalists, having an unprecedented ability for rampant host switching among distantly related vertebrates. In addition, we examine whether the distribution of ERVs can be explained by host factors predicted to influence viral transmission and find that internal fertilization has a pronounced effect on retroviral colonization of host genomes. By capturing the mode and pattern of retroviral evolution and contrasting ERV diversity with known retroviral diversity, our study provides a cohesive framework to understand host-virus coevolution better.

Friends-enemies: endogenous retroviruses are major transcriptional regulators of human DNA

NASA Astrophysics Data System (ADS)

Buzdin, Anton A.; Prassolov, Vladimir; Garazha, Andrew V.

2017-06-01

Endogenous retroviruses are mobile genetic elements hardly distinguishable from infectious, or “exogenous”, retroviruses at the time of insertion in the host DNA. Human endogenous retroviruses (HERVs) are not rare. They gave rise to multiple families of closely related mobile elements that occupy 8% of the human genome. Together, they shape genomic regulatory landscape by providing at least 320,000 human transcription factor binding sites (TFBS) located on 110,000 individual HERV elements. The HERVs host as many as 155,000 mapped DNaseI hypersensitivity sites, which denote loci active in the regulation of gene expression or chromatin structure. The contemporary view of the HERVs evolutionary dynamics suggests that at the early stages after insertion, the HERV is treated by the host cells as a foreign genetic element, and is likely to be suppressed by the targeted methylation and mutations. However, at the later stages, when significant number of mutations has been already accumulated and when the retroviral genes are broken, the regulatory potential of a HERV may be released and recruited to modify the genomic balance of transcription factor binding sites. This process goes together with further accumulation and selection of mutations, which reshape the regulatory landscape of the human DNA. However, developmental reprogramming, stress or pathological conditions like cancer, inflammation and infectious diseases, can remove the blocks limiting expression and HERV-mediated host gene regulation. This, in turn, can dramatically alter the gene expression equilibrium and shift it to a newer state, thus further amplifying instability and exacerbating the stressful situation.

Pantropic retroviruses as a transduction tool for sea urchin embryos

PubMed Central

Core, Amanda B.; Reyna, Arlene E.; Conaway, Evan A.; Bradham, Cynthia A.

2012-01-01

Sea urchins are an important model for experiments at the intersection of development and systems biology, and technical innovations that enhance the utility of this model are of great value. This study explores pantropic retroviruses as a transduction tool for sea urchin embryos, and demonstrates that pantropic retroviruses infect sea urchin embryos with high efficiency and genomically integrate at a copy number of one per cell. We successfully used a self-inactivation strategy to both insert a sea urchin-specific enhancer and disrupt the endogenous viral enhancer. The resulting self-inactivating viruses drive global and persistent gene expression, consistent with genomic integration during the first cell cycle. Together, these data provide substantial proof of principle for transduction technology in sea urchin embryos. PMID:22431628

Pan-vertebrate comparative genomics unmasks retrovirus macroevolution

PubMed Central

Hayward, Alexander; Cornwallis, Charlie K.; Jern, Patric

2015-01-01

Although extensive research has demonstrated host-retrovirus microevolutionary dynamics, it has been difficult to gain a deeper understanding of the macroevolutionary patterns of host–retrovirus interactions. Here we use recent technological advances to infer broad patterns in retroviral diversity, evolution, and host–virus relationships by using a large-scale phylogenomic approach using endogenous retroviruses (ERVs). Retroviruses insert a proviral DNA copy into the host cell genome to produce new viruses. ERVs are provirus insertions in germline cells that are inherited down the host lineage and consequently present a record of past host–viral associations. By mining ERVs from 65 host genomes sampled across vertebrate diversity, we uncover a great diversity of ERVs, indicating that retroviral sequences are much more prevalent and widespread across vertebrates than previously appreciated. The majority of ERV clades that we recover do not contain known retroviruses, implying either that retroviral lineages are highly transient over evolutionary time or that a considerable number of retroviruses remain to be identified. By characterizing the distribution of ERVs, we show that no major vertebrate lineage has escaped retroviral activity and that retroviruses are extreme host generalists, having an unprecedented ability for rampant host switching among distantly related vertebrates. In addition, we examine whether the distribution of ERVs can be explained by host factors predicted to influence viral transmission and find that internal fertilization has a pronounced effect on retroviral colonization of host genomes. By capturing the mode and pattern of retroviral evolution and contrasting ERV diversity with known retroviral diversity, our study provides a cohesive framework to understand host–virus coevolution better. PMID:25535393

[Efficient packaging retrovirus and construction of transgenic chicken technical platform].

PubMed

Man, Chaolai; Zhang, Qing; Chen, Yan; Zhu, Dahai

2007-10-01

Transgenic chicken and oviduct bioreactor are growing to be one of the hotspot of scientific study in the field of biology. The most successful method of producing transgenic chicken is pseudotyped retrovirus vector system, but no one has reported the production of transgenic chicken by retrovirus system recently in our country. In order to accelerate our study in this field, we introduced the relevant technical methods such as packaging retrovirus and vesicular stomatitis virus G glycoprotein (VSV-G) pseudotyped retrovirus, optimizing the conditions of packaging retrovirus, concentrating VSV-G pseudotyped retrovirus, helper virus assays, and microinjection of retrovirus. Furthermore, we successfully conducted in vivo study for detecting the marker gene EGFP of chicken embryo as well as in vitro study for detecting that gene of chicken embryo myoblast (CFM), thus we have provided an applied technical platform for studies of transgenic chicken in the future.

[Establishment of the retrovirus-mediated murine model with MLL-AF9 leukemia].

PubMed

Xu, Si-Miao; Yang, Yang; Zhou, Mi; Zhao, Xue-Jiao; Qin, Yu; Zhang, Pei-Ling; Yuan, Rui-Feng; Zhou, Jian-Feng; Fang, Yong

2013-10-01

This study was purposed to establish a retrovirus-mediated murine model with MLL-AF9 leukemia, so as to provide a basis for further investigation of the pathogenesis and therapeutic strategy of MLL associated leukemia. Murine (CD45.2) primary hematopoietic precursor positively selected for expression of the progenitor marker c-Kit by means of MACS were transduced with a retrovirus carrying MLL-AF9 fusion gene. After cultured in vitro, the transduced cells were injected intravenously through the tail vein into the lethally irradiated mice (CD45.1). PCR, flow cytometry and morphological observation were employed to evaluate the murine leukemia model system. The results showed that MLL-AF9 fusion gene was expressed in the infected cells, and the cells had a dramatically enhanced potential to generate myeloid colonies with primitive and immature morphology. Flow cytometric analysis revealed that the immortalized cells highly expressed myeloid lineage surface markers Gr-1 and Mac-1. Moreover, the expression levels of Hoxa9 and Meis1 mRNA were significantly higher in the MLL-AF9 cells than that in control. The mice transplanted with MLL-AF9 cells displayed typical signs of leukemia within 6-12 weeks. Extensive infiltration leukemic cells was observed in the Wright-Giemsa stained peripheral blood smear and bone marrow, and also in the histology of liver and spleen. Flow cytometric analysis of the bone marrow and spleen cells demonstrated that the CD45.2 populations expressed highly myeloid markers Gr-1 and Mac-1. The leukemic mice died within 12 weeks. It is concluded that the retrovirus-mediated murine model with MLL-AF9 leukemia is successfully established, which can be applied in the subsequent researches.

Tetherin promotes the innate and adaptive cell-mediated immune response against retrovirus infection in vivo.

PubMed

Li, Sam X; Barrett, Bradley S; Heilman, Karl J; Messer, Ronald J; Liberatore, Rachel A; Bieniasz, Paul D; Kassiotis, George; Hasenkrug, Kim J; Santiago, Mario L

2014-07-01

Tetherin/BST-2 is a host restriction factor that could directly inhibit retroviral particle release by tethering nascent virions to the plasma membrane. However, the immunological impact of Tetherin during retrovirus infection remains unknown. We now show that Tetherin influences antiretroviral cell-mediated immune responses. In contrast to the direct antiviral effects of Tetherin, which are dependent on cell surface expression, the immunomodulatory effects are linked to the endocytosis of the molecule. Mice encoding endocytosis-competent C57BL/6 Tetherin exhibited lower viremia and pathology at 7 d postinfection with Friend retrovirus (FV) compared with mice encoding endocytosis-defective NZW/LacJ Tetherin. Notably, antiretroviral protection correlated with stronger NK cell responses. In addition, Friend retrovirus infection levels were significantly lower in wild-type C57BL/6 mice than in Tetherin knockout mice at 2 wk postinfection, and antiretroviral protection correlated with stronger NK cell and virus-specific CD8+ T cell responses. The results demonstrate that Tetherin acts as a modulator of the cell-mediated immune response against retrovirus infection in vivo.

Complex codon usage pattern and compositional features of retroviruses.

PubMed

RoyChoudhury, Sourav; Mukherjee, Debaprasad

2013-01-01

Retroviruses infect a wide range of organisms including humans. Among them, HIV-1, which causes AIDS, has now become a major threat for world health. Some of these viruses are also potential gene transfer vectors. In this study, the patterns of synonymous codon usage in retroviruses have been studied through multivariate statistical methods on ORFs sequences from the available 56 retroviruses. The principal determinant for evolution of the codon usage pattern in retroviruses seemed to be the compositional constraints, while selection for translation of the viral genes plays a secondary role. This was further supported by multivariate analysis on relative synonymous codon usage. Thus, it seems that mutational bias might have dominated role over translational selection in shaping the codon usage of retroviruses. Codon adaptation index was used to identify translationally optimal codons among genes from retroviruses. The comparative analysis of the preferred and optimal codons among different retroviral groups revealed that four codons GAA, AAA, AGA, and GGA were significantly more frequent in most of the retroviral genes inspite of some differences. Cluster analysis also revealed that phylogenetically related groups of retroviruses have probably evolved their codon usage in a concerted manner under the influence of their nucleotide composition.

In between: gypsy in Drosophila melanogaster reveals new insights into endogenous retrovirus evolution.

PubMed

Touret, Franck; Guiguen, François; Greenland, Timothy; Terzian, Christophe

2014-12-09

Retroviruses are RNA viruses that are able to synthesize a DNA copy of their genome and insert it into a chromosome of the host cell. Sequencing of different eukaryote genomes has revealed the presence of many such endogenous retroviral sequences. The mechanisms by which these retroviral sequences have colonized the genome are still unknown, and the endogenous retrovirus gypsy of Drosophila melanogaster is a powerful experimental model for deciphering this process in vivo. Gypsy is expressed in a layer of somatic cells, and then transferred into the oocyte by an unknown mechanism. This critical step is the start of the endogenization process. Moreover gypsy has been shown to have infectious properties, probably due to its envelope gene acquired from a baculovirus. Recently we have also shown that gypsy maternal transmission is reduced in the presence of the endosymbiotic bacterium Wolbachia. These studies demonstrate that gypsy is a unique and powerful model for understanding the endogenization of retroviruses.

In between: Gypsy in Drosophila melanogaster Reveals New Insights into Endogenous Retrovirus Evolution

PubMed Central

Touret, Franck; Guiguen, François; Greenland, Timothy; Terzian, Christophe

2014-01-01

Retroviruses are RNA viruses that are able to synthesize a DNA copy of their genome and insert it into a chromosome of the host cell. Sequencing of different eukaryote genomes has revealed the presence of many such endogenous retroviral sequences. The mechanisms by which these retroviral sequences have colonized the genome are still unknown, and the endogenous retrovirus gypsy of Drosophila melanogaster is a powerful experimental model for deciphering this process in vivo. Gypsy is expressed in a layer of somatic cells, and then transferred into the oocyte by an unknown mechanism. This critical step is the start of the endogenization process. Moreover gypsy has been shown to have infectious properties, probably due to its envelope gene acquired from a baculovirus. Recently we have also shown that gypsy maternal transmission is reduced in the presence of the endosymbiotic bacterium Wolbachia. These studies demonstrate that gypsy is a unique and powerful model for understanding the endogenization of retroviruses. PMID:25502325

Tetherin/BST-2 promotes dendritic cell activation and function during acute retrovirus infection

PubMed Central

Li, Sam X.; Barrett, Bradley S.; Guo, Kejun; Kassiotis, George; Hasenkrug, Kim J.; Dittmer, Ulf; Gibbert, Kathrin; Santiago, Mario L.

2016-01-01

Tetherin/BST-2 is a host restriction factor that inhibits retrovirus release from infected cells in vitro by tethering nascent virions to the plasma membrane. However, contradictory data exists on whether Tetherin inhibits acute retrovirus infection in vivo. Previously, we reported that Tetherin-mediated inhibition of Friend retrovirus (FV) replication at 2 weeks post-infection correlated with stronger natural killer, CD4+ T and CD8+ T cell responses. Here, we further investigated the role of Tetherin in counteracting retrovirus replication in vivo. FV infection levels were similar between wild-type (WT) and Tetherin KO mice at 3 to 7 days post-infection despite removal of a potent restriction factor, Apobec3/Rfv3. However, during this phase of acute infection, Tetherin enhanced myeloid dendritic cell (DC) function. DCs from infected, but not uninfected, WT mice expressed significantly higher MHC class II and the co-stimulatory molecule CD80 compared to Tetherin KO DCs. Tetherin-associated DC activation during acute FV infection correlated with stronger NK cell responses. Furthermore, Tetherin+ DCs from FV-infected mice more strongly stimulated FV-specific CD4+ T cells ex vivo compared to Tetherin KO DCs. The results link the antiretroviral and immunomodulatory activity of Tetherin in vivo to improved DC activation and MHC class II antigen presentation. PMID:26846717

Tetherin/BST-2 promotes dendritic cell activation and function during acute retrovirus infection.

PubMed

Li, Sam X; Barrett, Bradley S; Guo, Kejun; Kassiotis, George; Hasenkrug, Kim J; Dittmer, Ulf; Gibbert, Kathrin; Santiago, Mario L

2016-02-05

Tetherin/BST-2 is a host restriction factor that inhibits retrovirus release from infected cells in vitro by tethering nascent virions to the plasma membrane. However, contradictory data exists on whether Tetherin inhibits acute retrovirus infection in vivo. Previously, we reported that Tetherin-mediated inhibition of Friend retrovirus (FV) replication at 2 weeks post-infection correlated with stronger natural killer, CD4+ T and CD8+ T cell responses. Here, we further investigated the role of Tetherin in counteracting retrovirus replication in vivo. FV infection levels were similar between wild-type (WT) and Tetherin KO mice at 3 to 7 days post-infection despite removal of a potent restriction factor, Apobec3/Rfv3. However, during this phase of acute infection, Tetherin enhanced myeloid dendritic cell (DC) function. DCs from infected, but not uninfected, WT mice expressed significantly higher MHC class II and the co-stimulatory molecule CD80 compared to Tetherin KO DCs. Tetherin-associated DC activation during acute FV infection correlated with stronger NK cell responses. Furthermore, Tetherin+ DCs from FV-infected mice more strongly stimulated FV-specific CD4+ T cells ex vivo compared to Tetherin KO DCs. The results link the antiretroviral and immunomodulatory activity of Tetherin in vivo to improved DC activation and MHC class II antigen presentation.

Retrovirus-based vectors for transient and permanent cell modification.

PubMed

Schott, Juliane W; Hoffmann, Dirk; Schambach, Axel

2015-10-01

Retroviral vectors are commonly employed for long-term transgene expression via integrating vector technology. However, three alternative retrovirus-based platforms are currently available that allow transient cell modification. Gene expression can be mediated from either episomal DNA or RNA templates, or selected proteins can be directly transferred through retroviral nanoparticles. The different technologies are functionally graded with respect to safety, expression magnitude and expression duration. Improvement of the initial technologies, including modification of vector designs, targeted increase in expression strength and duration as well as improved safety characteristics, has allowed maturation of retroviral systems into efficient and promising tools that meet the technological demands of a wide variety of potential application areas. Copyright © 2015 Elsevier Ltd. All rights reserved.

Retroviruses and amyotrophic lateral sclerosis.

PubMed

Alfahad, Tariq; Nath, Avindra

2013-08-01

Amyotrophic lateral sclerosis (ALS) is a progressive, invariably fatal neurologic disorder resulting from upper and lower motor neuron degeneration, which typically develops during the sixth or seventh decade of life, and is diagnosed based on standard clinical criteria. Its underlying cause remains undetermined. The disease may occur with increased frequency within certain families, often in association with specific genomic mutations, while some sporadic cases have been linked to environmental toxins or trauma. Another possibility, first proposed in the 1970s, is that retroviruses play a role in pathogenesis. In this paper, we review the published literature for evidence that ALS is associated either with infection by an exogenous retrovirus or with the expression of human endogenous retroviral (HERV) sequences in cells of the central nervous system. A small percentage of persons infected with the human immunodeficiency virus-1 (HIV-1) or human T cell leukemia virus-1 (HTLV-1) develop ALS-like syndromes. While HTLV-1 associated ALS-like syndrome has several features that may distinguish it from classical ALS, HIV-infected patients may develop neurological manifestations that resemble classical ALS although it occurs at a younger age and they may show a dramatic improvement following the initiation of antiretroviral therapy. However, most patients with probable or definite ALS show no evidence of HIV-1 or HTLV-1 infection. In contrast, recent reports have shown a stronger association with HERV, as analysis of serum samples, and postmortem brain tissue from a number of patients with a classical ALS has revealed significantly increased expression of HERV-K, compared to controls. These findings suggest that endogenous retroviral elements are involved in the pathophysiology of ALS, but there is no evidence that they are the primary cause of the syndrome. Published by Elsevier B.V.

Ancestry of a human endogenous retrovirus family.

PubMed Central

Mariani-Costantini, R; Horn, T M; Callahan, R

1989-01-01

The human endogenous retrovirus type II (HERVII) family of HERV genomes has been found by Southern blot analysis to be characteristic of humans, apes, and Old World monkeys. New World monkeys and prosimians lack HERVII proviral genomes. Cellular DNAs of humans, common chimpanzees, gorillas, and orangutans, but not lesser ape lar gibbons, appear to contain the HERVII-related HLM-2 proviral genome integrated at the same site (HLM-2 maps to human chromosome 1). This suggests that the ancestral HERVII retrovirus(es) entered the genomes of Old World anthropoids by infection after the divergence of New World monkeys (platyrrhines) but before the evolutionary radiation of large hominoids. Images PMID:2507793

Convergent Evolution of Ribonuclease H in LTR Retrotransposons and Retroviruses

PubMed Central

Ustyantsev, Kirill; Novikova, Olga; Blinov, Alexander; Smyshlyaev, Georgy

2015-01-01

Ty3/Gypsy long terminals repeat (LTR) retrotransposons are structurally and phylogenetically close to retroviruses. Two notable structural differences between these groups of genetic elements are 1) the presence in retroviruses of an additional envelope gene, env, which mediates infection, and 2) a specific dual ribonuclease H (RNH) domain encoded by the retroviral pol gene. However, similar to retroviruses, many Ty3/Gypsy LTR retrotransposons harbor additional env-like genes, promoting concepts of the infective mode of these retrotransposons. Here, we provide a further line of evidence of similarity between retroviruses and some Ty3/Gypsy LTR retrotransposons. We identify that, together with their additional genes, plant Ty3/Gypsy LTR retrotransposons of the Tat group have a second RNH, as do retroviruses. Most importantly, we show that the resulting dual RNHs of Tat LTR retrotransposons and retroviruses emerged independently, providing strong evidence for their convergent evolution. The convergent resemblance of Tat LTR retrotransposons and retroviruses may indicate similar selection pressures acting on these diverse groups of elements and reveal potential evolutionary constraints on their structure. We speculate that dual RNH is required to accelerate retrotransposon evolution through increased rates of strand transfer events and subsequent recombination events. PMID:25605791

The retrovirus HTLV-1 inserts an ectopic CTCF-binding site into the human genome.

PubMed

Satou, Yorifumi; Miyazato, Paola; Ishihara, Ko; Yaguchi, Hiroko; Melamed, Anat; Miura, Michi; Fukuda, Asami; Nosaka, Kisato; Watanabe, Takehisa; Rowan, Aileen G; Nakao, Mitsuyoshi; Bangham, Charles R M

2016-03-15

Human T-lymphotropic virus type 1 (HTLV-1) is a retrovirus that causes malignant and inflammatory diseases in ∼10% of infected people. A typical host has between 10(4) and 10(5) clones of HTLV-1-infected T lymphocytes, each clone distinguished by the genomic integration site of the single-copy HTLV-1 provirus. The HTLV-1 bZIP (HBZ) factor gene is constitutively expressed from the minus strand of the provirus, whereas plus-strand expression, required for viral propagation to uninfected cells, is suppressed or intermittent in vivo, allowing escape from host immune surveillance. It remains unknown what regulates this pattern of proviral transcription and latency. Here, we show that CTCF, a key regulator of chromatin structure and function, binds to the provirus at a sharp border in epigenetic modifications in the pX region of the HTLV-1 provirus in T cells naturally infected with HTLV-1. CTCF is a zinc-finger protein that binds to an insulator region in genomic DNA and plays a fundamental role in controlling higher order chromatin structure and gene expression in vertebrate cells. We show that CTCF bound to HTLV-1 acts as an enhancer blocker, regulates HTLV-1 mRNA splicing, and forms long-distance interactions with flanking host chromatin. CTCF-binding sites (CTCF-BSs) have been propagated throughout the genome by transposons in certain primate lineages, but CTCF binding has not previously been described in present-day exogenous retroviruses. The presence of an ectopic CTCF-BS introduced by the retrovirus in tens of thousands of genomic locations has the potential to cause widespread abnormalities in host cell chromatin structure and gene expression.

Convergent evolution of ribonuclease h in LTR retrotransposons and retroviruses.

PubMed

Ustyantsev, Kirill; Novikova, Olga; Blinov, Alexander; Smyshlyaev, Georgy

2015-05-01

Ty3/Gypsy long terminals repeat (LTR) retrotransposons are structurally and phylogenetically close to retroviruses. Two notable structural differences between these groups of genetic elements are 1) the presence in retroviruses of an additional envelope gene, env, which mediates infection, and 2) a specific dual ribonuclease H (RNH) domain encoded by the retroviral pol gene. However, similar to retroviruses, many Ty3/Gypsy LTR retrotransposons harbor additional env-like genes, promoting concepts of the infective mode of these retrotransposons. Here, we provide a further line of evidence of similarity between retroviruses and some Ty3/Gypsy LTR retrotransposons. We identify that, together with their additional genes, plant Ty3/Gypsy LTR retrotransposons of the Tat group have a second RNH, as do retroviruses. Most importantly, we show that the resulting dual RNHs of Tat LTR retrotransposons and retroviruses emerged independently, providing strong evidence for their convergent evolution. The convergent resemblance of Tat LTR retrotransposons and retroviruses may indicate similar selection pressures acting on these diverse groups of elements and reveal potential evolutionary constraints on their structure. We speculate that dual RNH is required to accelerate retrotransposon evolution through increased rates of strand transfer events and subsequent recombination events. © The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Multiple groups of endogenous epsilon-like retroviruses conserved across primates.

PubMed

Brown, Katherine; Emes, Richard D; Tarlinton, Rachael E

2014-11-01

Several types of cancer in fish are caused by retroviruses, including those responsible for major outbreaks of disease, such as walleye dermal sarcoma virus and salmon swim bladder sarcoma virus. These viruses form a phylogenetic group often described as the epsilonretrovirus genus. Epsilon-like retroviruses have become endogenous retroviruses (ERVs) on several occasions, integrating into germ line cells to become part of the host genome, and sections of fish and amphibian genomes are derived from epsilon-like retroviruses. However, epsilon-like ERVs have been identified in very few mammals. We have developed a pipeline to screen full genomes for ERVs, and using this pipeline, we have located over 800 endogenous epsilon-like ERV fragments in primate genomes. Genomes from 32 species of mammals and birds were screened, and epsilon-like ERV fragments were found in all primate and tree shrew genomes but no others. These viruses appear to have entered the genome of a common ancestor of Old and New World monkeys between 42 million and 65 million years ago. Based on these results, there is an ancient evolutionary relationship between epsilon-like retroviruses and primates. Clearly, these viruses had the potential to infect the ancestors of primates and were at some point a common pathogen in these hosts. Therefore, this result raises questions about the potential of epsilonretroviruses to infect humans and other primates and about the evolutionary history of these retroviruses. Epsilonretroviruses are a group of retroviruses that cause several important diseases in fish. Retroviruses have the ability to become a permanent part of the DNA of their host by entering the germ line as endogenous retroviruses (ERVs), where they lose their infectivity over time but can be recognized as retroviruses for millions of years. Very few mammals are known to have epsilon-like ERVs; however, we have identified over 800 fragments of endogenous epsilon-like ERVs in the genomes of all major

Endogenous retroviruses of sheep: a model system for understanding physiological adaptation to an evolving ruminant genome.

PubMed

Spencer, Thomas E; Palmarini, Massimo

2012-01-01

Endogenous retroviruses (ERVs) are present in the genome of all vertebrates and are remnants of ancient exogenous retroviral infections of the host germline transmitted vertically from generation to generation. Sheep betaretroviruses offer a unique model system to study the complex interaction between retroviruses and their host. The sheep genome contains 27 endogenous betaretroviruses (enJSRVs) related to the exogenous and pathogenic Jaagsiekte sheep retrovirus (JSRV), the causative agent of a transmissible lung cancer in sheep. The enJSRVs can protect their host against JSRV infection by blocking early and late steps of the JSRV replication cycle. In the female reproductive tract, enJSRVs are specifically expressed in the uterine luminal and glandular epithelia as well as in the conceptus (embryo and associated extraembryonic membranes) trophectoderm and in utero loss-of-function experiments found the enJSRVs envelope (env) to be essential for conceptus elongation and trophectoderm growth and development. Collectively, available evidence in sheep and other mammals indicate that ERVs coevolved with their hosts for millions of years and were positively selected for biological roles in genome plasticity and evolution, protection of the host against infection of related pathogenic and exogenous retroviruses, and placental development.

[The effect of retrovirus-mediated hTRT transfection into cultured oral keratinocytes].

PubMed

Huang, Ji-yan; Liu, Wei; Zhou, Zeng-tong; Zhou, Hai-wen

2014-06-01

Human telomerase reverse transcriptase (hTRT) was transfected into cultured oral keratinocytes (OKC) mediated by pBABE-tert recombined retrovirus to investigate the effect on OKC lifespan. pBABE-tert recombined retrovirus loaded with hTRT gene was amplified by transfected PT67 cells, and then transfected into cultured OKC in vitro. The positive clones of OKC were separated by puromycin and subcultured. Telomerase activity was analyzed by telomerase PCR-ELISA and PCR-PAGE. The hTRT positive clones of OKC showed telomerase expression, with extending lifespan to 8-9 passages. The hTRT transfected OKC can prolong doubly lifespan but not be immortalized, which indicates that cellular immortality mechanism is complicated and multi-controled. Telomerase activity is the key for cell immortalization but not the only impact factor.

Examining the discovery of the human retrovirus.

PubMed

Kontaratos, N; Sourvinos, G; Spandidos, D A

2010-01-01

Retroviruses have been found in many bird and animal species where they often cause various types of cancer. Dr. Robert Gallo's contribution to the field of retrovirology and the link he established between RNA viruses and cancer has been significant. Historical aspects of his discoveries in the area of human retroviruses are presented and an attempt is made to focus attention on his outstanding role.

The aliens inside us: HERV-W endogenous retroviruses and multiple sclerosis.

PubMed

Dolei, Antonina

2018-01-01

Two human endogenous retroviruses of the HERV-W family are proposed as multiple sclerosis (MS) co-factors: MS-associated retrovirus (MSRV) and ERVWE1, whose env proteins showed several potentially neuropathogenic features, in vitro and in animal models. Phase II clinical trials against HERV-Wenv are ongoing. HERV-W/MSRV was repeatedly found in MS patients, in striking parallel with MS stages, active/remission phases, and therapy outcome. The HERV-Wenv protein is highly expressed in active MS plaques. Early MSRV presence in spinal fluids predicted worst MS progression 10 years in advance. Effective anti-MS therapies strongly reduced MSRV/Syncytin-1/HERV-W expression. The Epstein-Barr virus (EBV) activates HERV-W/MSRV in vitro and in vivo, in patients with infectious mononucleosis and controls with high anti-EBNA1-IgG titers. Thus, the two main EBV/MS links (infectious mononucleosis and high anti-EBNA1-IgG titers) are paralleled by activation of HERV-W/MSRV. It is hypothesized that EBV may act as initial trigger of future MS, years later, by activating MSRV, which would act as direct neuropathogenic effector, before and during MS.

Characterizing novel endogenous retroviruses from genetic variation inferred from short sequence reads

PubMed Central

Mourier, Tobias; Mollerup, Sarah; Vinner, Lasse; Hansen, Thomas Arn; Kjartansdóttir, Kristín Rós; Guldberg Frøslev, Tobias; Snogdal Boutrup, Torsten; Nielsen, Lars Peter; Willerslev, Eske; Hansen, Anders J.

2015-01-01

From Illumina sequencing of DNA from brain and liver tissue from the lion, Panthera leo, and tumor samples from the pike-perch, Sander lucioperca, we obtained two assembled sequence contigs with similarity to known retroviruses. Phylogenetic analyses suggest that the pike-perch retrovirus belongs to the epsilonretroviruses, and the lion retrovirus to the gammaretroviruses. To determine if these novel retroviral sequences originate from an endogenous retrovirus or from a recently integrated exogenous retrovirus, we assessed the genetic diversity of the parental sequences from which the short Illumina reads are derived. First, we showed by simulations that we can robustly infer the level of genetic diversity from short sequence reads. Second, we find that the measures of nucleotide diversity inferred from our retroviral sequences significantly exceed the level observed from Human Immunodeficiency Virus infections, prompting us to conclude that the novel retroviruses are both of endogenous origin. Through further simulations, we rule out the possibility that the observed elevated levels of nucleotide diversity are the result of co-infection with two closely related exogenous retroviruses. PMID:26493184

Gamma-Retrovirus Integration Marks Cell Type-Specific Cancer Genes: A Novel Profiling Tool in Cancer Genomics.

PubMed

Gilroy, Kathryn L; Terry, Anne; Naseer, Asif; de Ridder, Jeroen; Allahyar, Amin; Wang, Weiwei; Carpenter, Eric; Mason, Andrew; Wong, Gane K-S; Cameron, Ewan R; Kilbey, Anna; Neil, James C

2016-01-01

Retroviruses have been foundational in cancer research since early studies identified proto-oncogenes as targets for insertional mutagenesis. Integration of murine gamma-retroviruses into the host genome favours promoters and enhancers and entails interaction of viral integrase with host BET/bromodomain factors. We report that this integration pattern is conserved in feline leukaemia virus (FeLV), a gamma-retrovirus that infects many human cell types. Analysis of FeLV insertion sites in the MCF-7 mammary carcinoma cell line revealed strong bias towards active chromatin marks with no evidence of significant post-integration growth selection. The most prominent FeLV integration targets had little overlap with the most abundantly expressed transcripts, but were strongly enriched for annotated cancer genes. A meta-analysis based on several gamma-retrovirus integration profiling (GRIP) studies in human cells (CD34+, K562, HepG2) revealed a similar cancer gene bias but also remarkable cell-type specificity, with prominent exceptions including a universal integration hotspot at the long non-coding RNA MALAT1. Comparison of GRIP targets with databases of super-enhancers from the same cell lines showed that these have only limited overlap and that GRIP provides unique insights into the upstream drivers of cell growth. These observations elucidate the oncogenic potency of the gamma-retroviruses and support the wider application of GRIP to identify the genes and growth regulatory circuits that drive distinct cancer types.

[Cell lineage tracing of regenerating cells after partial pancreatectomy using pseudo-type retrovirus].

PubMed

Zhang, Lixin; Ju, Xiaofang; Wang, Fa; Guo, Zhiwei; Piao, Shanhua; Teng, Chunbo

2008-04-01

Pancreas is an important mixed gland having both endocrine and exocrine functions, and has been proven regeneration after injury. To explore the cell lineage tracing methods in pancreas in vivo and the regenerate cells source, we used pseudo-type retrovirus to transfect adult mouse pancreas which had been partially pancreatectomized by rubbing the kerf using a cotton stick saturated with retrovirus suspension then injecting 100 microL retrovirus suspension into pancreas, injecting 100 microL retrovirus by caudal vein, or interperitoneally injecting retrovirus respectively. The results showed that the method of rubbing the kerf then injection of retrovirus suspension into pancreas could more effectively mark the pancreatic cells than the caudal vein injection and the intraperitoneal injection did in vivo. Furthermore, this study also found that some acinus cells could accept injury stimulus signals to regenerate through resuming mitosis after pancreatic injury. This study establishes a cell lineage tracing method in pancreas in vivo using retrovirus and offers a clue for gene therapy of pancreatic diseases using retrovirus vectors.

Characterization of chicken c-ski oncogene products expressed by retrovirus vectors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sutrave, P.; Copeland, T.D.; Hughes, S.H.

1990-06-01

The authors have constructed replication-competent avian retrovirus vectors that contain two of the three known types of chicken c-{ital ski} cDNAs and a third vector that contains a truncated c-{ital ski} cDNA. They developed antisera that recognize the c-{ital ski} proteins made by the three transforming c-{ital ski} viruses. All three proteins (apparent molecular masses, 50, 60, and 90 kilodaltons) are localized primarily in the nucleus. The proteins are differentially phosphorylated; immunofluorescence also suggests that there are differences in subnuclear localization of the c-{ital ski} proteins and that c-{ital ski} protein is associated with condensed chromatin in dividing cells.

An elastic model of partial budding of retroviruses

NASA Astrophysics Data System (ADS)

Zhang, Rui; Nguyen, Toan

2008-03-01

Retroviruses are characterized by their unique infection strategy of reverse transcription, in which the genetic information flows from RNA back to DNA. The most well known representative is the human immunodeficiency virus (HIV). Unlike budding of traditional enveloped viruses, retrovirus budding happens together with the formation of spherical virus capsids at the cell membrane. Led by this unique budding mechanism, we proposed an elastic model of retrovirus budding in this work. We found that if the lipid molecules of the membrane are supplied fast enough from the cell interior, the budding always proceeds to completion. In the opposite limit, there is an optimal size of partially budded virions. The zenith angle of these partially spherical capsids, α, is given by α˜(2̂/κσ)^1/4, where κ is the bending modulus of the membrane, σ is the surface tension of the membrane, and τ characterizes the strength of capsid protein interaction. If τ is large enough such that α˜π, the budding is complete. Our model explained many features of retrovirus partial budding observed in experiments.

Endogenous Retrovirus 3 – History, Physiology, and Pathology

PubMed Central

Bustamante Rivera, Yomara Y.; Brütting, Christine; Schmidt, Caroline; Volkmer, Ines; Staege, Martin S.

2018-01-01

Endogenous viral elements (EVE) seem to be present in all eukaryotic genomes. The composition of EVE varies between different species. The endogenous retrovirus 3 (ERV3) is one of these elements that is present only in humans and other Catarrhini. Conservation of ERV3 in most of the investigated Catarrhini and the expression pattern in normal tissues suggest a putative physiological role of ERV3. On the other hand, ERV3 has been implicated in the pathogenesis of auto-immunity and cancer. In the present review we summarize knowledge about this interesting EVE. We propose the model that expression of ERV3 (and probably other EVE loci) under pathological conditions might be part of a metazoan SOS response. PMID:29379485

APOBEC3-mediated hypermutation of retroviral vectors produced from some retrovirus packaging cell lines.

PubMed

Miller, A D; Metzger, M J

2011-05-01

APOBEC3 proteins are packaged into retrovirus virions and can hypermutate retroviruses during reverse transcription. We found that HT-1080 human fibrosarcoma cells hypermutate retroviruses, and that the HT-1080 cell-derived FLYA13 retrovirus packaging cells also hypermutate a retrovirus vector produced using these cells. We found no hypermutation of the same vector produced by the mouse cell-derived packaging line PT67 or by human 293 cells transfected with the vector and retrovirus packaging plasmids. We expect that avoidance of vector hypermutation will be particularly important for vectors used in gene therapy, wherein mutant proteins might stimulate deleterious immune responses.

Wolbachia influences the maternal transmission of the gypsy endogenous retrovirus in Drosophila melanogaster.

PubMed

Touret, Franck; Guiguen, François; Terzian, Christophe

2014-09-02

The endosymbiotic bacteria of the genus Wolbachia are present in most insects and are maternally transmitted through the germline. Moreover, these intracellular bacteria exert antiviral activity against insect RNA viruses, as in Drosophila melanogaster, which could explain the prevalence of Wolbachia bacteria in natural populations. Wolbachia is maternally transmitted in D. melanogaster through a mechanism that involves distribution at the posterior pole of mature oocytes and then incorporation into the pole cells of the embryos. In parallel, maternal transmission of several endogenous retroviruses is well documented in D. melanogaster. Notably, gypsy retrovirus is expressed in permissive follicle cells and transferred to the oocyte and then to the offspring by integrating into their genomes. Here, we show that the presence of Wolbachia wMel reduces the rate of gypsy insertion into the ovo gene. However, the presence of Wolbachia does not modify the expression levels of gypsy RNA and envelope glycoprotein from either permissive or restrictive ovaries. Moreover, Wolbachia affects the pattern of distribution of the retroviral particles and the gypsy envelope protein in permissive follicle cells. Altogether, our results enlarge the knowledge of the antiviral activity of Wolbachia to include reducing the maternal transmission of endogenous retroviruses in D. melanogaster. Animals have established complex relationships with bacteria and viruses that spread horizontally among individuals or are vertically transmitted, i.e., from parents to offspring. It is well established that members of the genus Wolbachia, maternally inherited symbiotic bacteria present mainly in arthropods, reduce the replication of several RNA viruses transmitted horizontally. Here, we demonstrate for the first time that Wolbachia diminishes the maternal transmission of gypsy, an endogenous retrovirus in Drosophila melanogaster. We hypothesize that gypsy cannot efficiently integrate into the germ

Modulating drug resistance by targeting BCRP/ABCG2 using retrovirus-mediated RNA interference.

PubMed

Xie, Ni; Mou, Lisha; Yuan, Jianhui; Liu, Wenlan; Deng, Tingting; Li, Zigang; Jing, Yi; Jin, Yi; Hu, Zhangli

2014-01-01

The BCRP/ABCG2 transporter, which mediates drug resistance in many types of cells, depends on energy provided by ATP hydrolysis. Here, a retrovirus encoding a shRNA targeting the ATP-binding domain of this protein was used to screen for highly efficient agents that could reverse drug resistance and improve cell sensitivity to drugs, thus laying the foundation for further studies and applications. To target the ATP-binding domain of BCRP/ABCG2, pLenti6/BCRPsi shRNA recombinant retroviruses, with 20 bp target sequences starting from the 270th, 745th and 939th bps of the 6th exon, were constructed and packaged. The pLenti6/BCRPsi retroviruses (V-BCRPi) that conferred significant knockdown effects were screened using a drug-sensitivity experiment and flow cytometry. The human choriocarcinoma cell line JAR, which highly expresses endogenous BCRP/ABCG2, was injected under the dorsal skin of a hairless mouse to initiate a JAR cytoma. After injecting V-BCRPi-infected JAR tumor cells into the dorsal skin of hairless mice, BCRP/ABCG2 expression in the tumor tissue was determined using immunohistochemistry, fluorescent quantitative RT-PCR and Western blot analyses. After intraperitoneal injection of BCRP/ABCG2-tolerant 5-FU, the tumor volume, weight change, and apoptosis rate of the tumor tissue were determined using in situ hybridization. V-BCRPi increased the sensitivity of the tumor histiocytes to 5-FU and improved the cell apoptosis-promoting effects of 5-FU in the tumor. The goal of the in vivo and in vitro studies was to screen for an RNA interference recombinant retrovirus capable of stably targeting the ATP-binding domain of BCRP/ABCG2 (V-BCRPi) to inhibit its function. A new method to improve the chemo-sensitivity of breast cancer and other tumor cells was discovered, and this method could be used for gene therapy and functional studies of malignant tumors.

Efficient retrovirus-mediated transfer and expression of a human adenosine deaminase gene in diploid skin fibroblasts from an adenosine deaminase-deficient human

DOE Office of Scientific and Technical Information (OSTI.GOV)

Palmer, T.D.; Hock, R.A.; Osborne, W.R.A.

1987-02-01

Skin fibroblasts might be considered suitable recipients for therapeutic genes to cure several human genetic diseases; however, these cells are resistant to gene transfer by most methods. The authors studied the ability of retroviral vectors to transfer genes into normal human diploid skin fibroblasts. Retroviruses carrying genes for neomycin or hygromycin B resistance conferred drug resistance to greater than 50% of the human fibroblasts after a single exposure to virus-containing medium. This represents at least a 500-fold increase in efficiency over other methods. Transfer was achieved in the absence of helper virus by using amphotropic retrovirus-packaging cells. A retrovirus vectormore » containing a human adenosine deaminase (ADA) cDNA was constructed and used to infect ADA/sup -/ fibroblasts from a patient with ADA deficiency. The infected cells produced 12-fold more ADA enzyme than fibroblasts from normal individuals and were able to rapidly metabolize exogenous deoxyadenosine and adenosine, metabolites that accumulate in plasma in ADA-deficient patients and are responsible for the severe combined immunodeficiency in these patients. These experiments indicate the potential of retrovirus-mediated gene transfer into human fibroblasts for gene therapy.« less

Detection of the human endogenous retrovirus ERV3-encoded Env-protein in human tissues using antibody-based proteomics.

PubMed

Fei, Chen; Atterby, Christina; Edqvist, Per-Henrik; Pontén, Fredrik; Zhang, Wei Wei; Larsson, Erik; Ryan, Frank P

2014-01-01

There is growing evidence to suggest that human endogenous retroviruses (HERVs) have contributed to human evolution, being expressed in development, normal physiology and disease. A key difficulty in the scientific evaluation of this potential viral contribution is the accurate demonstration of virally expressed protein in specific human cells and tissues. In this study, we have adopted the endogenous retrovirus, ERV3, as our test model in developing a reliable high-capacity methodology for the expression of such endogenous retrovirus-coded protein. Two affinity-purified polyclonal antibodies to ERV3 Env-encoded protein were generated to detect the corresponding protein expression pattern in specific human cells, tissues and organs. Sampling included normal tissues from 144 individuals ranging from childhood to old age. This included more than forty different tissues and organs and some 216 different cancer tissues representing the twenty commonest forms of human cancer. The Rudbeck Laboratory, Uppsala University and Uppsala University Hospital, Uppsala, Sweden. The potential expression at likely physiological level of the ERV3Env encoded protein in a wide range of human cells, tissues and organs. We found that ERV3 encoded Env protein is expressed at substantive levels in placenta, testis, adrenal gland, corpus luteum, Fallopian tubes, sebaceous glands, astrocytes, bronchial epithelium and the ducts of the salivary glands. Substantive expression was also seen in a variety of epithelial cells as well as cells known to undergo fusion in inflammation and in normal physiology, including fused macrophages, myocardium and striated muscle. This contrasted strongly with the low levels expressed in other tissues types. These findings suggest that this virus plays a significant role in human physiology and may also play a possible role in disease. This technique can now be extended to the study of other HERV genomes within the human chromosomes that may have contributed to

tirant, a newly discovered active endogenous retrovirus in Drosophila simulans.

PubMed

Akkouche, Abdou; Rebollo, Rita; Burlet, Nelly; Esnault, Caroline; Martinez, Sonia; Viginier, Barbara; Terzian, Christophe; Vieira, Cristina; Fablet, Marie

2012-04-01

Endogenous retroviruses have the ability to become permanently integrated into the genomes of their host, and they are generally transmitted vertically from parent to progeny. With the exception of gypsy, few endogenous retroviruses have been identified in insects. In this study, we describe the tirant endogenous retrovirus in a subset of Drosophila simulans natural populations. By focusing on the envelope gene, we show that the entire retroviral cycle (transcription, translation, and retrotransposition) can be completed for tirant within one population of this species.

Seroprevalence of retrovirus in North American captive macropodidae.

PubMed

Georoff, Timothy A; Joyner, Priscilla H; Hoover, John P; Payton, Mark E; Pogranichniy, Roman M

2008-09-01

Laboratory records of serology results from captive macropodidae sampled between 1997 and 2005 were reviewed to assess the seroprevalence of retrovirus exposure. Serum samples from 269 individuals (136 males, 133 females) representing 10 species of macropods housed in 31 North American captive collections were analyzed for retrovirus antibody using an indirect immunofluorescent assay. The prevalence of positive antibody titers comparing male versus female, between species, between age groups, and among animals with identified parentage was examined by nonparametric statistical analyses. Median age of animals at time of sample collection was 36 mo (range 2-201 mo). Total percentage seropositive was 20.4%. Serum antibody was detected in 31 of 47 (66.0%) tammar wallaby (Macropus eugenii), nine of 24 (37.5%) yellow-footed rock wallaby (Petrogale xanthopus), four of 11 (36.4%) swamp wallaby (Wallabia bicolor), 10 of 80 (12.5%) red-necked wallaby (Macropus rufogriseus), and one of 54 (1.9%) parma wallaby (Macropus parma). No individuals of western gray kangaroo (n=3) (Macropus fuliginosus), eastern gray kangaroo (n=19) (Macropus giganteus), common wallaroo (n=6) (Macropus robustus), red kangaroo (n=11) (Macropus rufus), or Matschie's tree kangaroo (n=14) (Dendrolagus matschiei) were positive for retrovirus antibody. These results demonstrate that five species of captive macropods have a history of exposure to retrovirus, with the highest percentage seropositive and highest statistical correlation in M. eugenii (pair-wise Fisher's exact test, alpha = 0.05). Additionally, one wild-caught M. eugenii was confirmed seropositive during quarantine period, indicating that retrovirus exposure may exist in wild populations.

Investigation of Endogenous Retrovirus Sequences in the Neighborhood of Genes Up-regulated in a Neuroblastoma Model after Treatment with Hypoxia-Mimetic Cobalt Chloride

PubMed Central

Brütting, Christine; Narasimhan, Harini; Hoffmann, Frank; Kornhuber, Malte E.; Staege, Martin S.; Emmer, Alexander

2018-01-01

Human endogenous retroviruses (ERVs) have been found to be associated with different diseases, e.g., multiple sclerosis (MS). Most human ERVs integrated in our genome are not competent to replicate and these sequences are presumably silent. However, transcription of human ERVs can be reactivated, e.g., by hypoxia. Interestingly, MS has been linked to hypoxia since decades. As some patterns of demyelination are similar to white matter ischemia, hypoxic damage is discussed. Therefore, we are interested in the association between hypoxia and ERVs. As a model, we used human SH-SY5Y neuroblastoma cells after treatment with the hypoxia-mimetic cobalt chloride and analyzed differences in the gene expression profiles in comparison to untreated cells. The vicinity of up-regulated genes was scanned for endogenous retrovirus-derived sequences. Five genes were found to be strongly up-regulated in SH-SY5Y cells after treatment with cobalt chloride: clusterin, glutathione peroxidase 3, insulin-like growth factor 2, solute carrier family 7 member 11, and neural precursor cell expressed developmentally down-regulated protein 9. In the vicinity of these genes we identified large (>1,000 bp) open reading frames (ORFs). Most of these ORFs showed only low similarities to proteins from retro-transcribing viruses. However, we found very high similarity between retrovirus envelope sequences and a sequence in the vicinity of neural precursor cell expressed developmentally down-regulated protein 9. This sequence encodes the human endogenous retrovirus group FRD member 1, the encoded protein product is called syncytin 2. Transfection of syncytin 2 into the well-characterized Ewing sarcoma cell line A673 was not able to modulate the low immunostimulatory activity of this cell line. Future research is needed to determine whether the identified genes and the human endogenous retrovirus group FRD member 1 might play a role in the etiology of MS. PMID:29515560

Investigation of Endogenous Retrovirus Sequences in the Neighborhood of Genes Up-regulated in a Neuroblastoma Model after Treatment with Hypoxia-Mimetic Cobalt Chloride.

PubMed

Brütting, Christine; Narasimhan, Harini; Hoffmann, Frank; Kornhuber, Malte E; Staege, Martin S; Emmer, Alexander

2018-01-01

Human endogenous retroviruses (ERVs) have been found to be associated with different diseases, e.g., multiple sclerosis (MS). Most human ERVs integrated in our genome are not competent to replicate and these sequences are presumably silent. However, transcription of human ERVs can be reactivated, e.g., by hypoxia. Interestingly, MS has been linked to hypoxia since decades. As some patterns of demyelination are similar to white matter ischemia, hypoxic damage is discussed. Therefore, we are interested in the association between hypoxia and ERVs. As a model, we used human SH-SY5Y neuroblastoma cells after treatment with the hypoxia-mimetic cobalt chloride and analyzed differences in the gene expression profiles in comparison to untreated cells. The vicinity of up-regulated genes was scanned for endogenous retrovirus-derived sequences. Five genes were found to be strongly up-regulated in SH-SY5Y cells after treatment with cobalt chloride: clusterin, glutathione peroxidase 3, insulin-like growth factor 2, solute carrier family 7 member 11, and neural precursor cell expressed developmentally down-regulated protein 9. In the vicinity of these genes we identified large (>1,000 bp) open reading frames (ORFs). Most of these ORFs showed only low similarities to proteins from retro-transcribing viruses. However, we found very high similarity between retrovirus envelope sequences and a sequence in the vicinity of neural precursor cell expressed developmentally down-regulated protein 9. This sequence encodes the human endogenous retrovirus group FRD member 1, the encoded protein product is called syncytin 2. Transfection of syncytin 2 into the well-characterized Ewing sarcoma cell line A673 was not able to modulate the low immunostimulatory activity of this cell line. Future research is needed to determine whether the identified genes and the human endogenous retrovirus group FRD member 1 might play a role in the etiology of MS.

Retrovirus-mediated conditional immortalization and analysis of established cell lines of osteoclast precursor cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kawata, Shigehisa; Suzuki, Jun; Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0871

2006-11-10

Osteoclast precursor cells (OPCs) have previously been established from bone marrow cells of SV40 temperature-sensitive T antigen-expressing transgenic mice. Here, we use retrovirus-mediated gene transfer to conditionally immortalize OPCs by expressing temperature-sensitive large T antigen (tsLT) from wild type bone marrow cells. The immortalized OPCs proliferated at the permissive temperature of 33.5 deg. C, but stopped growing at the non-permissive temperature of 39 deg. C. In the presence of receptor activator of NF{kappa}B ligand (RANKL), the OPCs differentiated into tartrate-resistant acid phosphatase (TRAP)-positive cells and formed multinucleate osteoclasts at 33.5 deg. C. From these OPCs, we cloned two types ofmore » cell lines. Both differentiated into TRAP-positive cells, but one formed multinucleate osteoclasts while the other remained unfused in the presence of RANKL. These results indicate that the established cell lines are useful for analyzing mechanisms of differentiation, particularly multinucleate osteoclast formation. Retrovirus-mediated conditional immortalization should be a useful method to immortalize OPCs from primary bone marrow cells.« less

KOALA RETROVIRUS: A REVIEW.

PubMed

Kinney, Matthew E; Pye, Geoffrey W

2016-06-01

Koala retrovirus (KoRV) is a gammaretrovirus that has been identified in both captive and free-ranging koalas ( Phascolarctos cinereus ) with variable geographic distribution in Australia. KoRV is capable of both exogenous and endogenous transmission, which provides an interesting research platform for scientists to study active retrovirus endogenization into a host genome and offers veterinary scientists an opportunity to examine the clinical consequences of KoRV infection in koalas. Causation between KoRV and frequently recognized clinical conditions associated with immune suppression and neoplasia in koalas has not been definitively established, however research continues to evaluate a potential association. Three KoRV variants, KoRV-A, KoRV-B, and KoRV-J, have been the most thoroughly described and preliminary evidence suggests KoRV variability may be fundamental in host pathogenicity. In addition to reviewing what is currently known about KoRV, this article discusses treatment, management, and future research directions.

Identification, characterization, and comparative genomic distribution of the HERV-K (HML-2) group of human endogenous retroviruses

PubMed Central

2011-01-01

Background Integration of retroviral DNA into a germ cell may lead to a provirus that is transmitted vertically to that host's offspring as an endogenous retrovirus (ERV). In humans, ERVs (HERVs) comprise about 8% of the genome, the vast majority of which are truncated and/or highly mutated and no longer encode functional genes. The most recently active retroviruses that integrated into the human germ line are members of the Betaretrovirus-like HERV-K (HML-2) group, many of which contain intact open reading frames (ORFs) in some or all genes, sometimes encoding functional proteins that are expressed in various tissues. Interestingly, this expression is upregulated in many tumors ranging from breast and ovarian tissues to lymphomas and melanomas, as well as schizophrenia, rheumatoid arthritis, and other disorders. Results No study to date has characterized all HML-2 elements in the genome, an essential step towards determining a possible functional role of HML-2 expression in disease. We present here the most comprehensive and accurate catalog of all full-length and partial HML-2 proviruses, as well as solo LTR elements, within the published human genome to date. Furthermore, we provide evidence for preferential maintenance of proviruses and solo LTR elements on gene-rich chromosomes of the human genome and in proximity to gene regions. Conclusions Our analysis has found and corrected several errors in the annotation of HML-2 elements in the human genome, including mislabeling of a newly identified group called HML-11. HML-elements have been implicated in a wide array of diseases, and characterization of these elements will play a fundamental role to understand the relationship between endogenous retrovirus expression and disease. PMID:22067224

Complementation between avirulent Newcastle disease virus and a fusion protein gene expressed from a retrovirus vector: requirements for membrane fusion.

PubMed Central

Morrison, T; McQuain, C; McGinnes, L

1991-01-01

The cDNA derived from the fusion gene of the virulent AV strain of Newcastle disease virus (NDV) was expressed in chicken embryo cells by using a retrovirus vector. The fusion protein expressed in this system was transported to the cell surface and was efficiently cleaved into the disulfide-linked F1-F2 form found in infectious virions. The cells expressing the fusion gene grew normally and could be passaged many times. Monolayers of these cells would plaque, in the absence of trypsin, avirulent NDV strains (strains which encode a fusion protein which is not cleaved in tissue culture). Fusion protein-expressing cells would not fuse if mixed with uninfected cells or uninfected cells expressing the hemagglutinin-neuraminidase (HN) protein. However, the fusion protein-expressing cells, if infected with avirulent strains of NDV, would fuse with uninfected cells, suggesting that fusion requires both the fusion protein and another viral protein expressed in the same cell. Fusion was also seen after transfection of the HN protein gene into fusion protein-expressing cells. Thus, the expressed fusion protein gene is capable of complementing the virus infection, providing an active cleaved fusion protein required for the spread of infection. However, the fusion protein does not mediate cell fusion unless the cell also expresses the HN protein. Fusion protein-expressing cells would not plaque influenza virus in the absence of trypsin, nor would influenza virus-infected fusion protein-expressing cells fuse with uninfected cells. Thus, the influenza virus HA protein will not substitute for the NDV HN protein in cell-to-cell fusion. Images PMID:1987376

Human endogenous retroviruses and multiple sclerosis: innocent bystanders or disease determinants?

PubMed

Antony, Joseph M; Deslauriers, Andre M; Bhat, Rakesh K; Ellestad, Kristofer K; Power, Christopher

2011-02-01

Human endogenous retroviruses (HERVs) constitute 5-8% of human genomic DNA and are replication incompetent despite expression of individual HERV genes from different chromosomal loci depending on the specific tissue. Several HERV genes have been detected as transcripts and proteins in the central nervous system, frequently in the context of neuroinflammation. The HERV-W family has received substantial attention in large part because of associations with diverse syndromes including multiple sclerosis (MS) and several psychiatric disorders. A HERV-W-related retroelement, multiple sclerosis retrovirus (MSRV), has been reported in MS patients to be both a biomarker as well as an effector of aberrant immune responses. HERV-H and HERV-K have also been implicated in MS and other neurological diseases but await delineation of their contributions to disease. The HERV-W envelope-encoded glycosylated protein, syncytin-1, is encoded by chromosome 7q21 and exhibits increased glial expression within MS lesions. Overexpression of syncytin-1 in glia induces endoplasmic reticulum stress leading to neuroinflammation and the induction of free radicals, which damage proximate cells. Syncytin-1's receptor, ASCT1 is a neutral amino acid transporter expressed on glia and is suppressed in white matter of MS patients. Of interest, antioxidants ameliorate syncytin-1's neuropathogenic effects raising the possibility of using these agents as therapeutics for neuroinflammatory diseases. Given the multiple insertion sites of HERV genes as complete and incomplete open reading frames, together with their differing capacity to be expressed and the complexities of individual HERVs as both disease markers and bioactive effectors, HERV biology is a compelling area for understanding neuropathogenic mechanisms and developing new therapeutic strategies. 2010 Elsevier B.V. All rights reserved.

A Paradigm for Virus–Host Coevolution: Sequential Counter-Adaptations between Endogenous and Exogenous Retroviruses

PubMed Central

Arnaud, Frederick; Caporale, Marco; Varela, Mariana; Biek, Roman; Chessa, Bernardo; Alberti, Alberto; Golder, Matthew; Mura, Manuela; Zhang, Ya-ping; Yu, Li; Pereira, Filipe; DeMartini, James C; Leymaster, Kreg; Spencer, Thomas E; Palmarini, Massimo

2007-01-01

Endogenous retroviruses (ERVs) are remnants of ancient retroviral infections of the host germline transmitted vertically from generation to generation. It is hypothesized that some ERVs are used by the host as restriction factors to block the infection of pathogenic retroviruses. Indeed, some ERVs efficiently interfere with the replication of related exogenous retroviruses. However, data suggesting that these mechanisms have influenced the coevolution of endogenous and/or exogenous retroviruses and their hosts have been more difficult to obtain. Sheep are an interesting model system to study retrovirus-host coevolution because of the coexistence in this animal species of two exogenous (i.e., horizontally transmitted) oncogenic retroviruses, Jaagsiekte sheep retrovirus and Enzootic nasal tumor virus, with highly related and biologically active endogenous retroviruses (enJSRVs). Here, we isolated and characterized the evolutionary history and molecular virology of 27 enJSRV proviruses. enJSRVs have been integrating in the host genome for the last 5–7 million y. Two enJSRV proviruses (enJS56A1 and enJSRV-20), which entered the host genome within the last 3 million y (before and during speciation within the genus Ovis), acquired in two temporally distinct events a defective Gag polyprotein resulting in a transdominant phenotype able to block late replication steps of related exogenous retroviruses. Both transdominant proviruses became fixed in the host genome before or around sheep domestication (∼ 9,000 y ago). Interestingly, a provirus escaping the transdominant enJSRVs has emerged very recently, most likely within the last 200 y. Thus, we determined sequentially distinct events during evolution that are indicative of an evolutionary antagonism between endogenous and exogenous retroviruses. This study strongly suggests that endogenization and selection of ERVs acting as restriction factors is a mechanism used by the host to fight retroviral infections. PMID

A historical reflection on the discovery of human retroviruses.

PubMed

Vahlne, Anders

2009-05-01

The discovery of HIV-1 as the cause of AIDS was one of the major scientific achievements during the last century. Here the events leading to this discovery are reviewed with particular attention to priority and actual contributions by those involved. Since I would argue that discovering HIV was dependent on the previous discovery of the first human retrovirus HTLV-I, the history of this discovery is also re-examined. The first human retroviruses (HTLV-I) was first reported by Robert C. Gallo and coworkers in 1980 and reconfirmed by Yorio Hinuma and coworkers in 1981. These discoveries were in turn dependent on the previous discovery by Gallo and coworkers in 1976 of interleukin 2 or T-cell growth factor as it was called then. HTLV-II was described by Gallo's group in 1982. A human retrovirus distinct from HTLV-I and HTLV-II in that it was shown to have the morphology of a lentivirus was in my mind described for the first time by Luc Montagnier in an oral presentation at Cold Spring Harbor in September of 1983. This virus was isolated from a patient with lymphadenopathy using the protocol previously described for HTLV by Gallo. The first peer reviewed paper by Montagnier's group of such a retrovirus, isolated from two siblings of whom one with AIDS, appeared in Lancet in April of 1984. However, the proof that a new human retrovirus (HIV-1) was the cause of AIDS was first established in four publications by Gallo's group in the May 4th issue of Science in 1984.

Infectious disease risks in xenotransplantation.

PubMed

Fishman, Jay A

2018-03-07

Hurdles exist to clinical xenotransplantation including potential infectious transmission from nonhuman species to xenograft recipients. In anticipation of clinical trials of xenotransplantation, the associated infectious risks have been investigated. Swine and immunocompromised humans share some potential pathogens. Swine herpesviruses including porcine cytomegalovirus (PCMV) and porcine lymphotropic herpesvirus (PLHV) are largely species-specific and do not, generally, infect human cells. Human cellular receptors exist for porcine endogenous retrovirus (PERV), which infects certain human-derived cell lines in vitro. PERV-inactivated pigs have been produced recently. Human infection due to PERV has not been described. A screening paradigm can be applied to exclude potential human pathogens from "designated pathogen free" breeding colonies. Various microbiological assays have been developed for screening and diagnosis including antibody-based tests and qualitative and quantitative molecular assays for viruses. Additional assays may be required to diagnose pig-specific organisms in human xenograft recipients. Significant progress has been made in the evaluation of the potential infectious risks of clinical xenotransplantation. Infectious risk would be amplified by intensive immunosuppression. The available data suggest that risks of xenotransplant-associated recipient infection are manageable and that clinical trials can be performed safely. Possible infectious risks of xenotransplantation to the community at large are undefined but merit consideration. © 2018 The American Society of Transplantation and the American Society of Transplant Surgeons.

Distinct Particle Morphologies Revealed through Comparative Parallel Analyses of Retrovirus-Like Particles.

PubMed

Martin, Jessica L; Cao, Sheng; Maldonado, Jose O; Zhang, Wei; Mansky, Louis M

2016-09-15

The Gag protein is the main retroviral structural protein, and its expression alone is usually sufficient for production of virus-like particles (VLPs). In this study, we sought to investigate-in parallel comparative analyses-Gag cellular distribution, VLP size, and basic morphological features using Gag expression constructs (Gag or Gag-YFP, where YFP is yellow fluorescent protein) created from all representative retroviral genera: Alpharetrovirus, Betaretrovirus, Deltaretrovirus, Epsilonretrovirus, Gammaretrovirus, Lentivirus, and Spumavirus. We analyzed Gag cellular distribution by confocal microscopy, VLP budding by thin-section transmission electron microscopy (TEM), and general morphological features of the VLPs by cryogenic transmission electron microscopy (cryo-TEM). Punctate Gag was observed near the plasma membrane for all Gag constructs tested except for the representative Beta- and Epsilonretrovirus Gag proteins. This is the first report of Epsilonretrovirus Gag localizing to the nucleus of HeLa cells. While VLPs were not produced by the representative Beta- and Epsilonretrovirus Gag proteins, the other Gag proteins produced VLPs as confirmed by TEM, and morphological differences were observed by cryo-TEM. In particular, we observed Deltaretrovirus-like particles with flat regions of electron density that did not follow viral membrane curvature, Lentivirus-like particles with a narrow range and consistent electron density, suggesting a tightly packed Gag lattice, and Spumavirus-like particles with large envelope protein spikes and no visible electron density associated with a Gag lattice. Taken together, these parallel comparative analyses demonstrate for the first time the distinct morphological features that exist among retrovirus-like particles. Investigation of these differences will provide greater insights into the retroviral assembly pathway. Comparative analysis among retroviruses has been critically important in enhancing our understanding of

Real-time quantitative PCR for retrovirus-like particle quantification in CHO cell culture.

PubMed

de Wit, C; Fautz, C; Xu, Y

2000-09-01

Chinese hamster ovary (CHO) cells have been widely used to manufacture recombinant proteins intended for human therapeutic uses. Retrovirus-like particles, which are apparently defective and non-infectious, have been detected in all CHO cells by electron microscopy (EM). To assure viral safety of CHO cell-derived biologicals, quantification of retrovirus-like particles in production cell culture and demonstration of sufficient elimination of such retrovirus-like particles by the down-stream purification process are required for product market registration worldwide. EM, with a detection limit of 1x10(6) particles/ml, is the standard retrovirus-like particle quantification method. The whole process, which requires a large amount of sample (3-6 litres), is labour intensive, time consuming, expensive, and subject to significant assay variability. In this paper, a novel real-time quantitative PCR assay (TaqMan assay) has been developed for the quantification of retrovirus-like particles. Each retrovirus particle contains two copies of the viral genomic particle RNA (pRNA) molecule. Therefore, quantification of retrovirus particles can be achieved by quantifying the pRNA copy number, i.e. every two copies of retroviral pRNA is equivalent to one retrovirus-like particle. The TaqMan assay takes advantage of the 5'-->3' exonuclease activity of Taq DNA polymerase and utilizes the PRISM 7700 Sequence Detection System of PE Applied Biosystems (Foster City, CA, U.S.A.) for automated pRNA quantification through a dual-labelled fluorogenic probe. The TaqMan quantification technique is highly comparable to the EM analysis. In addition, it offers significant advantages over the EM analysis, such as a higher sensitivity of less than 600 particles/ml, greater accuracy and reliability, higher sample throughput, more flexibility and lower cost. Therefore, the TaqMan assay should be used as a substitute for EM analysis for retrovirus-like particle quantification in CHO cell

Effect of Type-I Interferon on Retroviruses

PubMed Central

Gómez-Lucía, Esperanza; Collado, Victorio M.; Miró, Guadalupe; Doménech, Ana

2009-01-01

Type-I interferons (IFN-I) play an important role in the innate immune response to several retroviruses. They seem to be effective in controlling the in vivo infection, though many of the clinical signs of retroviral infection may be due to their continual presence which over-stimulates the immune system and activates apoptosis. IFN-I not only affect the immune system, but also operate directly on virus replication. Most data suggest that the in vitro treatment with IFN-I of retrovirus infected cells inhibits the final stages of virogenesis, avoiding the correct assembly of viral particles and their budding, even though the mechanism is not well understood. However, in some retroviruses IFN-I may also act at a previous stage as some retroviral LTRs posses sequences homologous to the IFN-stimulated response element (ISRE). When stimulated, ISREs control viral transcription. HIV-1 displays several mechanisms for evading IFN-I, such as through Tat and Nef. Besides IFN-α and IFN-β, some other type I IFN, such as IFN-τ and IFN-ω, have potent antiviral activity and are promising treatment drugs. PMID:21994560

Molecular cloning and long terminal repeat sequences of human endogenous retrovirus genes related to types A and B retrovirus genes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ono, M.

1986-06-01

By using a DNA fragment primarily encoding the reverse transcriptase (pol) region of the Syrian hamster intracisternal A particle (IAP; type A retrovirus) gene as a probe, human endogenous retrovirus genes, tentatively termed HERV-K genes, were cloned from a fetal human liver gene library. Typical HERV-K genes were 9.1 or 9.4 kilobases in length, having long terminal repeats (LTRs) of ca. 970 base pairs. Many structural features commonly observed on the retrovirus LTRs, such as the TATAA box, polyadenylation signal, and terminal inverted repeats, were present on each LTR, and a lysine (K) tRNA having a CUU anticodon was identifiedmore » as a presumed primer tRNA. The HERV-K LTR, however, had little sequence homology to either the IAP LTR or other typical oncovirus LTRs. By filter hybridization, the number of HERV-K genes was estimated to be ca. 50 copies per haploid human genome. The cloned mouse mammary tumor virus (type B) gene was found to hybridize with both the HERV-K and IAP genes to essentially the same extent.« less

Retrovirus-mediated siRNA targeting TRPM7 gene induces apoptosis in RBL-2H3 cells.

PubMed

Ng, N-M; Jiang, S-P; Lv, Z-Q

2012-09-01

Calcium signaling is important for both normal physiologic processes and pathology of various diseases. Transient receptor potential melastatin 7 (TRPM7) gene has been reported to be a potential candidate for calcium influx. The present study aimed to investigate the possible role of TRPM7 channels in apoptosis in rat basophilic leukemia mast cell line (RBL-2H3), which is widely used in mast cell-associated studies. A recombinant retrovirus vector siRNA targeting rat TRPM7 gene was constructed and identified. Cellular survival was assessed by MTT. Cell apoptosis was evaluated by flow cytometry and TUNEL-FITC/Hoechst 33258 staining. The transfection efficiency by retrovirus vector was about 60%-70%. Transfection with TRPM7 siRNA significantly reduced TRPM7 expression both at mRNA and protein levels. Suppression of TRPM7 expression by siRNA led to significantly decreased cellular survival rates and increased apoptosis rates in RBL-2H3 cells. This study indicates that TRPM7 is involved in the apoptosis process in RBL-2H3 cells.

Inhibition of West Nile Virus replication by retrovirus-delivered small interfering RNA in human neuroblastoma cells.

PubMed

Yang, Yongbo; Wu, Chengxiang; Wu, Jianguo; Nerurkar, Vivek R; Yanagihara, Richard; Lu, Yuanan

2008-05-01

West Nile virus (WNV) has been responsible for the largest outbreaks of arboviral encephalitis in U.S. history. No specific drug is currently available for the effective treatment of WNV infection. To exploit RNA interference as a potential therapeutic approach, a Moloney murine leukemia virus-based retrovirus vector was used to effectively deliver WNV-specific small interfering RNA (siRNA) into human neuroblastoma HTB-11 cells. Viral plaque assays demonstrated that transduced cells were significantly refractory to WNV replication, as compared to untransduced control cells (P < 0.05), which correlated with the reduced expression of target viral genes and respective viral proteins. Therefore, retrovirus-mediated delivery of siRNA for gene silencing can be used to study the specific functions of viral genes associated with replication and may have potential therapeutic applications.

The role of BST2/tetherin in infection with the feline retroviruses

PubMed Central

Dietrich, Isabelle; Hosie, Margaret J.; Willett, Brian J.

2014-01-01

The recently identified host restriction factor tetherin (BST-2, CD317) potently inhibits the release of nascent retrovirus particles from infected cells. Recently, we reported the identification and characterization of tetherin as a novel feline retroviral restriction factor. Based on homology to human tetherin we identified a putative tetherin gene in the genome of the domestic cat (Felis catus) which was found to be expressed in different feline cell lines both prior to and post treatment with either type I or type II interferon (IFN). The predicted structure of feline tetherin (feTHN) was that of a type II single-pass transmembrane protein encoding an N-terminal transmembrane anchor, central predicted coiled-coil bearing extracellular domain to promote dimerization, and a C-terminal GPI-anchor, consistent with conservation of structure between human and feline tetherin. FeTHN displayed potent inhibition of feline immunodeficiency virus (FIV) and human immunodeficiency virus type 1 (HIV-1) particle release in single-cycle replication assays. Notably, feTHN activity was resistant to antagonism by HIV-1 Vpu. However, stable ectopic expression of feTHN mRNA in different feline cell lines had no inhibitory effect on the growth of diverse primary or cell culture-adapted strains of FIV. Hence, whereas feline tetherin efficiently blocks viral particle release in single-cycle replication assays, it might not prevent dissemination of feline retroviruses in vivo. PMID:21715020

Characterization of African Human Retroviruses Related to HTLV-III/LAV

DTIC Science & Technology

1988-06-15

the prototype AIDS virus , HIV -1 (4,5). This new human retrovirus has now been termed Human Immunodeficiency Virus Type...lImmunodeficiency Virus and its Relationship to the Human Immunodeficiency Viruses . Nature 32:539-543, 1987. (12) Guyader, M., Emerman, M., Sonigo, P...populations the number of AIDS cases is still quite low suggesting a distinct pathobiology to this new human retrovirus . Cohort studies of

Broad-scale phylogenomics provides insights into retrovirus-host evolution.

PubMed

Hayward, Alexander; Grabherr, Manfred; Jern, Patric

2013-12-10

Genomic data provide an excellent resource to improve understanding of retrovirus evolution and the complex relationships among viruses and their hosts. In conjunction with broad-scale in silico screening of vertebrate genomes, this resource offers an opportunity to complement data on the evolution and frequency of past retroviral spread and so evaluate future risks and limitations for horizontal transmission between different host species. Here, we develop a methodology for extracting phylogenetic signal from large endogenous retrovirus (ERV) datasets by collapsing information to facilitate broad-scale phylogenomics across a wide sample of hosts. Starting with nearly 90,000 ERVs from 60 vertebrate host genomes, we construct phylogenetic hypotheses and draw inferences regarding the designation, host distribution, origin, and transmission of the Gammaretrovirus genus and associated class I ERVs. Our results uncover remarkable depths in retroviral sequence diversity, supported within a phylogenetic context. This finding suggests that current infectious exogenous retrovirus diversity may be underestimated, adding credence to the possibility that many additional exogenous retroviruses may remain to be discovered in vertebrate taxa. We demonstrate a history of frequent horizontal interorder transmissions from a rodent reservoir and suggest that rats may have acted as important overlooked facilitators of gammaretrovirus spread across diverse mammalian hosts. Together, these results demonstrate the promise of the methodology used here to analyze large ERV datasets and improve understanding of retroviral evolution and diversity for utilization in wider applications.

A majority of mice show long-term expression of a human. beta. -globin gene after retrovirus transfer into hematopoietic stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bender, M.A.; Gelinas, R.E.; Miller

1989-04-01

Murine bone marrow was infected with a high-titer retrovirus vector containing the human {beta}-globin and neomycin phosphotransferase genes. Anemic W/W/sup v/ mice were transplanted with infected marrow which in some cases had been exposed to the selective agent G418. Human {beta}-globin expression was monitored in transplanted animals by using a monoclonal antibody specific for human {beta}-globin polypeptide, and hematopoietic reconstitution was monitored by using donor and recipient mice which differed in hemoglobin type. In some experiments all transplanted mice expressed the human {beta}-globin polypeptide for over 4 months, and up to 50% of peripheral erythrocytes contained detectable levels of polypeptide.more » DNA analysis of transplanted animals revealed that virtually every myeloid cell contained a provirus. Integration site analysis and reconstitution of secondary marrow recipients suggested that every mouse was reconstituted with at least one infected stem cell which had extensive repopulation capability. The ability to consistently transfer an active {beta}-globin gene into mouse hematopoietic cells improves the feasibility of using these techniques for somatic cell gene therapy in humans.« less

[Inhibitory effect of migration-inducing gene-7-shRNA recombinant retrovirus combined with endostatin on growth and metastasis of hepatoma xenograft].

PubMed

Qu, B; Chen, G N; Sheng, G N; Yu, F; Lyu, Q; Gu, Y J; Guo, L; Lyu, Y

2016-09-20

Objective: To investigate the inhibitory effect of migration-inducing gene-7(Mig-7)interfered with retrovirus-mediated RNA(shRNA)combined with recombinant human endostatin(ES)on the growth and metastasis of subcutaneous xenograft of human hepatoma cells in nude mice. Methods: Two Mig-7-mRNA oligonucleotide sequences(Mig-7-shRNA-1 and Mig-7-shRNA-2)and one sequence as a negative control(Mig-7-shRNA-N)were designed. The specific Mig-7-shRNA recombinant retrovirus expression vector plasmid was constructed and used for the transfection of human hepatoma MHCC-97H cells with high expression of Mig-7. The subcutaneous xenograft tumor model of human hepatocellular carcinoma(HCC)in nude mice was established, and according to the condition of transfection and administration, the nude mice were divided into pSIREN-M1 group, pSIREN-MN group, ES group, and pSIREN-M1+ES group. The xenograft tumor volume, mass, and metastasis were compared between groups. Immunohistochemistry was used to observe the formation of vasculogenic mimicry(VM)in xenograft tumor and the difference in tumor microvascular density(MVD), and Western blot was used to measure the expression of Mig-7 and vascular endothelial growth factor(VEGF)in each group. A one-way analysis of variance was used for comparison between groups, and the Fisher's exact test was used for comparison of continuous data between groups. Results: Compared with the pSIREN-MN group, the pSIREN-M1 group had significantly lower xenograft tumor volume, mass, and metastasis rate, Mig-7 expression, and formation of VM( P < 0.05), as well as significantly higher VEGF expression and MVD( P < 0.05). Compared with the pSIREN-MN group, the ES group had significantly lower xenograft tumor volume, mass, and metastasis rate, VEGF expression, and MVD( P < 0.05), as well as significantly higher Mig-7 expression and formation of VM( P < 0.05). Compared with the pSIREN-M1 group and the ES group, the pSIREN-M1+ES group had significantly lower xenograft

A New Approach to Establish a Cell Line with Reduced Risk of Endogenous Retroviruses

PubMed Central

Fukuma, Aiko; Yoshikawa, Rokusuke; Miyazawa, Takayuki; Yasuda, Jiro

2013-01-01

Endogenous retroviruses (ERVs) are integrated as DNA proviruses in the genomes of all mammalian species. Several ERVs are replication-competent and produced as fully infectious viruses from host cell. Thus, live-attenuated vaccines and biological substances have been prepared using the cell lines which may produce ERV. Indeed, we recently reported that several commercial live-attenuated vaccines for pets were contaminated with the infectious feline endogenous retrovirus, RD-114. In this study, to establish a cell line for vaccine manufacture with reduced risk of ERVs, we generated a cell line stably expressing human tetherin (Teth-CRFK cells). The release of infectious ERV from Teth-CRFK cells was suppressed to undetectable levels, while the production of parvovirus in Teth-CRFK cells was similar to that in parental CRFK cells. These observations suggest that Teth-CRFK cells will be useful as a cell line for the manufacture of live-attenuated vaccines or biological substances with reduced risk of ERV. PMID:23585909

A new approach to establish a cell line with reduced risk of endogenous retroviruses.

PubMed

Fukuma, Aiko; Yoshikawa, Rokusuke; Miyazawa, Takayuki; Yasuda, Jiro

2013-01-01

Endogenous retroviruses (ERVs) are integrated as DNA proviruses in the genomes of all mammalian species. Several ERVs are replication-competent and produced as fully infectious viruses from host cell. Thus, live-attenuated vaccines and biological substances have been prepared using the cell lines which may produce ERV. Indeed, we recently reported that several commercial live-attenuated vaccines for pets were contaminated with the infectious feline endogenous retrovirus, RD-114. In this study, to establish a cell line for vaccine manufacture with reduced risk of ERVs, we generated a cell line stably expressing human tetherin (Teth-CRFK cells). The release of infectious ERV from Teth-CRFK cells was suppressed to undetectable levels, while the production of parvovirus in Teth-CRFK cells was similar to that in parental CRFK cells. These observations suggest that Teth-CRFK cells will be useful as a cell line for the manufacture of live-attenuated vaccines or biological substances with reduced risk of ERV.

Widespread and highly persistent gene transfer to the CNS by retrovirus vector in utero: implication for gene therapy to Krabbe disease.

PubMed

Shen, Jin-Song; Meng, Xing-Li; Yokoo, Takashi; Sakurai, Ken; Watabe, Kazuhiko; Ohashi, Toya; Eto, Yoshikatsu

2005-05-01

Brain-directed prenatal gene therapy may benefit some lysosomal storage diseases that affect the central nervous system (CNS) before birth. Our previous study showed that intrauterine introduction of recombinant adenoviruses into cerebral ventricles results in efficient gene transfer to the CNS in the mouse. However, transgene expression decreased with time due to the non-integrative property of adenoviral vectors. In this study, in order to obtain permanent gene transduction, we investigated the feasibility of retrovirus-mediated in utero gene transduction. Concentrated retrovirus encoding the LacZ gene was injected into the cerebral ventricles of the embryos of normal and twitcher mice (a murine model of Krabbe disease) at embryonic day 12. The distribution and maintenance of the transgene expression in the recipient brain were analyzed histochemically, biochemically and by the quantitative polymerase chain reaction method pre- and postnatally. Efficient and highly persistent gene transduction to the brain was achieved both in normal and the twitcher mouse. Transduced neurons, astrocytes and oligodendrocytes were distributed throughout the brain. The transduced LacZ gene, its transcript and protein expression in the brain were maintained for 14 months without decrement. In addition, gene transduction to multiple tissues other than the brain was also detected at low levels. This study suggests that brain-directed in utero gene transfer using retrovirus vector may be beneficial to the treatment of lysosomal storage diseases with severe brain damage early in life, such as Krabbe disease. Copyright (c) 2005 John Wiley & Sons, Ltd.

Marine origin of retroviruses in the early Palaeozoic Era

NASA Astrophysics Data System (ADS)

Aiewsakun, Pakorn; Katzourakis, Aris

2017-01-01

Very little is known about the ancient origin of retroviruses, but owing to the discovery of their ancient endogenous viral counterparts, their early history is beginning to unfold. Here we report 36 lineages of basal amphibian and fish foamy-like endogenous retroviruses (FLERVs). Phylogenetic analyses reveal that ray-finned fish FLERVs exhibit an overall co-speciation pattern with their hosts, while amphibian FLERVs might not. We also observe several possible ancient viral cross-class transmissions, involving lobe-finned fish, shark and frog FLERVs. Sequence examination and analyses reveal two major lineages of ray-finned fish FLERVs, one of which had gained two novel accessory genes within their extraordinarily large genomes. Our phylogenetic analyses suggest that this major retroviral lineage, and therefore retroviruses as a whole, have an ancient marine origin and originated together with, if not before, their jawed vertebrate hosts >450 million years ago in the Ordovician period, early Palaeozoic Era.

Prevention of 3-methylcholanthrene-induced fibrosarcomas in rats pre-inoculated with endogenous rat retrovirus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fish, D.C.; Demarais, J.T.; Djurickovic, D.B.

1981-04-01

Weanling Fischer 344 rats received a single intraperitoneal injection of a 1000-fold concentrated preparation of endogenous nontransforming rat retrovirus. Ten days later, the rats were each given a single subcutaneous injection of 3-methylcholanthrene. The rats inoculated with the endogenous rat retrovirus were significantly protected against the development of cancer, whereas uninoculated rats and rats given one of several murine tetroviruses or baboon retrovirus were not protected.

Identification and Characterization of Two Closely Related Unclassifiable Endogenous Retroviruses in Pythons (Python molurus and Python curtus)

PubMed Central

Huder, Jon B.; Böni, Jürg; Hatt, Jean-Michel; Soldati, Guido; Lutz, Hans; Schüpbach, Jörg

2002-01-01

Boid inclusion body disease (BIBD) is a fatal disorder of boid snakes that is suspected to be caused by a retrovirus. In order to identify this agent, leukocyte cultures (established from Python molurus specimens with symptoms of BIBD or kept together with such diseased animals) were assessed for reverse transcriptase (RT) activity. Virus from cultures exhibiting high RT activity was banded on sucrose density gradients, and the RT peak fraction was subjected to highly efficient procedures for the identification of unknown particle-associated retroviral RNA. A 7-kb full retroviral sequence was identified, cloned, and sequenced. This virus contained intact open reading frames (ORFs) for gag, pro, pol, and env, as well as another ORF of unknown function within pol. Phylogenetic analysis showed that the virus is distantly related to viruses from both the B and D types and the mammalian C type but cannot be classified. It is present as a highly expressed endogenous retrovirus in all P. molurus individuals; a closely related, but much less expressed virus was found in all tested Python curtus individuals. All other boid snakes tested, including Python regius, Python reticulatus, Boa constrictor, Eunectes notaeus, and Morelia spilota, were virus negative, independent of whether they had BIBD or not. Virus isolated from P. molurus could not be transmitted to the peripheral blood mononuclear cells of B. constrictor or P. regius. Thus, there is no indication that this novel virus, which we propose to name python endogenous retrovirus (PyERV), is causally linked with BIBD. PMID:12097574

Interspecies radioimmunoassay for the major structural proteins of primate type-D retroviruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Colcher, D.; Teramoto, Y.A.; Schlom, J.

1977-12-01

A competition radioimmunoassay has been developed in which type-D retroviruses from three primate species compete. The assay utilizes the major structural protein (36,000 daltons) of the endogenous squirrel monkey retrovirus and antisera directed against the major structural protein (27,000 daltons) of the Mason-Pfizer monkey virus isolated from rhesus monkeys. Purified preparations of both viruses grown in heterologous cells, as well as extracts of heterologous cells infected with squirrel monkey retrovirus or Mason-Pfizer monkey virus, compete completely in the assay. Addition of an endogenous virus of the langur monkey also results in complete blocking. No blocking in the assay is observedmore » with type-C baboon viruses, woolly monkey virus, and gibbon virus. Various other type-C and type-B viruses also showed no reactivity. An interspecies assay has thus been developed that recognizes the type-D retroviruses from both Old World monkey (rhesus and langur) and New World monkey (squirrel) species.« less

New Phase of Growth for Xenogeneic-Based Bioartificial Organs

PubMed Central

Pitkin, Zorina

2016-01-01

In this article, we examine the advanced clinical development of bioartificial organs and describe the challenges to implementing such systems into patient care. The case for bioartificial organs is evident: they are meant to reduce patient morbidity and mortality caused by the persistent shortage of organs available for allotransplantation. The widespread introduction and adoption of bioengineered organs, incorporating cells and tissues derived from either human or animal sources, would help address this shortage. Despite the decades of development, the variety of organs studied and bioengineered, and continuous progress in the field, only two bioengineered systems are currently commercially available: Apligraf® and Dermagraft® are both approved by the FDA to treat diabetic foot ulcers, and Apligraf® is approved to treat venous leg ulcers. Currently, no products based on xenotransplantation have been approved by the FDA. Risk factors include immunological barriers and the potential infectivity of porcine endogenous retrovirus (PERV), which is unique to xenotransplantation. Recent breakthroughs in gene editing may, however, mitigate risks related to PERV. Because of its primary role in interrupting progress in xenotransplantation, we present a risk assessment for PERV infection, and conclude that the formerly high risk has been reduced to a moderate level. Advances in gene editing, and more broadly in the field, may make it more likely than ever before that bioartificial organs will alleviate the suffering of patients with organ failure. PMID:27657057

Treatment against human endogenous retrovirus: a possible personalized medicine approach for multiple sclerosis.

PubMed

Curtin, François; Perron, Hervé; Faucard, Raphael; Porchet, Hervé; Lang, Alois B

2015-10-01

Human endogenous retroviruses (HERV) represent about 8 % of the human genome. Some of these genetic elements are expressed in pathological circumstances. A HERV protein, the multiple sclerosis-associated retrovirus (MSRV) envelope protein (MSRV-Env), is expressed in the blood and active brain lesions of multiple sclerosis (MS) patients. It possesses pro-inflammatory and myelinotoxic properties. The patterns of expression and pathogenic properties of MSRV-Env make it a relevant drug target for MS therapeutics-in particular for preventing neurodegeneration, a key component of progressive forms of MS. An immunoglobulin G4 monoclonal antibody (mAb), called GNbAC1, has been developed to neutralize this pathogenic target. After showing neutralizing effects in vitro and in mouse models of MS, GNbAC1 is now in phase II clinical development. MSRV-related biomarkers such as MSRV-Env and MSRV polymerase (MSRV-Pol) gene transcripts are overexpressed in the blood and cerebrospinal fluid of patients with MS. These biomarkers may have prognostic value for long-term MS evolution, and their transcription levels in blood decline during treatments with GNbAC1, which has also been reported in patients administered reference MS drugs such as natalizumab or interferon-β. GNbAC1 as a new MSRV-Env-antagonist mAb could be a specific and causal treatment for MS, with a particular application for progressive forms of the disease. For possible use in companion diagnostic tests, MSRV-associated biomarkers could open the door to a personalized therapeutic approach for MS.

The impact of p53 on the early stage replication of retrovirus.

PubMed

Kinnetz, Michaela; Alghamdi, Faris; Racz, Michael; Hu, Wenwei; Shi, Binshan

2017-08-09

The function of p53 in cancer biology has been studied extensively, but its role in anti-retrovirus infection has been elusive for many years. The restriction of retrovirus early stage replication by p53 was investigated in this study. VSV-G pseudotyped retrovirus with GFP reporter gene was used to infect both HCT116 p53 +/+ cells and its isogenic p53 knockout HCT116 p53 -/- cells. The infection was detected by flow cytometry. Reverse transcription products were quantified by real time PCR. Mutation analysis was performed after 1-LTR cycle and 2-LTR cycle DNA were amplified and PCR products were sequenced. Transcription and translation of cyclin-dependent kinase inhibitor 1 (p21 Cip1 ) and SAM domain and HD domain-containing protein 1 (SAMHD1) were analyzed by TaqMan PCR and Western blot experiments. siRNA experiment was applied to study the role of p53 downstream gene p21 Cip1 in the restriction of retrovirus infection. It was found that the block of retrovirus infection in non-cycling cells was significantly attenuated in HCT116 p53 -/- cells when compared to HCT116 p53 +/+ cells. It was found that both late reverse transcription products and viral 2-LTR cycle DNA were significantly increased in infected non-cycling HCT116 p53 -/- cells. Furthermore, the mutation frequency detected in 1-LTR DNA from HCT116 p53 +/+ cells were significantly decreased in comparison to HCT116 p53 -/- cells. A higher number of insertion and deletion mutations were detected in the joint region of 2-LTR cycle DNA in infected p53 +/+ cells. Cell cycle analysis showed retrovirus infection promoted host cell replication. Higher levels of mRNA and protein of p21 Cip1 were found in HCT116 p53 +/+ cells in comparison to the HCT116 p53 -/- cells. Furthermore, knockdown of p21 Cip1 in non-cycling HCT116 p53 +/+ cells significantly increased the infection. The results of this study showed that p53 is an important restriction factor that interferes with retrovirus infection in its early stage of

Synthesis and assembly of retrovirus Gag precursors into immature capsids in vitro.

PubMed Central

Sakalian, M; Parker, S D; Weldon, R A; Hunter, E

1996-01-01

The assembly of retroviral particles is mediated by the product of the gag gene; no other retroviral gene products are necessary for this process. While most retroviruses assemble their capsids at the plasma membrane, viruses of the type D class preassemble immature capsids within the cytoplasm of infected cells. This has allowed us to determine whether immature capsids of the prototypical type D retrovirus, Mason-Pfizer monkey virus (M-PMV), can assemble in a cell-free protein synthesis system. We report here that assembly of M-PMV Gag precursor proteins can occur in this in vitro system. Synthesized particles sediment in isopycnic gradients to the appropriate density and in thin-section electron micrographs have a size and appearance consistent with those of immature retrovirus capsids. The in vitro system described in this report appears to faithfully mimic the process of assembly which occurs in the host cell cytoplasm, since M-PMV gag mutants defective in in vivo assembly also fail to assemble in vitro. Likewise, the Gag precursor proteins of retroviruses that undergo type C morphogenesis, Rous sarcoma virus and human immunodeficiency virus, which do not preassemble capsids in vivo, fail to assemble particles in this system. Additionally, we demonstrate, with the use of anti-Gag antibodies, that this cell-free system can be utilized for analysis in vitro of potential inhibitors of retrovirus assembly. PMID:8648705

A novel approach to achieving modular retrovirus clearance for a parvovirus filter.

PubMed

Stuckey, Juliana; Strauss, Daniel; Venkiteshwaran, Adith; Gao, Jinxin; Luo, Wen; Quertinmont, Michelle; O'Donnell, Sean; Chen, Dayue

2014-01-01

Viral filtration is routinely incorporated into the downstream purification processes for the production of biologics produced in mammalian cell cultures (MCC) to remove potential viral contaminants. In recent years, the use of retentive filters designed for retaining parvovirus (~20 nm) has become an industry standard in a conscious effort to further improve product safety. Since retentive filters remove viruses primarily by the size exclusion mechanism, it is expected that filters designed for parvovirus removal can effectively clear larger viruses such as retroviruses (~100 nm). In an attempt to reduce the number of viral clearance studies, we have taken a novel approach to demonstrate the feasibility of claiming modular retrovirus clearance for Asahi Planova 20N filters. Porcine parvovirus (PPV) and xenotropic murine leukemia virus (XMuLV) were co-spiked into six different feedstreams and then subjected to laboratory scale Planova 20N filtration. Our results indicate that Planova 20N filters consistently retain retroviruses and no retrovirus has ever been detected in the filtrates even when significant PPV breakthrough is observed. Based on the data from multiple in-house viral validation studies and the results from the co-spiking experiments, we have successfully claimed a modular retrovirus clearance of greater than 6 log10 reduction factors (LRF) to support clinical trial applications in both USA and Europe. © 2013 American Institute of Chemical Engineers.

Retrovirus maturation-an extraordinary structural transformation.

PubMed

Mattei, Simone; Schur, Florian Km; Briggs, John Ag

2016-06-01

Retroviruses such as HIV-1 assemble and bud from infected cells in an immature, non-infectious form. Subsequently, a series of proteolytic cleavages catalysed by the viral protease leads to a spectacular structural rearrangement of the viral particle into a mature form that is competent to fuse with and infect a new cell. Maturation involves changes in the structures of protein domains, in the interactions between protein domains, and in the architecture of the viral components that are assembled by the proteins. Tight control of proteolytic cleavages at different sites is required for successful maturation, and the process is a major target of antiretroviral drugs. Here we will describe what is known about the structures of immature and mature retrovirus particles, and about the maturation process by which one transitions into the other. Despite a wealth of available data, fundamental questions about retroviral maturation remain unanswered. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Innate immunity in the pathogenesis of polytropic retrovirus infection in the central nervous system.

PubMed

Peterson, Karin E; Du, Min

2009-01-01

Neuroinflammation, including astrogliosis, microgliosis, and the production of proinflammatory cytokines and chemokines is a common response in the central nervous system (CNS) to virus infection, including retrovirus infection. However, the contribution of this innate immune response in disease pathogenesis remains unresolved. Analysis of the neuroinflammatory response to polytropic retrovirus infection in the mouse has provided insight into the potential contribution of the innate immune response to retrovirus-induced neurologic disease. In this model, retroviral pathogenesis correlates with the induction of neuroinflammatory responses including the activation of astrocytes and microglia, as well as the production of proinflammatory cytokines and chemokines. Studies of the neurovirulent determinants of the polytropic envelope protein as well as studies with knockout mice suggest that retroviral pathogenesis in the brain is multifaceted and that cytokine and chemokine production may be only one mechanism of disease pathogenesis. Analysis of the activation of the innate immune response to retrovirus infection in the CNS indicates that toll-like receptor 7 (TLR7) is a contributing factor to retrovirus-induced neuroinflammation, but that other factors can compensate for the lack of TLR7 in inducing both neuroinflammation and neurologic disease.

Ubiquitin is part of the retrovirus budding machinery

NASA Astrophysics Data System (ADS)

Patnaik, Akash; Chau, Vincent; Wills, John W.

2000-11-01

Retroviruses contain relatively large amounts of ubiquitin, but the significance of this finding has been unknown. Here, we show that drugs that are known to reduce the level of free ubiquitin in the cell dramatically reduced the release of Rous sarcoma virus, an avian retrovirus. This effect was suppressed by overexpressing ubiquitin and also by directly fusing ubiquitin to the C terminus of Gag, the viral protein that directs budding and particle release. The block to budding was found to be at the plasma membrane, and electron microscopy revealed that the reduced level of ubiquitin results in a failure of mature virus particles to separate from each other and from the plasma membrane during budding. These data indicate that ubiquitin is actually part of the budding machinery.

Sequence of retrovirus provirus resembles that of bacterial transposable elements

NASA Astrophysics Data System (ADS)

Shimotohno, Kunitada; Mizutani, Satoshi; Temin, Howard M.

1980-06-01

The nucleotide sequences of the terminal regions of an infectious integrated retrovirus cloned in the modified λ phage cloning vector Charon 4A have been elucidated. There is a 569-base pair direct repeat at both ends of the viral DNA. The cell-virus junctions at each end consist of a 5-base pair direct repeat of cell DNA next to a 3-base pair inverted repeat of viral DNA. This structure resembles that of a transposable element and is consistent with the protovirus hypothesis that retroviruses evolved from the cell genome.

Human endogenous retrovirus W in brain lesions: Rationale for targeted therapy in multiple sclerosis.

PubMed

van Horssen, Jack; van der Pol, Susanne; Nijland, Philip; Amor, Sandra; Perron, Hervé

2016-07-01

Attempts to identify a causative agent of Multiple Sclerosis (MS) among environmental viruses have consistently failed suggesting that development of MS is a result from gene-environment interactions. A new pathogenic player within human genes, a human endogenous retrovirus (HERV) was identified from MS cells, named MS-associated retrovirus element (MSRV) and unveiled homologous multicopy HERVs (HERV-W). As independent studies revealed biological features of HERV-W on immune-mediated inflammation and on remyelinating cells, the present study characterized the presence of HERV-W envelope protein (MSRV-Env) at the cellular level, in different MS lesion stages to extend and validate previous studies. Immunohistological analysis of HERV-W envelope cellular expression in different lesion stages from a cohort of MS brains versus controls, using well-characterized and highly specific monoclonal antibodies. HERV-W envelope protein was detected in all MS brains and quite essentially in lesions. Immunohistochemistry showed dominant expression in macrophages and microglia, coinciding with areas of active demyelination, spread over the active lesions, or limited to the rim of active microglia in chronic active lesions or in few surviving astrocytes of inactive plaques. Weak expression was seen in MS normal appearing white matter. In active plaques, few lymphoid cells and astrocytes were also stained. This HERV-W expression was not observed in control brains. HERV-W was expressed in demyelinated lesions from MS brains, which were all positive for this endogenous pathogenic protein. Pronounced HERV-W immunoreactivity in active MS lesions was intimately associated with areas of active demyelination throughout the successive stages of lesion evolution in MS brains. Based on its pathogenic potential, this HERV-W (MSRV) endogenous toxin thus appears to be a novel therapeutic target in MS. It also has a unique positioning as an early and lifelong expressed pathogenic agonist, acting

Loss of retrovirus production in JB/RH melanoma cells transfected with H-2Kb and TAP-1 genes.

PubMed

Li, M; Xu, F; Muller, J; Huang, X; Hearing, V J; Gorelik, E

1999-01-20

JB/RH1 melanoma cells, as well as other melanomas of C57BL/6 mice (B16 and JB/MS), express a common melanoma-associated antigen (MAA) encoded by an ecotropic melanoma-associated retrovirus (MelARV). JB/RH1 cells do not express the H-2Kb molecules due to down-regulation of the H-2Kb and TAP-1 genes. When JB/RH1 cells were transfected with the H-2Kb and cotransfected with the TAP-1 gene, it resulted in the appearance of H-2Kb molecules and an increase in their immunogenicity, albeit they lost expression of retrovirus-encoded MAA recognized by MM2-9B6 mAb. Loss of MAA was found to result from a complete and stable elimination of ecotropic MelARV production in the H-2Kb/TAP-1-transfected JB/RH1 cells. Northern blot analysis showed no differences in ecotropic retroviral messages in MelARV-producing and -nonproducing melanoma cells, suggesting that loss of MelARV production was not due to down-regulation of MelARV transcription. Southern blot analysis revealed several rearrangements in the proviral DNA of H-2Kb-positive JB/RH1 melanoma cells. Sequence analysis of the ecotropic proviral DNA from these cells showed numerous nucleotide substitutions, some of which resulted in the appearance of a novel intraviral PstI restriction site and the loss of a HindIII restriction site in the pol region. PCR amplification of the proviral DNAs indicates that an ecotropic provirus found in the H-2Kb-positive cells is novel and does not preexist in the parental H-2Kb-negative melanoma cells. Conversely, the ecotropic provirus of the parental JB/RH1 cells was not amplifable from the H-2Kb-positive cells. Our data indicate that stable loss of retroviral production in the H-2Kb/TAP-1-transfected melanoma cells is probably due to the induction of recombination between a productive ecotropic MelARV and a defective nonecotropic provirus leading to the generation of a defective ecotropic provirus and the loss of MelARV production and expression of the retrovirus-encoded MAA. Copyright 1999

Myeloid-derived suppressor cells in murine retrovirus-induced AIDS inhibit T- and B-cell responses in vitro that are used to define the immunodeficiency.

PubMed

Green, Kathy A; Cook, W James; Green, William R

2013-02-01

Myeloid-derived suppressor cells (MDSCs) have been characterized in several disease settings, especially in many tumor systems. Compared to their involvement in tumor microenvironments, however, MDSCs have been less well studied in their responses to infectious disease processes, in particular to retroviruses that induce immunodeficiency. Here, we demonstrate for the first time the development of a highly immunosuppressive MDSC population that is dependent on infection by the LP-BM5 retrovirus, which causes murine acquired immunodeficiency. These MDSCs express a cell surface marker signature (CD11b(+) Gr-1(+) Ly6C(+)) characteristic of monocyte-type MDSCs. Such MDSCs profoundly inhibit immune responsiveness by a cell dose- and substantially inducible nitric oxide synthase (iNOS)-dependent mechanism that is independent of arginase activity, PD-1-PD-L1 expression, and interleukin 10 (IL-10) production. These MDSCs display levels of immunosuppressive function in parallel with the extent of disease in LP-BM5-infected wild-type (w.t.) versus knockout mouse strains that are differentially susceptible to pathogenesis. These MDSCs suppressed not only T-cell but also B-cell responses, which are an understudied target for MDSC inhibition. The MDSC immunosuppression of B-cell responses was confirmed by the use of purified B responder cells, multiple B-cell stimuli, and independent assays measuring B-cell expansion. Retroviral load measurements indicated that the suppressive Ly6G(low/±) Ly6C(+) CD11b(+)-enriched MDSC subset was positive for LP-BM5, albeit at a significantly lower level than that of nonfractionated splenocytes from LP-BM5-infected mice. These results, including the strong direct MDSC inhibition of B-cell responsiveness, are novel for murine retrovirus-induced immunosuppression and, as this broadly suppressive function mirrors that of the LP-BM5-induced disease syndrome, support a possible pathogenic effector role for these retrovirus-induced MDSCs.

The Retrovirus pol Gene Encodes a Product Required for DNA Integration: Identification of a Retrovirus int Locus

NASA Astrophysics Data System (ADS)

Panganiban, Antonito T.; Temin, Howard M.

1984-12-01

We mutagenized cloned spleen necrosis virus DNA to identify a region of the retrovirus genome encoding a polypeptide required for integration of viral DNA. Five plasmids bearing different lesions in the 3' end of the pol gene were examined for the ability to integrate or replicate following transfection of chicken embryo fibroblasts. Transfection with one of these DNAs resulted in the generation of mutant virus incapable of integrating but able to replicate at low levels; this phenotype is identical to that of mutants bearing alterations in the cis-acting region, att. To determine whether the 3' end of the pol gene encodes a protein that interacts with att, we did a complementation experiment. Cells were first infected with an att- virus and then superinfected with the integration-deficient virus containing a lesion in the pol gene and a wild-type att site. The results showed that the att- virus provided a trans-acting function allowing integration of viral DNA derived from the mutant bearing a wild-type att site. Thus, the 3' end of the pol gene serves as an ``int'' locus and encodes a protein mediating integration of retrovirus DNA through interaction with att.

Different modes of retrovirus restriction by human APOBEC3A and APOBEC3G in vivo.

PubMed

Stavrou, Spyridon; Crawford, Daniel; Blouch, Kristin; Browne, Edward P; Kohli, Rahul M; Ross, Susan R

2014-05-01

The apolipoprotein B editing complex 3 (A3) cytidine deaminases are among the most highly evolutionarily selected retroviral restriction factors, both in terms of gene copy number and sequence diversity. Primate genomes encode seven A3 genes, and while A3F and 3G are widely recognized as important in the restriction of HIV, the role of the other genes, particularly A3A, is not as clear. Indeed, since human cells can express multiple A3 genes, and because of the lack of an experimentally tractable model, it is difficult to dissect the individual contribution of each gene to virus restriction in vivo. To overcome this problem, we generated human A3A and A3G transgenic mice on a mouse A3 knockout background. Using these mice, we demonstrate that both A3A and A3G restrict infection by murine retroviruses but by different mechanisms: A3G was packaged into virions and caused extensive deamination of the retrovirus genomes while A3A was not packaged and instead restricted infection when expressed in target cells. Additionally, we show that a murine leukemia virus engineered to express HIV Vif overcame the A3G-mediated restriction, thereby creating a novel model for studying the interaction between these proteins. We have thus developed an in vivo system for understanding how human A3 proteins use different modes of restriction, as well as a means for testing therapies that disrupt HIV Vif-A3G interactions.

Discovery of a novel retrovirus sequence in an Australian native rodent (Melomys burtoni): a putative link between gibbon ape leukemia virus and koala retrovirus.

PubMed

Simmons, Greg; Clarke, Daniel; McKee, Jeff; Young, Paul; Meers, Joanne

2014-01-01

Gibbon ape leukaemia virus (GALV) and koala retrovirus (KoRV) share a remarkably close sequence identity despite the fact that they occur in distantly related mammals on different continents. It has previously been suggested that infection of their respective hosts may have occurred as a result of a species jump from another, as yet unidentified vertebrate host. To investigate possible sources of these retroviruses in the Australian context, DNA samples were obtained from 42 vertebrate species and screened using PCR in order to detect proviral sequences closely related to KoRV and GALV. Four proviral partial sequences totalling 2880 bases which share a strong similarity with KoRV and GALV were detected in DNA from a native Australian rodent, the grassland melomys, Melomys burtoni. We have designated this novel gammaretrovirus Melomys burtoni retrovirus (MbRV). The concatenated nucleotide sequence of MbRV shares 93% identity with the corresponding sequence from GALV-SEATO and 83% identity with KoRV. The geographic ranges of the grassland melomys and of the koala partially overlap. Thus a species jump by MbRV from melomys to koalas is conceivable. However the genus Melomys does not occur in mainland South East Asia and so it appears most likely that another as yet unidentified host was the source of GALV.

Human endogenous retrovirus W env increases nitric oxide production and enhances the migration ability of microglia by regulating the expression of inducible nitric oxide synthase.

PubMed

Xiao, Ran; Li, Shan; Cao, Qian; Wang, Xiuling; Yan, Qiujin; Tu, Xiaoning; Zhu, Ying; Zhu, Fan

2017-06-01

Human endogenous retrovirus W env (HERV-W env) plays a critical role in many neuropsychological diseases such as schizophrenia and multiple sclerosis (MS). These diseases are accompanied by immunological reactions in the central nervous system (CNS). Microglia are important immunocytes in brain inflammation that can produce a gasotransmitter-nitric oxide (NO). NO not only plays a role in the function of neuronal cells but also participates in the pathogenesis of various neuropsychological diseases. In this study, we reported increased NO production in CHME-5 microglia cells after they were transfected with HERV-W env. Moreover, HERV-W env increased the expression and function of human inducible nitric oxide synthase (hiNOS) and enhanced the promoter activity of hiNOS. Microglial migration was also enhanced. These data revealed that HERV-W env might contribute to increase NO production and microglial migration ability in neuropsychological disorders by regulating the expression of inducible NOS. Results from this study might lead to the identification of novel targets for the treatment of neuropsychological diseases, including neuroinflammatory diseases, stroke, and neurodegenerative diseases.

Identification of an ancient endogenous retrovirus, predating the divergence of the placental mammals.

PubMed

Lee, Adam; Nolan, Alison; Watson, Jason; Tristem, Michael

2013-09-19

The evolutionary arms race between mammals and retroviruses has long been recognized as one of the oldest host-parasite interactions. Rapid evolution rates in exogenous retroviruses have often made accurate viral age estimations highly problematic. Endogenous retroviruses (ERVs), however, integrate into the germline of their hosts, and are subjected to their evolutionary rates. This study describes, for the first time, a retroviral orthologue predating the divergence of placental mammals, giving it a minimum age of 104-110 Myr. Simultaneously, other orthologous selfish genetic elements (SGEs), inserted into the ERV sequence, provide evidence for the oldest individual mammalian-wide interspersed repeat and medium-reiteration frequency interspersed repeat mammalian repeats, with the same minimum age. The combined use of shared SGEs and reconstruction of viral orthologies defines new limits and increases maximum 'lookback' times, with subsequent implications for the field of paleovirology.

Cell-to-cell transmission of retroviruses: Innate immunity and interferon-induced restriction factors

PubMed Central

Jolly, Clare

2011-01-01

It has been known for some time that retroviruses can disseminate between immune cells either by conventional cell-free transmission or by directed cell-to-cell spread. Over the past few years there has been increasing interest in how retroviruses may use cell-to-cell spread to promote more rapid infection kinetics and circumvent humoral immunity. Effective humoral immune responses are intimately linked with innate immunity and the interplay between retroviruses and innate immunity is a rapidly expanding area of research that has been advanced considerably by the identification of cellular restriction factors that provide barriers to retroviral infection. The effect of innate immunity and restriction factors on retroviral cell-to-cell spread has been comparatively little studied; however recent work suggests this maybe changing. Here I will review some recent advances in what is a budding area of retroviral research. PMID:21247613

SAMHD1 knockout mice: modeling retrovirus restriction in vivo.

PubMed

Wu, Li

2013-11-20

The host dNTP hydrolase SAMHD1 acts as a viral restriction factor to inhibit the replication of several retroviruses and DNA viruses in non-cycling human immune cells. However, understanding the physiological role of mammalian SAMHD1 has been elusive due to the lack of an animal model. Two recent studies reported the generation of samhd1 knockout mouse models for investigating the restriction of HIV-1 vectors and endogenous retroviruses in vivo. Both studies suggest that SAMHD1 is important for regulating the intracellular dNTP pool and the intrinsic immunity against retroviral infection, despite different outcomes of HIV-1 vector transduction in these mouse models. Here I discuss the significance of these new findings and the future directions in studying SAMHD1-mediated retroviral restriction.

LTRs of Endogenous Retroviruses as a Source of Tbx6 Binding Sites.

PubMed

Yasuhiko, Yukuto; Hirabayashi, Yoko; Ono, Ryuichi

2017-01-01

Retrotransposons are abundant in mammalian genomes and can modulate the gene expression of surrounding genes by disrupting endogenous binding sites for transcription factors (TFs) or providing novel TFs binding sites within retrotransposon sequences. Here, we show that a (C/T)CACACCT sequence motif in ORR1A, ORR1B, ORR1C, and ORR1D, Long Terminal Repeats (LTRs) of MaLR endogenous retrovirus (ERV), is the direct target of Tbx6, an evolutionary conserved family of T-box TFs. Moreover, by comparing gene expression between control mice (Tbx6 +/-) and Tbx6-deficient mice (Tbx6 -/-), we demonstrate that at least four genes, Twist2, Pitx2, Oscp1 , and Nfxl1 , are down-regulated with Tbx6 deficiency. These results suggest that ORR1A, ORR1B, ORR1C and ORR1D may contribute to the evolution of mammalian embryogenesis.

LTRs of endogenous retroviruses as a source of Tbx6 binding sites

NASA Astrophysics Data System (ADS)

Yasuhiko, Yukuto; Hirabayashi, Yoko; Ono, Ryuichi

2017-06-01

Retrotransposons are abundant in mammalian genomes and can modulate the gene expression of surrounding genes by disrupting endogenous binding sites for transcription factors (TFs) or providing novel TFs binding sites within retrotransposon sequences. Here, we show that a (C/T)CACACCT sequence motif in ORR1A, ORR1B, ORR1C and ORR1D, Long Terminal Repeats (LTRs) of MaLR endogenous retrovirus (ERV), is the direct target of Tbx6, an evolutionary conserved family of T-box transcription factors. Moreover, by comparing gene expression between control mice (Tbx6 +/-) and Tbx6-deficient mice (Tbx6 -/-), we demonstrate that at least four genes, Twist2, Pitx2, Oscp1, and Nfxl1, are down-regulated with Tbx6 deficiency. These results suggest that ORR1A, ORR1B, ORR1C and ORR1D may contribute to the evolution of mammalian embryogenesis.

Characterization and Complete Nucleotide Sequence of an Unusual Reptilian Retrovirus Recovered from the Order Crocodylia

PubMed Central

Martin, Joanne; Kabat, Peter; Herniou, Elisabeth; Tristem, Michael

2002-01-01

A novel group of retroviruses found within the order Crocodylia are described. Phylogenetic analyses demonstrate that they are probably the most divergent members of the Retroviridae described to date; even the most conserved regions of Pol show an average of only 23% amino acid identity when compared to other retroviruses. PMID:11932432

A computational model for predicting integrase catalytic domain of retrovirus.

PubMed

Wu, Sijia; Han, Jiuqiang; Zhang, Xinman; Zhong, Dexing; Liu, Ruiling

2017-06-21

Integrase catalytic domain (ICD) is an essential part in the retrovirus for integration reaction, which enables its newly synthesized DNA to be incorporated into the DNA of infected cells. Owing to the crucial role of ICD for the retroviral replication and the absence of an equivalent of integrase in host cells, it is comprehensible that ICD is a promising drug target for therapeutic intervention. However, annotated ICDs in UniProtKB database have still been insufficient for a good understanding of their statistical characteristics so far. Accordingly, it is of great importance to put forward a computational ICD model in this work to annotate these domains in the retroviruses. The proposed model then discovered 11,660 new putative ICDs after scanning sequences without ICD annotations. Subsequently in order to provide much confidence in ICD prediction, it was tested under different cross-validation methods, compared with other database search tools, and verified on independent datasets. Furthermore, an evolutionary analysis performed on the annotated ICDs of retroviruses revealed a tight connection between ICD and retroviral classification. All the datasets involved in this paper and the application software tool of this model can be available for free download at https://sourceforge.net/projects/icdtool/files/?source=navbar. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Computational Model for Predicting RNase H Domain of Retrovirus.

PubMed

Wu, Sijia; Zhang, Xinman; Han, Jiuqiang

2016-01-01

RNase H (RNH) is a pivotal domain in retrovirus to cleave the DNA-RNA hybrid for continuing retroviral replication. The crucial role indicates that RNH is a promising drug target for therapeutic intervention. However, annotated RNHs in UniProtKB database have still been insufficient for a good understanding of their statistical characteristics so far. In this work, a computational RNH model was proposed to annotate new putative RNHs (np-RNHs) in the retroviruses. It basically predicts RNH domains through recognizing their start and end sites separately with SVM method. The classification accuracy rates are 100%, 99.01% and 97.52% respectively corresponding to jack-knife, 10-fold cross-validation and 5-fold cross-validation test. Subsequently, this model discovered 14,033 np-RNHs after scanning sequences without RNH annotations. All these predicted np-RNHs and annotated RNHs were employed to analyze the length, hydrophobicity and evolutionary relationship of RNH domains. They are all related to retroviral genera, which validates the classification of retroviruses to a certain degree. In the end, a software tool was designed for the application of our prediction model. The software together with datasets involved in this paper can be available for free download at https://sourceforge.net/projects/rhtool/files/?source=navbar.

ROCK1 and LIM kinase modulate retrovirus particle release and cell-cell transmission events.

PubMed

Wen, Xiaoyun; Ding, Lingmei; Wang, Jaang-Jiun; Qi, Mingli; Hammonds, Jason; Chu, Hin; Chen, Xuemin; Hunter, Eric; Spearman, Paul

2014-06-01

The assembly and release of retroviruses from the host cells require dynamic interactions between viral structural proteins and a variety of cellular factors. It has been long speculated that the actin cytoskeleton is involved in retrovirus production, and actin and actin-related proteins are enriched in HIV-1 virions. However, the specific role of actin in retrovirus assembly and release remains unknown. Here we identified LIM kinase 1 (LIMK1) as a cellular factor regulating HIV-1 and Mason-Pfizer monkey virus (M-PMV) particle release. Depletion of LIMK1 reduced not only particle output but also virus cell-cell transmission and was rescued by LIMK1 replenishment. Depletion of the upstream LIMK1 regulator ROCK1 inhibited particle release, as did a competitive peptide inhibitor of LIMK1 activity that prevented cofilin phosphorylation. Disruption of either ROCK1 or LIMK1 led to enhanced particle accumulation on the plasma membrane as revealed by total internal reflection fluorescence microscopy (TIRFM). Electron microscopy demonstrated a block to particle release, with clusters of fully mature particles on the surface of the cells. Our studies support a model in which ROCK1- and LIMK1-regulated phosphorylation of cofilin and subsequent local disruption of dynamic actin turnover play a role in retrovirus release from host cells and in cell-cell transmission events. Viruses often interact with the cellular cytoskeletal machinery in order to deliver their components to the site of assembly and budding. This study indicates that a key regulator of actin dynamics at the plasma membrane, LIM kinase, is important for the release of viral particles for HIV as well as for particle release by a distantly related retrovirus, Mason-Pfizer monkey virus. Moreover, disruption of LIM kinase greatly diminished the spread of HIV from cell to cell. These findings suggest that LIM kinase and its dynamic modulation of the actin cytoskeleton in the cell may be an important host factor for

ENDOGENOUS RETROVIRUSES MOBILIZED DURING FRIEND MURINE LEUKEMIA VIRUS INFECTION

PubMed Central

Hansen, Ethan; Hendrick, Duncan; Malik, Frank; Evans, Leonard H.

2016-01-01

We have demonstrated in a mouse model that infection with a retrovirus can lead not only to the generation of recombinants between exogenous and endogenous gammaretrovirus, but also to the mobilization of endogenous proviruses by pseudotyping entire polytropic proviral transcripts and facilitating their infectious spread to new cells. However, the frequency of this occurrence, the kinetics, and the identity of mobilized endogenous proviruses was unclear. Here we find that these mobilized transcripts are detected after only one day of infection. They predominate over recombinant polytropic viruses early in infection, persist throughout the course of disease and are comprised of multiple different polytropic proviruses. Other endogenous retroviral elements such as intracisternal A particles (IAPs) were not detected. The integration of the endogenous transcripts into new cells could result in loss of transcriptional control and elevated expression which may facilitate pathogenesis, perhaps by contributing to the generation of polytropic recombinant viruses. PMID:27657834

Human endogenous retroviruses and cancer prevention: evidence and prospects.

PubMed

Cegolon, Luca; Salata, Cristiano; Weiderpass, Elisabete; Vineis, Paolo; Palù, Giorgio; Mastrangelo, Giuseppe

2013-01-03

Cancer is a significant and growing problem worldwide. While this increase may, in part, be attributed to increasing longevity, improved case notifications and risk-enhancing lifestyle (such as smoking, diet and obesity), hygiene-related factors resulting in immuno-regulatory failure may also play a major role and call for a revision of vaccination strategies to protect against a range of cancers in addition to infections. Human endogenous retroviruses (HERVs) are a significant component of a wider family of retroelements that constitutes part of the human genome. They were originated by the integration of exogenous retroviruses into the human genome millions of years ago. HERVs are estimated to comprise about 8% of human DNA and are ubiquitous in somatic and germinal tissues.Physiologic and pathologic processes are influenced by some biologically active HERV families. HERV antigens are only expressed at low levels by the host, but in circumstances of inappropriate control their genes may initiate or maintain pathological processes. Although the precise mechanism leading to abnormal HERVs gene expression has yet to be clearly elucidated, environmental factors seem to be involved by influencing the human immune system.HERV-K expression has been detected in different types of tumors.Among the various human endogenous retroviral families, the K series was the latest acquired by the human species. Probably because of its relatively recent origin, the HERV-K is the most complete and biologically active family.The abnormal expression of HERV-K seemingly triggers pathological processes leading to melanoma onset, but also contributes to the morphological and functional cellular modifications implicated in melanoma maintenance and progression.The HERV-K-MEL antigen is encoded by a pseudo-gene incorporated in the HERV-K env-gene. HERV-K-MEL is significantly expressed in the majority of dysplastic and normal naevi, as well as other tumors like sarcoma, lymphoma, bladder and

World Reference Center for Arboviruses and Retroviruses

DTIC Science & Technology

1988-05-01

Reference Center for Arboviruses and Retroviruses identified viruses from Thailand, Nepal, Egypt, Colombia, and Panama. Cache Valley virus from a recruit...ARBOVIRUSES . . 13 A. Study of viruses from Thailand and Nepal . . . . 13 B. Isolation of Sicilian sandfly fever virus from Egyptian phlebotomines...dengue viruses ..... . 30 VII. LOW PASSAGE VIRUS COLLECTION .... ............. 32 VIII. ARBOVIRUS BULLETIN BOARD, REFERENCE, AND DATA ACCESS . 32 IX

Human endogenous retroviruses and cancer: causality and therapeutic possibilities.

PubMed

Mullins, Christina S; Linnebacher, Michael

2012-11-14

A substantial part of the human genome is derived from transposable elements; remnants of ancient retroviral infections. Conservative estimates set the percentage of human endogenous retroviruses (HERVs) in the genome at 8%. For the most part, the interplay between mutations, epigenetic mechanisms and posttranscriptional regulations silence HERVs in somatic cells. We first highlight mechanisms by which activation of members of several HERV families may be associated with tumor development before discussing the arising chances for both diagnosis and therapy. It has been shown that at least in some cases, tumor cells expressing HERV open reading frames (ORFs) thus gain tumor-promoting functions. However, since these proteins are not expressed in healthy tissues, they become prime target structures. Of potential pharmacological interest are the prevention of HERV transposition, the inhibition of HERV-encoded protein expression and the interference with these proteins' activities. Evidence from recent studies unequivocally proves that HERV ORFs represent a very interesting source of novel tumor-specific antigens with even the potential to surpass entity boundaries. The development of new tumor (immune-) therapies is a very active field and true tumor-specific targets are of outstanding interest since they minimize the risk of autoimmunity and could reduce side effects. Finally, we postulate on main future research streams in order to stimulate discussion on this hot topic.

Genomic Flexibility of Human Endogenous Retrovirus Type K

PubMed Central

Dube, Derek; Contreras-Galindo, Rafael; He, Shirley; King, Steven R.; Gonzalez-Hernandez, Marta J.; Gitlin, Scott D.; Kaplan, Mark H.

2014-01-01

ABSTRACT Human endogenous retrovirus type K (HERV-K) proviruses are scattered throughout the human genome, but as no infectious HERV-K virus has been detected to date, the mechanism by which these viruses replicated and populated the genome remains unresolved. Here, we provide evidence that, in addition to the RNA genomes that canonical retroviruses package, modern HERV-K viruses can contain reverse-transcribed DNA (RT-DNA) genomes. Indeed, reverse transcription of genomic HERV-K RNA into the DNA form is able to occur in three distinct times and locations: (i) in the virus-producing cell prior to viral release, yielding a DNA-containing extracellular virus particle similar to the spumaviruses; (ii) within the extracellular virus particle itself, transitioning from an RNA-containing particle to a DNA-containing particle; and (iii) after entry of the RNA-containing virus into the target cell, similar to canonical retroviruses, such as murine leukemia virus and HIV. Moreover, using a resuscitated HERV-K virus construct, we show that both viruses with RNA genomes and viruses with DNA genomes are capable of infecting target cells. This high level of genomic flexibility historically could have permitted these viruses to replicate in various host cell environments, potentially assisting in their many integration events and resulting in their high prevalence in the human genome. Moreover, the ability of modern HERV-K viruses to proceed through reverse transcription and package RT-DNA genomes suggests a higher level of replication competency than was previously understood, and it may be relevant in HERV-K-associated human diseases. IMPORTANCE Retroviral elements comprise at least 8% of the human genome. Of all the endogenous retroviruses, HERV-K viruses are the most intact and biologically active. While a modern infectious HERV-K has yet to be found, HERV-K activation has been associated with cancers, autoimmune diseases, and HIV-1 infection. Thus, determining how this

The mechanism of retrovirus suppression of human T cell proliferation in vitro.

PubMed

Copelan, E A; Rinehart, J J; Lewis, M; Mathes, L; Olsen, R; Sagone, A

1983-10-01

Immunosuppression is commonly associated with retrovirus-induced animal tumors. Studies in the murine and feline retrovirus systems suggest that the 15,000-dalton envelope protein (p15E) of the virion may contribute to immunosuppression by interfering with normal lymphocyte function. We examined the effect of inactivated feline leukemia virus (UV-FeLV) and p15E derived from this virus on concanavalin A (Con A) driven human T cell proliferation. Virus and p15E markedly suppressed mononuclear cell proliferative response to Con A. Suppression was not due to inhibition of monocyte accessory cell function, or interleukin 1 (IL 1) secretion. In fact, the presence of monocytes partially protected T cells from UV-FeLV suppression. UV-FeLV, however, suppressed T cell secretion of and response to interleukin 2 (IL 2). We conclude that UV-FeLV and derived p15E inhibit T cell proliferation by direct inhibition of T cell function. These findings, extended to the in vivo situations, suggest that retrovirus-associated suppression of the immune response involves the induction of T cell but not monocyte dysfunction.

Reassessment of murine APOBEC1 as a retrovirus restriction factor in vivo.

PubMed

Barrett, Bradley S; Guo, Kejun; Harper, Michael S; Li, Sam X; Heilman, Karl J; Davidson, Nicholas O; Santiago, Mario L

2014-11-01

APOBEC1 is a cytidine deaminase involved in cholesterol metabolism that has been linked to retrovirus restriction, analogous to the evolutionarily-related APOBEC3 proteins. In particular, murine APOBEC1 was shown to inhibit Friend retrovirus (FV) in vitro, generating high levels of C-to-T and G-to-A mutations. These observations raised the possibility that FV infection might be altered in APOBEC1-null mice. To examine this question directly, we infected wild-type and APOBEC1-null mice with FV complex and evaluated acute infection levels. Surprisingly, APOBEC1-null mice exhibited similar cellular infection levels and plasma viremia relative to wild-type mice. Moreover, next-generation sequencing analyses revealed that in contrast to APOBEC3, APOBEC1 did not enhance retroviral C-to-T and G-to-A mutational frequencies in genomic DNA. Thus, APOBEC1 neither inhibited nor significantly drove the molecular evolution of FV in vivo. Our findings reinforce that not all retrovirus restriction factors characterized as potent in vitro may be functionally relevant in vivo. Copyright © 2014 Elsevier Inc. All rights reserved.

Multiple sclerosis and human T-cell lymphotropic retroviruses

NASA Astrophysics Data System (ADS)

Koprowski, Hilary; Defreitas, Elaine C.; Harper, Mary E.; Sandberg-Wollheim, Magnhild; Sheremata, William A.; Robert-Guroff, Marjorie; Saxinger, Carl W.; Feinberg, Mark B.; Wong-Staal, Flossie; Gallo, Robert C.

1985-11-01

A combination of different types of data suggests that some multiple sclerosis patients respond immunologically to, and have cerebrospinal T cells containing, a retrovirus that is related to, but distinct from, the three types of human T-cell lymphotropic viruses. The role of this virus in multiple sclerosis is uncertain.

Human endogenous retrovirus K Gag coassembles with HIV-1 Gag and reduces the release efficiency and infectivity of HIV-1.

PubMed

Monde, Kazuaki; Contreras-Galindo, Rafael; Kaplan, Mark H; Markovitz, David M; Ono, Akira

2012-10-01

Human endogenous retroviruses (HERVs), which are remnants of ancestral retroviruses integrated into the human genome, are defective in viral replication. Because activation of HERV-K and coexpression of this virus with HIV-1 have been observed during HIV-1 infection, it is conceivable that HERV-K could affect HIV-1 replication, either by competition or by cooperation, in cells expressing both viruses. In this study, we found that the release efficiency of HIV-1 Gag was 3-fold reduced upon overexpression of HERV-K(CON) Gag. In addition, we observed that in cells expressing Gag proteins of both viruses, HERV-K(CON) Gag colocalized with HIV-1 Gag at the plasma membrane. Furthermore, HERV-K(CON) Gag was found to coassemble with HIV-1 Gag, as demonstrated by (i) processing of HERV-K(CON) Gag by HIV-1 protease in virions, (ii) coimmunoprecipitation of virion-associated HERV-K(CON) Gag with HIV-1 Gag, and (iii) rescue of a late-domain-defective HERV-K(CON) Gag by wild-type (WT) HIV-1 Gag. Myristylation-deficient HERV-K(CON) Gag localized to nuclei, suggesting cryptic nuclear trafficking of HERV-K Gag. Notably, unlike WT HERV-K(CON) Gag, HIV-1 Gag failed to rescue myristylation-deficient HERV-K(CON) Gag to the plasma membrane. Efficient colocalization and coassembly of HIV-1 Gag and HERV-K Gag also required nucleocapsid (NC). These results provide evidence that HIV-1 Gag heteromultimerizes with HERV-K Gag at the plasma membrane, presumably through NC-RNA interaction. Intriguingly, HERV-K Gag overexpression reduced not only HIV-1 release efficiency but also HIV-1 infectivity in a myristylation- and NC-dependent manner. Altogether, these results indicate that Gag proteins of endogenous retroviruses can coassemble with HIV-1 Gag and modulate the late phase of HIV-1 replication.

LTRs of Endogenous Retroviruses as a Source of Tbx6 Binding Sites

PubMed Central

Yasuhiko, Yukuto; Hirabayashi, Yoko; Ono, Ryuichi

2017-01-01

Retrotransposons are abundant in mammalian genomes and can modulate the gene expression of surrounding genes by disrupting endogenous binding sites for transcription factors (TFs) or providing novel TFs binding sites within retrotransposon sequences. Here, we show that a (C/T)CACACCT sequence motif in ORR1A, ORR1B, ORR1C, and ORR1D, Long Terminal Repeats (LTRs) of MaLR endogenous retrovirus (ERV), is the direct target of Tbx6, an evolutionary conserved family of T-box TFs. Moreover, by comparing gene expression between control mice (Tbx6 +/−) and Tbx6-deficient mice (Tbx6 −/−), we demonstrate that at least four genes, Twist2, Pitx2, Oscp1, and Nfxl1, are down-regulated with Tbx6 deficiency. These results suggest that ORR1A, ORR1B, ORR1C and ORR1D may contribute to the evolution of mammalian embryogenesis. PMID:28664156

Enhanced proliferation of primary rat type II pneumocytes by Jaagsiekte sheep retrovirus envelope protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnson, Chassidy; Jahid, Sohail; Voelker, Dennis R.

Jaagsiekte sheep retrovirus (JSRV) is the causative agent of a contagious lung cancer in sheep. The envelope protein (Env) is the oncogene, as it can transform cell lines in culture and induce tumors in animals, although the mechanisms for transformation are not yet clear because a system to perform transformation assays in differentiated type II pneumocytes does not exist. In this study we report culture of primary rat type II pneumocytes in conditions that favor prolonged expression of markers for type II pneumocytes. Env-expressing cultures formed more colonies that were larger in size and were viable for longer periods ofmore » time compared to vector control samples. The cells that remained in culture longer were confirmed to be derived from type II pneumocytes because they expressed surfactant protein C, cytokeratin, displayed alkaline phosphatase activity and were positive for Nile red. This system will be useful to study JSRV Env in the targets of transformation.« less

Retroviruses facilitate the rapid evolution of the mammalian placenta

PubMed Central

Chuong, Edward B.

2015-01-01

The mammalian placenta exhibits elevated expression of endogenous retroviruses (ERVs), but the evolutionary significance of this feature remains unclear. I propose that ERV-mediated regulatory evolution was, and continues to be, an important mechanism underlying the evolution of placenta development. Many recent studies have focused on the co-option of ERV-derived genes for specific functional adaptations in the placenta. However, the co-option of ERV-derived regulatory elements has the potential to co-opt entire gene regulatory networks, which, I argue, would facilitate relatively rapid developmental evolution of the placenta. I suggest a model in which an ancient retroviral infection led to the establishment of the ancestral placental developmental gene network through the co-option of ERV-derived regulatory elements. Consequently, placenta development would require elevated tolerance to ERV activity, which in turn would expose a continuous stream of novel ERV mutations that may have catalyzed the developmental diversification of the mammalian placenta. PMID:23873343

p53 as a retrovirus-induced oxidative stress modulator.

PubMed

Kim, Soo Jin; Wong, Paul K Y

2015-01-01

Infection of astrocytes by the neuropathogenic mutant of Moloney murine leukemia virus, ts1, exhibits increased levels of reactive oxygen species (ROS) and signs of oxidative stress compared with uninfected astrocytes. Previously, we have demonstrated that ts1 infection caused two separate events of ROS upregulation. The first upregulation occurs during early viral establishment in host cells and the second during the virus-mediated apoptotic process. In this study, we show that virus-mediated ROS upregulation activates the protein kinase, ataxia telangiectasia mutated, which in turn phosphorylates serine 15 on p53. This activation of p53 however, is unlikely associated with ts1-induced cell death. Rather p53 appears to be involved in suppressing intracellular ROS levels in astrocytes under oxidative stress. The activated p53 appears to delay retroviral gene expression by suppressing NADPH oxidase, a superoxide-producing enzyme. These results suggest that p53 plays a role as a retrovirus-mediated oxidative stress modulator. © 2015 The Authors.

Escape from R-peptide deletion in a {gamma}-retrovirus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schneider, Irene C.; Eckhardt, Manon; Brynza, Julia

2011-09-30

The R peptide in the cytoplasmic tail (C-tail) of {gamma}-retroviral envelope proteins (Env) prevents membrane fusion before budding. To analyse its role in the formation of replication competent, infectious particles, we developed chimeric murine leukaemia viruses (MLV) with unmodified or R-peptide deleted Env proteins of the gibbon ape leukaemia virus (GaLV). While titres of these viruses were unaffected, R-peptide deficiency led to strongly impaired spreading. Most remarkably, we isolated an escape mutant which had restored an open reading frame for a C-terminal extension of the truncated C-tail. A reconstituted virus encoding this escape C-tail replicated in cell culture. In contrastmore » to R-peptide deficient Env, particle incorporation of the escape Env was effective due to an enhanced protein expression and restored intracellular co-localisation with Gag proteins. Our data demonstrate that the R peptide not only regulates membrane fusion but also mediates efficient Env protein particle incorporation in {gamma}-retrovirus infected cells.« less

Endogenous Retroviruses: With Us and Against Us

NASA Astrophysics Data System (ADS)

Meyer, Thomas J.; Rosenkrantz, Jimi L.; Carbone, Lucia; Chavez, Shawn L.

2017-04-01

Mammalian genomes are scattered with thousands of copies of endogenous retroviruses (ERVs), mobile genetic elements that are relics of ancient retroviral infections. After inserting copies into the germ line of a host, most ERVs accumulate mutations that prevent the normal assembly of infectious viral particles, becoming trapped in host genomes and unable to leave to infect other cells. While most copies of ERVs are inactive, some are transcribed and encode the proteins needed to generate new insertions at novel loci. In some cases, old copies are removed via recombination and other mechanisms. This creates a shifting landscape of ERV copies within host genomes. New insertions can disrupt normal expression of nearby genes via directly inserting into key regulatory elements or by containing regulatory motifs within their sequences. Further, the transcriptional silencing of ERVs via epigenetic modification may result in changes to the epigenetic regulation of adjacent genes. In these ways, ERVs can be potent sources of regulatory disruption as well as genetic innovation. Here, we provide a brief review of the association between ERVs and gene expression, especially as observed in pre-implantation development and placentation. Moreover, we will describe the roles ERVs may play in somatic tissues, mostly in the context of human disease, including cancer, neurodegenerative disorders, and schizophrenia. Lastly, we discuss the recent discovery that some ERVs may have been pressed into the service of their host genomes to aid in the innate immune response to exogenous viral infections.

Highly efficient in vitro and in vivo delivery of functional RNAs using new versatile MS2-chimeric retrovirus-like particles

PubMed Central

Prel, Anne; Caval, Vincent; Gayon, Régis; Ravassard, Philippe; Duthoit, Christine; Payen, Emmanuel; Maouche-Chretien, Leila; Creneguy, Alison; Nguyen, Tuan Huy; Martin, Nicolas; Piver, Eric; Sevrain, Raphaël; Lamouroux, Lucille; Leboulch, Philippe; Deschaseaux, Frédéric; Bouillé, Pascale; Sensébé, Luc; Pagès, Jean-Christophe

2015-01-01

RNA delivery is an attractive strategy to achieve transient gene expression in research projects and in cell- or gene-based therapies. Despite significant efforts investigating vector-directed RNA transfer, there is still a requirement for better efficiency of delivery to primary cells and in vivo. Retroviral platforms drive RNA delivery, yet retrovirus RNA-packaging constraints limit gene transfer to two genome-molecules per viral particle. To improve retroviral transfer, we designed a dimerization-independent MS2-driven RNA packaging system using MS2-Coat-retrovirus chimeras. The engineered chimeric particles promoted effective packaging of several types of RNAs and enabled efficient transfer of biologically active RNAs in various cell types, including human CD34+ and iPS cells. Systemic injection of high-titer particles led to gene expression in mouse liver and transferring Cre-recombinase mRNA in muscle permitted widespread editing at the ROSA26 locus. We could further show that the VLPs were able to activate an osteoblast differentiation pathway by delivering RUNX2- or DLX5-mRNA into primary human bone-marrow mesenchymal-stem cells. Thus, the novel chimeric MS2-lentiviral particles are a versatile tool for a wide range of applications including cellular-programming or genome-editing. PMID:26528487

Physical and Functional Interactions of Human Endogenous Retrovirus Proteins Np9 and Rec with the Promyelocytic Leukemia Zinc Finger Protein▿

PubMed Central

Denne, Miriam; Sauter, Marlies; Armbruester, Vivienne; Licht, Jonathan D.; Roemer, Klaus; Mueller-Lantzsch, Nikolaus

2007-01-01

Only few of the human endogenous retrovirus (HERV) sequences in the human genome can produce proteins. We have previously reported that (i) patients with germ cell tumors often make antibodies against proteins encoded by HERV-K elements, (ii) expression of the HERV-K rec gene in transgenic mice can interfere with germ cell development and induce carcinoma in situ, and (iii) HERV-K np9 transcript is overproduced in many tumors including breast cancers. Here we document that both Np9 and Rec physically and functionally interact with the promyelocytic leukemia zinc finger (PLZF) tumor suppressor, a transcriptional repressor and chromatin remodeler implicated in cancer and the self-renewal of spermatogonial stem cells. Interaction is mediated via two different central and C-terminal domains of Np9 and Rec and the C-terminal zinc fingers of PLZF. One major target of PLZF is the c-myc proto-oncogene. Coexpression of Np9 and Rec with PLZF abrogates the transcriptional repression of the c-myc gene promoter by PLZF and results in c-Myc overproduction, altered expression of c-Myc-regulated genes, and corresponding effects on cell proliferation and survival. Thus, the human endogenous retrovirus proteins Np9 and Rec may act oncogenically by derepressing c-myc through the inhibition of PLZF. PMID:17360752

Tetraspanins displayed in retrovirus-derived virus-like particles and their immunogenicity.

PubMed

Soares, H R; Castro, R; Tomás, H A; Rodrigues, A F; Gomes-Alves, P; Bellier, B; Klatzmann, D; Carrondo, M J T; Alves, P M; Coroadinha, A S

2016-03-18

Virus-like particles (VLPs) are a particular subset of subunit vaccines which are currently explored as safer alternatives to live attenuated or inactivated vaccines. VLPs derived from retrovirus (retroVLPs) are commonly used as scaffolds for vaccine candidates due to their ability to incorporate heterologous envelope proteins. Pseudotyping retroVLPs is however not a selective process therefore, host cellular proteins such as tetraspanins are also included in the membrane. The contribution of these host-proteins to retrovirus immunogenicity remains unclear. In this work, human cells silenced and not silenced for tetraspanin CD81 were used to produce CD81(-) or CD81(+) retroVLPs. We first analyzed mice immune response against human CD81. Despite effective silencing of CD81 in retroVLP producing cells, both humoral and cellular immune responses showed persistent anti-CD81 immunogenicity, suggesting cross reactivity to related antigens. We thus compared the incorporation of related tetraspanins in retroVLPs and showed that decreased CD81 incorporation in CD81(-) retro-VLPs is compensated by an increased incorporation of CD9 and CD63 tetraspanins. These results highlight the dynamic nature of host-derived proteins incorporation in retroVLPs membrane, which should be considered when retrovirus-based biopharmaceuticals are produced in xenogeneic cells. Copyright © 2015 Elsevier Ltd. All rights reserved.

The antiretrovirus drug 3'-azido-3'-deoxythymidine increases the retrovirus mutation rate.

PubMed Central

Julias, J G; Kim, T; Arnold, G; Pathak, V K

1997-01-01

It was previously observed that the nucleoside analog 5-azacytidine increased the spleen necrosis virus (SNV) mutation rate 13-fold in one cycle of retrovirus replication (V. K. Pathak and H. M. Temin, J. Virol. 66:3093-3100, 1992). Based on this observation, we hypothesized that nucleoside analogs used as antiviral drugs may also increase retrovirus mutation rates. We sought to determine if 3'-azido-3'-deoxythymidine (AZT), the primary treatment for human immunodeficiency virus type 1 (HIV-1) infection, increases the retrovirus mutation rate. Two assays were used to determine the effects of AZT on retrovirus mutation rates. The strategy of the first assay involved measuring the in vivo rate of inactivation of the lacZ gene in one replication cycle of SNV- and murine leukemia virus-based retroviral vectors. We observed 7- and 10-fold increases in the SNV mutant frequency following treatment of target cells with 0.1 and 0.5 microM AZT, respectively. The murine leukemia virus mutant frequency increased two- and threefold following treatment of target cells with 0.5 and 1.0 microM AZT, respectively. The second assay used an SNV-based shuttle vector containing the lacZ alpha gene. Proviruses were recovered as plasmids in Escherichia coli, and the rate of inactivation of lacZ alpha was measured. The results indicated that treatment of target cells increased the overall mutation rate two- to threefold. DNA sequence analysis of mutant proviruses indicated that AZT increased both the deletion and substitution rates. These results suggest that AZT treatment of HIV-1 infection may increase the degree of viral variation and alter virus evolution or pathogenesis. PMID:9151812

Human endogenous retrovirus expression is inversely related with the up-regulation of interferon-inducible genes in the skin of patients with lichen planus.

PubMed

Nogueira, Marcelle Almeida de Sousa; Gavioli, Camila Fátima Biancardi; Pereira, Nátalli Zanete; de Carvalho, Gabriel Costa; Domingues, Rosana; Aoki, Valéria; Sato, Maria Notomi

2015-04-01

Lichen planus (LP) is a common inflammatory skin disease of unknown etiology. Reports of a common transactivation of quiescent human endogenous retroviruses (HERVs) support the connection of viruses to the disease. HERVs are ancient retroviral sequences in the human genome and their transcription is often deregulated in cancer and autoimmune diseases. We explored the transcriptional activity of HERV sequences as well as the antiviral restriction factor and interferon-inducible genes in the skin from LP patients and healthy control (HC) donors. The study included 13 skin biopsies from patients with LP and 12 controls. Real-time PCR assay identified significant decrease in the HERV-K gag and env mRNA expression levels in LP subjects, when compared to control group. The expressions of HERV-K18 and HERV-W env were also inhibited in the skin of LP patients. We observed a strong correlation between HERV-K gag with other HERV sequences, regardless the down-modulation of transcripts levels in LP group. In contrast, a significant up-regulation of the cytidine deaminase APOBEC 3G (apolipoprotein B mRNA-editing), and the GTPase MxA (Myxovirus resistance A) mRNA expression level was identified in the LP skin specimens. Other transcript expressions, such as the master regulator of type I interferon-dependent immune responses, STING (stimulator of interferon genes) and IRF-7 (interferon regulatory factor 7), IFN-β and the inflammassome NALP3, had increased levels in LP, when compared to HC group. Our study suggests that interferon-inducible factors, in addition to their role in innate immunity against exogenous pathogens, contribute to the immune control of HERVs. Evaluation of the balance between HERV and interferon-inducible factor expression could possibly contribute to surveillance of inflammatory/malignant status of skin diseases.

Generation of fibrosarcomas in vivo by a retrovirus that expresses the normal B chain of platelet-derived growth factor and mimics the alternative splice pattern of the v-sis oncogene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pech, M.; Gazit, A.; Arnstein, P.

1989-04-01

A retrovirus containing the entire human platelet-derived growth factor B-chain (PDGF-B) gene was constructed in order to investigate the in vivo biological activity of its encoded growth factor. When this virus was introduced into newborn mice, it reproducibly generated fibrosarcomas at the site of inoculation. Proviruses in each fibrosarcoma analyzed had lost 149 nucleotides downstream of the PDGF-B coding region. This deletion originated from an alternative or aberrant splice event that occurred within exon 7 of the PDGF-B gene and mimicked the v-sis oncogene. Thus, deletion of this region may be necessary for efficient retrovirus replication or for more potentmore » transforming function. Evidence that the normal growth factor coding sequence was unaltered derived from RNase protection studies and immunoprecipitation analysis. Tumors were generally polyclonal but demonstrated clonal subpopulations. Moreover, tumor-derived cell lines became monoclonal within a few tissue culture passages and rapidly formed tumors in vivo. These findings argue that overexpression of the normal human PDGF-B gene product under retrovirus control can induce the fully malignant phenotype.« less

Vaccination directed against the human endogenous retrovirus-K (HERV-K) gag protein slows HERV-K gag expressing cell growth in a murine model system.

PubMed

Kraus, Benjamin; Fischer, Katrin; Sliva, Katja; Schnierle, Barbara S

2014-03-26

Human endogenous retroviruses (HERVs) are remnants of ancestral infections and chromosomally integrated in all cells of an individual, are transmitted only vertically and are defective in viral replication. However enhanced expression of HERV-K accompanied by the emergence of anti-HERV-K-directed immune responses has been observed inter-alia in HIV-infected individuals and tumor patients. Therefore HERV-K might serve as a tumor-specific antigen or even as a constant target for the development of an HIV vaccine. To verify our hypothesis, we tested the immunogenicity of HERV-K Gag by using a recombinant vaccinia virus (MVA-HKcon) expressing the HERV-K Gag protein and established an animal model to test its vaccination efficacy. Murine renal carcinoma cells (Renca) were genetically altered to express E. coli beta-galactosidase (RLZ cells) and the HERV-K Gag protein (RLZ-HKGag cells). Subcutaneous application of RLZ-HKGag cells into syngenic BALB/c mice resulted in the formation of local tumors in MVA vaccinated mice. MVA-HKcon vaccination reduced the tumor growth. Furthermore, intravenous injection of RLZ-HKGag cells led to the formation of pulmonary metastases. Vaccination of tumor-bearing mice with MVA-HKcon drastically reduced the number of pulmonary RLZ-HKGag tumor nodules compared to vaccination with wild-type MVA. The data demonstrate that HERV-K Gag is a useful target for vaccine development and might offer new treatment opportunities for cancer patients.

The evolutionary capacitor HSP90 buffers the regulatory effects of mammalian endogenous retroviruses.

PubMed

Hummel, Barbara; Hansen, Erik C; Yoveva, Aneliya; Aprile-Garcia, Fernando; Hussong, Rebecca; Sawarkar, Ritwick

2017-03-01

Understanding how genotypes are linked to phenotypes is important in biomedical and evolutionary studies. The chaperone heat-shock protein 90 (HSP90) buffers genetic variation by stabilizing proteins with variant sequences, thereby uncoupling phenotypes from genotypes. Here we report an unexpected role of HSP90 in buffering cis-regulatory variation affecting gene expression. By using the tripartite-motif-containing 28 (TRIM28; also known as KAP1)-mediated epigenetic pathway, HSP90 represses the regulatory influence of endogenous retroviruses (ERVs) on neighboring genes that are critical for mouse development. Our data based on natural variations in the mouse genome show that genes respond to HSP90 inhibition in a manner dependent on their genomic location with regard to strain-specific ERV-insertion sites. The evolutionary-capacitor function of HSP90 may thus have facilitated the exaptation of ERVs as key modifiers of gene expression and morphological diversification. Our findings add a new regulatory layer through which HSP90 uncouples phenotypic outcomes from individual genotypes.

Retrovirus purification: method that conserves envelope glycoprotein and maximizes infectivity.

PubMed Central

McGrath, M; Witte, O; Pincus, T; Weissman, I L

1978-01-01

A Sepharose 4B chromatographic method for purification of retroviruses is described which was less time consuming, increased purified virus yields, conserved viral glycoprotein, and increased recovery of biological infectivity in comparison with conventional sucrose gradient ultracentrifugation techniques. Images PMID:205680

Identification of novel, highly expressed retroviral microRNAs in cells infected by bovine foamy virus.

PubMed

Whisnant, Adam W; Kehl, Timo; Bao, Qiuying; Materniak, Magdalena; Kuzmak, Jacek; Löchelt, Martin; Cullen, Bryan R

2014-05-01

While numerous viral microRNAs (miRNAs) expressed by DNA viruses, especially herpesvirus family members, have been reported, there have been very few reports of miRNAs derived from RNA viruses. Here we describe three miRNAs expressed by bovine foamy virus (BFV), a member of the spumavirus subfamily of retroviruses, in both BFV-infected cultured cells and BFV-infected cattle. All three viral miRNAs are initially expressed in the form of an ∼ 122-nucleotide (nt) pri-miRNA, encoded within the BFV long terminal repeat U3 region, that is subsequently cleaved to generate two pre-miRNAs that are then processed to yield three distinct, biologically active miRNAs. The BFV pri-miRNA is transcribed by RNA polymerase III, and the three resultant mature miRNAs were found to contribute a remarkable ∼ 70% of all miRNAs expressed in BFV-infected cells. These data document the second example of a retrovirus that is able to express viral miRNAs by using embedded proviral RNA polymerase III promoters. Foamy viruses are a ubiquitous family of nonpathogenic retroviruses that have potential as gene therapy vectors in humans. Here we demonstrate that bovine foamy virus (BFV) expresses high levels of three viral microRNAs (miRNAs) in BFV-infected cells in culture and also in infected cattle. The BFV miRNAs are unusual in that they are initially transcribed by RNA polymerase III as a single, ∼ 122-nt pri-miRNA that is subsequently processed to release three fully functional miRNAs. The observation that BFV, a foamy virus, is able to express viral miRNAs in infected cells adds to emerging evidence that miRNA expression is a common, albeit clearly not universal, property of retroviruses and suggests that these miRNAs may exert a significant effect on viral replication in vivo.

Identification of Novel, Highly Expressed Retroviral MicroRNAs in Cells Infected by Bovine Foamy Virus

PubMed Central

Whisnant, Adam W.; Kehl, Timo; Bao, Qiuying; Materniak, Magdalena; Kuzmak, Jacek; Löchelt, Martin

2014-01-01

ABSTRACT While numerous viral microRNAs (miRNAs) expressed by DNA viruses, especially herpesvirus family members, have been reported, there have been very few reports of miRNAs derived from RNA viruses. Here we describe three miRNAs expressed by bovine foamy virus (BFV), a member of the spumavirus subfamily of retroviruses, in both BFV-infected cultured cells and BFV-infected cattle. All three viral miRNAs are initially expressed in the form of an ∼122-nucleotide (nt) pri-miRNA, encoded within the BFV long terminal repeat U3 region, that is subsequently cleaved to generate two pre-miRNAs that are then processed to yield three distinct, biologically active miRNAs. The BFV pri-miRNA is transcribed by RNA polymerase III, and the three resultant mature miRNAs were found to contribute a remarkable ∼70% of all miRNAs expressed in BFV-infected cells. These data document the second example of a retrovirus that is able to express viral miRNAs by using embedded proviral RNA polymerase III promoters. IMPORTANCE Foamy viruses are a ubiquitous family of nonpathogenic retroviruses that have potential as gene therapy vectors in humans. Here we demonstrate that bovine foamy virus (BFV) expresses high levels of three viral microRNAs (miRNAs) in BFV-infected cells in culture and also in infected cattle. The BFV miRNAs are unusual in that they are initially transcribed by RNA polymerase III as a single, ∼122-nt pri-miRNA that is subsequently processed to release three fully functional miRNAs. The observation that BFV, a foamy virus, is able to express viral miRNAs in infected cells adds to emerging evidence that miRNA expression is a common, albeit clearly not universal, property of retroviruses and suggests that these miRNAs may exert a significant effect on viral replication in vivo. PMID:24522910

Murine endogenous retroviruses

PubMed Central

2016-01-01

Up to 10% of the mouse genome is comprised of endogenous retrovirus (ERV) sequences, and most represent the remains of ancient germ line infections. Our knowledge of the three distinct classes of ERVs is inversely correlated with their copy number, and their characterization has benefited from the availability of divergent wild mouse species and subspecies, and from ongoing analysis of the Mus genome sequence. In contrast to human ERVs, which are nearly all extinct, active mouse ERVs can still be found in all three ERV classes. The distribution and diversity of ERVs has been shaped by host-virus interactions over the course of evolution, but ERVs have also been pivotal in shaping the mouse genome by altering host genes through insertional mutagenesis, by adding novel regulatory and coding sequences, and by their co-option by host cells as retroviral resistance genes. We review mechanisms by which an adaptive coexistence has evolved. (Part of a Multi-author Review) PMID:18818872

Fate of the surface protein gp70 during entry of retrovirus into mouse fibroblasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Andersen, K.B.

1985-04-15

The kinetics of the viral surface protein gp70 and the viral core proteins p30 and p15C were followed during retrovirus entry into mouse fibroblasts. All three proteins were internalized, but whereas essentially all the gp70 was degraded, approximately one-third of the core proteins remained stable in the cells. These diverging routes of the different proteins are in agreement with the proposed route, that retrovirus enters the cells by endocytosis followed by a membrane fusion between the virus membrane and the vesicle membrane.

PML mediates the interferon-induced antiviral state against a complex retrovirus via its association with the viral transactivator

PubMed Central

Regad, Tarik; Saib, Ali; Lallemand-Breitenbach, Valérie; Pandolfi, Pier Paolo; de Thé, Hugues; Chelbi-Alix, Mounira K.

2001-01-01

The promyelocytic leukaemia (PML) protein localizes in the nucleus both in the nucleoplasm and in matrix-associated multiprotein complexes known as nuclear bodies (NBs). The number and the intensity of PML NBs increase in response to interferon (IFN). Overexpression of PML affects the replication of vesicular stomatitis virus and influenza virus. However, PML has a less powerful antiviral activity against these viruses than the IFN mediator MxA. Here, we show that overexpression of PML, but not that of Mx1 or MxA, leads to a drastic decrease of a complex retrovirus, the human foamy virus (HFV), gene expression. PML represses HFV transcription by complexing the HFV transactivator, Tas, preventing its direct binding to viral DNA. This physical interaction requires the N-terminal region of Tas and the RING finger of PML, but does not necessitate PML localization in NBs. Finally, we show that IFN treatment inhibits HFV replication in wild-type but not in PML–/– cells. These findings point to a role for PML in transcriptional repression and suggest that PML could play a key role in mediating an IFN-induced antiviral state against a complex retrovirus. PMID:11432836

Dysfunction of bovine endogenous retrovirus K2 envelope glycoprotein is related to unsuccessful intracellular trafficking.

PubMed

Nakaya, Yuki; Miyazawa, Takayuki

2014-06-01

Endogenous retroviruses (ERVs) are the remnants of retroviral infection of ancestral germ cells. Mutations introduced into ERVs halt the production of infectious agents, but their effects on the function of retroviral proteins are not fully understood. Retroviral envelope glycoproteins (Envs) are utilized in membrane fusion during viral entry, and we recently identified intact coding sequences for bovine endogenous retrovirus K1 (BERV-K1) and BERV-K2 Envs. Amino acid sequences of BERV-K1 Env (also called Fematrin-1) and BERV-K2 Env are similar, and both viruses are classified in the genus Betaretrovirus. While Fematrin-1 plays an important role in cell-to-cell fusion in bovine placenta, the BERV-K2 envelope gene is marginally expressed in vivo, and its recombinant Env protein is defective in membrane fusion due to inefficient cleavage of surface (SU) and transmembrane subunits. Here, we conducted chimeric analyses of Fematrin-1 and BERV-K2 Envs and revealed that defective maturation of BERV-K2 Env contributed to failed intracellular trafficking. Fluorescence microscopy and flow cytometric analysis suggested that in contrast to Fematrin-1 Env, BERV-K2 Env could not be transported from the endoplasmic reticulum to the trans-Golgi network, where cellular proteases required for processing retroviral Envs are localized. We also identified that one of the responsive regions of this phenomenon resided within a 65-amino-acid region of BERV-K2 SU. This is the first report to identify that retroviral Env SU is involved in the regulation of intracellular trafficking, and it may help to elucidate the maturation process of Fematrin-1 and other related Envs. Retroviruses utilize envelope glycoproteins (Envs) to enter host target cells. Mature retroviral Env is a heterodimer, which consists of surface (SU) and transmembrane (TM) subunits that are generated by the cleavage of an Env precursor protein in the trans-Golgi network. SU and TM mediate the recognition of the entry

Dysfunction of Bovine Endogenous Retrovirus K2 Envelope Glycoprotein Is Related to Unsuccessful Intracellular Trafficking

PubMed Central

2014-01-01

ABSTRACT Endogenous retroviruses (ERVs) are the remnants of retroviral infection of ancestral germ cells. Mutations introduced into ERVs halt the production of infectious agents, but their effects on the function of retroviral proteins are not fully understood. Retroviral envelope glycoproteins (Envs) are utilized in membrane fusion during viral entry, and we recently identified intact coding sequences for bovine endogenous retrovirus K1 (BERV-K1) and BERV-K2 Envs. Amino acid sequences of BERV-K1 Env (also called Fematrin-1) and BERV-K2 Env are similar, and both viruses are classified in the genus Betaretrovirus. While Fematrin-1 plays an important role in cell-to-cell fusion in bovine placenta, the BERV-K2 envelope gene is marginally expressed in vivo, and its recombinant Env protein is defective in membrane fusion due to inefficient cleavage of surface (SU) and transmembrane subunits. Here, we conducted chimeric analyses of Fematrin-1 and BERV-K2 Envs and revealed that defective maturation of BERV-K2 Env contributed to failed intracellular trafficking. Fluorescence microscopy and flow cytometric analysis suggested that in contrast to Fematrin-1 Env, BERV-K2 Env could not be transported from the endoplasmic reticulum to the trans-Golgi network, where cellular proteases required for processing retroviral Envs are localized. We also identified that one of the responsive regions of this phenomenon resided within a 65-amino-acid region of BERV-K2 SU. This is the first report to identify that retroviral Env SU is involved in the regulation of intracellular trafficking, and it may help to elucidate the maturation process of Fematrin-1 and other related Envs. IMPORTANCE Retroviruses utilize envelope glycoproteins (Envs) to enter host target cells. Mature retroviral Env is a heterodimer, which consists of surface (SU) and transmembrane (TM) subunits that are generated by the cleavage of an Env precursor protein in the trans-Golgi network. SU and TM mediate the recognition

Squamous epithelial proliferation induced by walleye dermal sarcoma retrovirus cyclin in transgenic mice

PubMed Central

Lairmore, Michael D.; Stanley, James R.; Weber, Stacy A.; Holzschu, Donald L.

2000-01-01

Walleye dermal sarcoma (WDS) is a common disease of walleye fish in the United States and Canada. These proliferative lesions are present autumn through winter and regress in the spring. Walleye dermal sarcoma virus (WDSV), a retrovirus distantly related to other members of the family Retroviridae, has been etiologically linked to the development of WDS. We have reported that the D-cyclin homologue [retroviral (rv) cyclin] encoded by WDSV rescues yeast conditionally deficient for cyclin synthesis from growth arrest and that WDSV-cyclin mRNA is present in developing tumors. These data strongly suggest that the rv-cyclin plays a central role in the development of WDS. To test the ability of the WDSV rv-cyclin to induce cell proliferation, we have generated transgenic mice expressing the rv-cyclin in squamous epithelia from the bovine keratin-5 promoter. The transgenic animals were smaller than littermates, had reduced numbers of hair follicles, and transgenic females did not lactate properly. Following injury the transgenic animals developed severe squamous epithelial hyperplasia and dysplasia with ultrastructural characteristics of neoplastic squamous epithelium. Immunocytochemistry studies demonstrated that the hyperplastic epithelium stained positive for cytokeratin and were abnormally differentiated. Furthermore, the rv-cyclin protein was detected in the thickened basal cell layers of the proliferating lesions. These data are the first to indicate that the highly divergent WDSV rv-cyclin is a very potent stimulator of eukaryotic cell proliferation and to demonstrate the potential of a cyclin homologue encoded by a retrovirus to induce hyperplastic skin lesions. PMID:10811912

Comparison between S+L- assay and LacZ marker rescue assay for detecting replication-competent gammaretroviruses.

PubMed

Hashimoto-Gotoh, A; Yoshikawa, R; Miyazawa, T

2015-09-01

To avoid contamination of adventitious gammaretroviruses in biological products such as vaccines, it is necessary to check the master seed cells for manufacturing. There are several assays to detect infectious gammaretroviruses. Among these, sarcoma-positive, leukemia-negative (S+L-) assay is a classical infectivity assay, which is often recommended in governmental guidelines. The S+L- cells used in S+L- assay generate unique focus upon the infection of replication-competent gammaretroviruses. Although S+L- assay is well recognized for the detection, their applicability is questionable in some cases. On the other hand, LacZ marker rescue (LMR) assay detects infectious gammaretroviruses by transducing LacZ marker gene to the target cells, which shows lacZ-positive foci if the infectious virus is present. In this study, we compared LMR and S+L- assays for detection of a variety of endogenous and exogenous gammaretroviruses. As results, LMR assay could detect all gammaretroviruses examined. On the other hand, S+L- assay using feline S+L- cells, termed QN10S, could not detect porcine endogenous retrovirus (PERV) subgroups A/B. Further, S+L- mink cells could not detect feline leukemia virus subgroups B in addition to PERV-A/B. These data indicate that LMR assay is better suited to detect wider range of gammaretroviruses. Copyright © 2015 The International Alliance for Biological Standardization. Published by Elsevier Ltd. All rights reserved.

Detection of koala retrovirus subgroup B (KoRV-B) in animals housed at European zoos.

PubMed

Fiebig, Uwe; Keller, Martina; Denner, Joachim

2016-12-01

Many koalas carry an endogenous retrovirus, KoRV-A, in their genome. Recently, a second retrovirus, KoRV-B, was detected in koalas in Japanese and U.S. zoos. However, this virus is not endogenous, differs in the receptor binding site of the surface envelope protein, and uses a receptor different from that of KoRV-A. We describe here a KoRV-B found in koalas at zoos in Germany and Belgium that differs slightly from that found in the Los Angeles zoo.

Apparatus for testing for infection by a retrovirus

DOEpatents

Layne, Scott P.; Beugelsdijk, Tony J.

1999-01-01

An apparatus for testing specimens for infection by a retrovirus is described. The apparatus comprises a process controller including a communications module for translating user commands into test instrument suite commands and a means for communicating specimen test results to a user. The apparatus further comprises a test instrument suite including a means for treating the specimen to manifest an observable result and a detector for measuring the observable result.

Lack of detection of a putative retrovirus associated with haemic neoplasia in the soft shell clam Mya arenaria.

PubMed

AboElkhair, M; Iwamoto, T; Clark, K F; McKenna, P; Siah, A; Greenwood, S J; Berthe, F C J; Casey, J W; Cepica, A

2012-01-01

Haemic neoplasia (HN) is a leukemia-like disease that affects at least 20 species of marine bivalves including soft shell clam, Mya arenaria. Since the disease was discovered in 1969, the etiology remains unknown. A retroviral etiology has been suggested based on the detection of reverse transcriptase activity and electron microscopic observation of retroviral-like particles using negative staining. To date, however no virus isolate and no retroviral sequence from HN has been obtained. Moreover, transmission of the disease by cell-free filtrate from affected clams has not been reproduced. In the current study, we reinvestigated the association of HN with a putative retrovirus. Sucrose gradient centrifugation followed by assessment of reverse transcriptase activity, electrophoretic analysis of protein and RNA, and electron microscopic examinations of fractions corresponding to retroviral density were employed. Detection of retroviral pol sequences using degenerate RT-PCR approaches was also attempted. Our results showed visible bands at the expected density of retrovirus in HN-positive and HN-negative clam tissues and both with reverse transcriptase activity. Electron microscopy, RNA analysis, protein analysis, and PCR systems targeting the pol gene of retroviruses did not however provide clear evidence supporting presence of a retrovirus. We point out that the retrovirus etiology of HN of Mya arenaria proposed some 25 years ago should be reconsidered in the absence of a virus isolate or virus sequences. Copyright © 2011 Elsevier Inc. All rights reserved.

Foamy virus reverse transcriptase is expressed independently from the Gag protein.

PubMed Central

Enssle, J; Jordan, I; Mauer, B; Rethwilm, A

1996-01-01

In the foamy virus (FV) subgroup of retroviruses the pol genes are located in the +1 reading frame relative to the gag genes and possess potential ATG initiation codons in their 5' regions. This genome organization suggests either a + 1 ribosomal frameshift to generate a Gag-Pol fusion protein, similar to all other retroviruses studied so far, or new initiation of Pol translation, as used by pararetroviruses, to express the Pol protein. By using a genetic approach we have ruled out the former possibility and provide evidence for the latter. Two down-mutations (M53 and M54) of the pol ATG codon were found to abolish replication and Pol protein expression of the human FV isolate. The introduction of a new ATG in mutation M55, 3' to the down-mutated ATG of mutation M53, restored replication competence, indicating that the pol ATG functions as a translational initiation codon. Two nonsense mutants (M56 and M57), which functionally separated gag and pol with respect to potential frame-shifting sites, were also replication-competent, providing further genetic evidence that FVs express the Pol protein independently from Gag. Our results show that during a particular step of the replication cycle, FVs differ fundamentally from all other retroviruses. Images Fig. 3 PMID:8633029

Retrovirus-Based Virus-Like Particle Immunogenicity and Its Modulation by Toll-Like Receptor Activation.

PubMed

Pitoiset, Fabien; Vazquez, Thomas; Levacher, Beatrice; Nehar-Belaid, Djamel; Dérian, Nicolas; Vigneron, James; Klatzmann, David; Bellier, Bertrand

2017-11-01

Retrovirus-derived virus-like particles (VLPs) are particularly interesting vaccine platforms, as they trigger efficient humoral and cellular immune responses and can be used to display heterologous antigens. In this study, we characterized the intrinsic immunogenicity of VLPs and investigated their possible adjuvantization by incorporation of Toll-like receptor (TLR) ligands. We designed a noncoding single-stranded RNA (ncRNA) that could be encapsidated by VLPs and induce TLR7/8 signaling. We found that VLPs efficiently induce in vitro dendritic cell activation, which can be improved by ncRNA encapsidation ( ncRNA VLPs). Transcriptome studies of dendritic cells harvested from the spleens of immunized mice identified antigen presentation and immune activation as the main gene expression signatures induced by VLPs, while TLR signaling and Th1 signatures characterize ncRNA VLPs. In vivo and compared with standard VLPs, ncRNA VLPs promoted Th1 responses and improved CD8 + T cell proliferation in a MyD88-dependent manner. In an HIV vaccine mouse model, HIV-pseudotyped ncRNA VLPs elicited stronger antigen-specific cellular and humoral responses than VLPs. Altogether, our findings provide molecular evidence for a strong vaccine potential of retrovirus-derived VLPs that can be further improved by harnessing TLR-mediated immune activation. IMPORTANCE We previously reported that DNA vaccines encoding antigens displayed in/on retroviral VLPs are more efficient than standard DNA vaccines at inducing cellular and humoral immune responses. We aimed to decipher the mechanisms and investigated the VLPs' immunogenicity independently of DNA vaccination. We show that VLPs have the ability to activate antigen-presenting cells directly, thus confirming their intrinsic immunostimulatory properties and their potential to be used as an antigenic platform. Notably, this immunogenicity can be further improved and/or oriented by the incorporation into VLPs of ncRNA, which provides further

A soluble envelope protein of endogenous retrovirus (FeLIX) present in serum of domestic cats mediates infection of a pathogenic variant of feline leukemia virus.

PubMed

Sakaguchi, Shoichi; Shojima, Takayuki; Fukui, Daisuke; Miyazawa, Takayuki

2015-03-01

T-lymphotropic feline leukemia virus (FeLV-T), a highly pathogenic variant of FeLV, induces severe immunosuppression in cats. FeLV-T is fusion defective because in its PHQ motif, a gammaretroviral consensus motif in the N terminus of an envelope protein, histidine is replaced with aspartate. Infection by FeLV-T requires FeLIX, a truncated envelope protein encoded by an endogenous FeLV, for transactivation of infectivity and Pit1 for binding FeLIX. Although Pit1 is present in most tissues in cats, the expression of FeLIX is limited to certain cells in lymphoid organs. Therefore, the host cell range of FeLV-T was thought to be restricted to cells expressing FeLIX. However, because FeLIX is a soluble factor and is expressed constitutively in lymphoid organs, we presumed it to be present in blood and evaluated its activities in sera of various mammalian species using a pseudotype assay. We demonstrated that cat serum has FeLIX activity at a functional level, suggesting that FeLIX is present in the blood and that FeLV-T may be able to infect cells expressing Pit1 regardless of the expression of FeLIX in vivo. In addition, FeLIX activities in sera were detected only in domestic cats and not in other feline species tested. To our knowledge, this is the first report to prove that a large amount of truncated envelope protein of endogenous retrovirus is circulating in the blood to facilitate the infection of a pathogenic exogenous retrovirus. © 2015 The Authors.

Expression of human argininosuccinate synthetase after retroviral-mediated gene transfer.

PubMed

Wood, P A; Partridge, C A; O'Brien, W E; Beaudet, A L

1986-09-01

The cDNA sequence for human argininosuccinate synthetase (AS) was introduced into plasmid expression vectors with an SV40 promoter or Rous sarcoma virus promoter to construct pSV2-AS and pRSV-AS, respectively, and human enzyme was synthesized after gene transfer into Chinese hamster cells. The functional cDNA was inserted into the retroviral vectors pZIP-NeoSV(X) and pZIP-NeoSV(B). Ecotropic AS retrovirus was produced after calcium-phosphate-mediated gene transfer of these constructions into the packaging cell line psi-2, and viral titers up to 10(5) CFU/ml were obtained. Recombinant AS retrovirus was evaluated by detecting G-418-resistant colonies after infection of the rodent cells, XC, NRK, and 3T3. Colonies were also obtained when infected XC cells were selected in citrulline medium for expression of AS activity. Southern blot analysis of infected cells demonstrated that the recombinant retroviral genome was not altered grossly after infecting some rodent cells, while other cells showed evidence of rearrangement. A rapid assay for detecting AS retrovirus was developed based on the incorporation of [14C]citrulline into protein by intact 3T3 cells or XC cells.

Human retroviruses: their role in cancer.

PubMed

Blattner, W A

1999-01-01

Viruses are etiologically linked to approximately 20% of all malignancies worldwide. Retroviruses account for approximately 8%-10% of the total. For human T-cell leukemia virus 1 (HTLV-I), the viral regulatory tax gene product is responsible for enhanced transcription of viral and cellular genes that promote cell growth by stimulating various growth factors and through dysregulation of cellular regulatory suppressor genes, such as p53. After a long latent period, adult T-cell leukemia/lymphoma (ATL) occurs in 1 per 1000 carriers per year, resulting in 2500-3000 cases per year worldwide and over half of the adult lymphoid malignancies in endemic areas. Human immunodeficiency virus 1 (HIV-1) accounts for a significant cancer burden, and its transactivating regulatory protein Tat enhances direct and indirect cytokine and immunological dysregulation to cause diverse cancers. Kaposi's sarcoma (KS) is a very rare tumor except after HIV-1 infection, when its incidence is greatly amplified reaching seventy thousand-fold in HIV-infected homosexual men. Human herpesvirus 8 (HHV-8), which is also known as Kaposi's sarcoma-associated virus (KSHV), is a necessary but not sufficient etiological factor in KS. The dramatic decline of KS since the introduction of highly active antiretroviral therapy (HAART) could be due to suppression of HIV-1 tat. B-cell non-Hodgkin's lymphoma occurs as their first acquired immunodeficiency syndrome-defining diagnosis in 3%-4% of HIV-infected patients. Hodgkin's lymphoma is also associated with HIV infection but at a lower risk. Human papillomaviruses are linked to invasive cervical cancer and anogenital cancers among HIV-infected patients. Human retroviruses cause malignancy via direct effects as well as through interactions with other oncogenic herpesviruses and other viruses.

Human retroviruses and AIDS, 1991. [CONTAINS GLOSSARY

DOE Office of Scientific and Technical Information (OSTI.GOV)

Myers, G.; Korber, B.; Berzofsky, J.A.

1991-05-01

This compendium and the accompanying floppy diskettes are the result of an effort to compile and rapidly publish all relevant molecular data concerning the human immunodeficiency viruses (HIV) and related retroviruses.The scope of the compendium and database is best summarized by the five parts that it comprises: (1) HIV and SIV Nucleotide Sequences; (2) Amino Acid Sequences; (3) Analyses; (4) Related Sequences; and (5) Database Communications. Information within all the parts is updated at least twice in each year, which accounts for the modes of binding and pagination in the compendium.

Serum Antibody Response to Koala Retrovirus Antigens Varies in Free-Ranging Koalas ( Phascolarctos cinereus ) in Australia: Implications for Vaccine Design.

PubMed

Waugh, Courtney; Gillett, Amber; Polkinghorne, Adam; Timms, Peter

2016-04-28

Little is known about the immune response in the koala ( Phascolarctos cinereus ) to its retroviruses. Koala retroviruses (KoRVs) have been linked to neoplasia in wild and captive koalas, but there is no treatment available. We tested the KoRV-specific serum immunoglobulin G antibody response in nonimmunized and immunized koalas.

Transcriptional and functional studies of Human Endogenous Retrovirus envelope EnvP(b) and EnvV genes in human trophoblasts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vargas, Amandine, E-mail: amandine.vargas@voila.fr; Thiery, Maxime, E-mail: thiery.maxime@courrier.uqam.ca; Lafond, Julie, E-mail: lafond.julie@uqam.ca

2012-03-30

HERV (Human Endogenous Retrovirus)-encoded envelope proteins are implicated in the development of the placenta. Indeed, Syncytin-1 and -2 play a crucial role in the fusion of human trophoblasts, a key step in placentation. Other studies have identified two other HERV env proteins, namely EnvP(b) and EnvV, both expressed in the placenta. In this study, we have fully characterized both env transcripts and their expression pattern and have assessed their implication in trophoblast fusion. Through RACE analyses, standard spliced transcripts were detected, while EnvV transcripts demonstrated alternative splicing at its 3 Prime end. Promoter activity and expression of both genes weremore » induced in forskolin-stimulated BeWo cells and in primary trophoblasts. Although we have confirmed the fusogenic activity of EnvP(b), overexpression or silencing experiments revealed no impact of this protein on trophoblast fusion. Our results demonstrate that both env genes are expressed in human trophoblasts but are not required for syncytialization.« less

Murine Endogenous Retroviruses Are Detectable in Patient-Derived Xenografts but Not in Patient-Individual Cell Lines of Human Colorectal Cancer.

PubMed

Bock, Stephanie; Mullins, Christina S; Klar, Ernst; Pérot, Philippe; Maletzki, Claudia; Linnebacher, Michael

2018-01-01

Endogenous retroviruses are remnants of retroviral infections. In contrast to their human counterparts, murine endogenous retroviruses (mERV) still can synthesize infectious particles and retrotranspose. Xenotransplanted human cells have occasionally been described to be mERV infected. With genetic engineered mice and patient-derived xenografts (PDXs) on the rise as eminent research tools, we here systematically investigated, if different tumor models harbor mERV infections. Relevant mERV candidates were first preselected by next generation sequencing (NGS) analysis of spontaneous lymphomas triggered by colorectal cancer (CRC) PDX tissue. Two primer systems were designed for each of these candidates (AblMLV, EcoMLV, EndoPP, MLV, and preXMRV) and implemented in an quantitative real-time (RT-qPCR) screen using murine tissues ( n = 11), PDX-tissues ( n = 22), PDX-derived cell lines ( n = 13), and patient-derived tumor cell lines ( n = 14). The expression levels of mERV varied largely both in the PDX samples and in the mouse tissues. No mERV signal was, however, obtained from cDNA or genomic DNA of CRC cell lines. Expression of EcoMLV was higher in PDX than in murine tissues; for EndoPP it was the opposite. These two were thus further investigated in 40 additional PDX. In addition, four patient-derived cell lines free of any mERV expression were subcutaneously injected into immunodeficient mice. Outgrowing cell-derived xenografts barely expressed EndoPP. In contrast, the expression of EcoMLV was even higher than in surrounding mouse tissues. This expression gradually vanished within few passages of re-cultivated cells. In summary, these results strongly imply that: (i) PDX and murine tissues in general are likely to be contaminated by mERV, (ii) mERV are expressed transiently and at low level in fresh PDX-derived cell cultures, and (iii) mERV integration into the genome of human cells is unlikely or at least a very rare event. Thus, mERVs are stowaways present in

Posttranscriptional regulation of retroviral gene expression: primary RNA transcripts play three roles as pre-mRNA, mRNA, and genomic RNA

PubMed Central

LeBlanc, Jason; Weil, Jason; Beemon, Karen

2013-01-01

After reverse transcription of the retroviral RNA genome and integration of the DNA provirus into the host genome, host machinery is used for viral gene expression along with viral proteins and RNA regulatory elements. Here, we discuss co-transcriptional and posttranscriptional regulation of retroviral gene expression, comparing simple and complex retroviruses. Cellular RNA polymerase II synthesizes full-length viral primary RNA transcripts that are capped and polyadenylated. All retroviruses generate a singly spliced env mRNA from this primary transcript, which encodes the viral glycoproteins. In addition, complex viral RNAs are alternatively spliced to generate accessory proteins, such as Rev, which is involved in posttranscriptional regulation of HIV-1 RNA. Importantly, the splicing of all retroviruses is incomplete; they must maintain and export a fraction of their primary RNA transcripts. This unspliced RNA functions both as the major mRNA for Gag and Pol proteins and as the packaged genomic RNA. Different retroviruses export their unspliced viral RNA from the nucleus to the cytoplasm by either Tap-dependent or Rev/CRM1-dependent routes. Translation of the unspliced mRNA involves frame-shifting or termination codon suppression so that the Gag proteins, which make up the capsid, are expressed more abundantly than the Pol proteins, which are the viral enzymes. After the viral polyproteins assemble into viral particles and bud from the cell membrane, a viral encoded protease cleaves them. Some retroviruses have evolved mechanisms to protect their unspliced RNA from decay by nonsense-mediated RNA decay and to prevent genome editing by the cellular APOBEC deaminases. PMID:23754689

The effect of bovine BST2A1 on the release and cell-to-cell transmission of retroviruses.

PubMed

Liang, Zhibin; Zhang, Yang; Song, Jie; Zhang, Hui; Zhang, Suzhen; Li, Yue; Tan, Juan; Qiao, Wentao

2017-09-06

Human BST2 (hBST2, also called Tetherin) is a host restriction factor that blocks the release of various enveloped viruses. BST2s from different mammals also possess antiviral activity. Bovine BST2s (bBST2s), bBST2A1 and bBST2A2, reduce production of cell-free bovine leukemia virus (BLV) and vesicular stomatitis virus (VSV). However, the effect of bBST2 on other retroviruses remains unstudied. Here, we studied the antiviral activity of wildtype and mutant bBST2A1 proteins on retroviruses including human immunodeficiency virus type 1 (HIV-1), prototypic foamy virus (PFV), bovine foamy virus (BFV) and bovine immunodeficiency virus (BIV). The results showed that wildtype bBST2A1 suppressed the release of HIV-1, PFV and BFV. We also generated bBST2A1 mutants, and found that GPI anchor and dimerization, but not glycosylation, are essential for antiviral activity of bBST2A1. Moreover, unlike hBST2, bBST2A1 displayed no inhibitory effect on cell-to-cell transmission of PFV, BFV and BIV. Our data suggested that bBST2A1 inhibited retrovirus release, however, had no effect on cell-to-cell transmission of retroviruses.

The Population History of Endogenous Retroviruses in Mule Deer (Odocoileus hemionus)

PubMed Central

2014-01-01

Mobile elements are powerful agents of genomic evolution and can be exceptionally informative markers for investigating species and population-level evolutionary history. While several studies have utilized retrotransposon-based insertional polymorphisms to resolve phylogenies, few population studies exist outside of humans. Endogenous retroviruses are LTR-retrotransposons derived from retroviruses that have become stably integrated in the host genome during past infections and transmitted vertically to subsequent generations. They offer valuable insight into host-virus co-evolution and a unique perspective on host evolutionary history because they integrate into the genome at a discrete point in time. We examined the evolutionary history of a cervid endogenous gammaretrovirus (CrERVγ) in mule deer (Odocoileus hemionus). We sequenced 14 CrERV proviruses (CrERV-in1 to -in14), and examined the prevalence and distribution of 13 proviruses in 262 deer among 15 populations from Montana, Wyoming, and Utah. CrERV absence in white-tailed deer (O. virginianus), identical 5′ and 3′ long terminal repeat (LTR) sequences, insertional polymorphism, and CrERV divergence time estimates indicated that most endogenization events occurred within the last 200000 years. Population structure inferred from CrERVs (F ST = 0.008) and microsatellites (θ = 0.01) was low, but significant, with Utah, northwestern Montana, and a Helena herd being particularly differentiated. Clustering analyses indicated regional structuring, and non-contiguous clustering could often be explained by known translocations. Cluster ensemble results indicated spatial localization of viruses, specifically in deer from northeastern and western Montana. This study demonstrates the utility of endogenous retroviruses to elucidate and provide novel insight into both ERV evolutionary history and the history of contemporary host populations. PMID:24336966

The population history of endogenous retroviruses in mule deer (Odocoileus heminous)

USGS Publications Warehouse

Kamath, Pauline L.; Elleder, Daniel; Bao, Le; Cross, Paul C.; Powell, John H.; Poss, Mary

2013-01-01

Mobile elements are powerful agents of genomic evolution and can be exceptionally informative markers for investigating species and population-level evolutionary history. While several studies have utilized retrotransposon-based insertional polymorphisms to resolve phylogenies, few population studies exist outside of humans. Endogenous retroviruses are LTR-retrotransposons derived from retroviruses that have become stably integrated in the host genome during past infections and transmitted vertically to subsequent generations. They offer valuable insight into host-virus co-evolution and a unique perspective on host evolutionary history because they integrate into the genome at a discrete point in time. We examined the evolutionary history of a cervid endogenous gammaretrovirus (CrERVγ) in mule deer (Odocoileus hemionus). We sequenced 14 CrERV proviruses (CrERV-in1 to -in14), and examined the prevalence and distribution of 13 proviruses in 262 deer among 15 populations from Montana, Wyoming, and Utah. CrERV absence in white-tailed deer (O. virginianus), identical 5′ and 3′ long terminal repeat (LTR) sequences, insertional polymorphism, and CrERV divergence time estimates indicated that most endogenization events occurred within the last 200000 years. Population structure inferred from CrERVs (F ST = 0.008) and microsatellites (θ = 0.01) was low, but significant, with Utah, northwestern Montana, and a Helena herd being particularly differentiated. Clustering analyses indicated regional structuring, and non-contiguous clustering could often be explained by known translocations. Cluster ensemble results indicated spatial localization of viruses, specifically in deer from northeastern and western Montana. This study demonstrates the utility of endogenous retroviruses to elucidate and provide novel insight into both ERV evolutionary history and the history of contemporary host populations.

Presence of a Shared 5'-Leader Sequence in Ancestral Human and Mammalian Retroviruses and Its Transduction into Feline Leukemia Virus.

PubMed

Kawasaki, Junna; Kawamura, Maki; Ohsato, Yoshiharu; Ito, Jumpei; Nishigaki, Kazuo

2017-10-15

Recombination events induce significant genetic changes, and this process can result in virus genetic diversity or in the generation of novel pathogenicity. We discovered a new recombinant feline leukemia virus (FeLV) gag gene harboring an unrelated insertion, termed the X region, which was derived from Felis catus endogenous gammaretrovirus 4 (FcERV-gamma4). The identified FcERV-gamma4 proviruses have lost their coding capabilities, but some can express their viral RNA in feline tissues. Although the X-region-carrying recombinant FeLVs appeared to be replication-defective viruses, they were detected in 6.4% of tested FeLV-infected cats. All isolated recombinant FeLV clones commonly incorporated a middle part of the FcERV-gamma4 5'-leader region as an X region. Surprisingly, a sequence corresponding to the portion contained in all X regions is also present in at least 13 endogenous retroviruses (ERVs) observed in the cat, human, primate, and pig genomes. We termed this shared genetic feature the commonly shared (CS) sequence. Despite our phylogenetic analysis indicating that all CS-sequence-carrying ERVs are classified as gammaretroviruses, no obvious closeness was revealed among these ERVs. However, the Shannon entropy in the CS sequence was lower than that in other parts of the provirus genome. Notably, the CS sequence of human endogenous retrovirus T had 73.8% similarity with that of FcERV-gamma4, and specific signals were detected in the human genome by Southern blot analysis using a probe for the FcERV-gamma4 CS sequence. Our results provide an interesting evolutionary history for CS-sequence circulation among several distinct ancestral viruses and a novel recombined virus over a prolonged period. IMPORTANCE Recombination among ERVs or modern viral genomes causes a rapid evolution of retroviruses, and this phenomenon can result in the serious situation of viral disease reemergence. We identified a novel recombinant FeLV gag gene that contains an unrelated

MCP-1 and CCR2 Contribute to Non-Lymphocyte-Mediated Brain Disease Induced by Fr98 Polytropic Retrovirus Infection in Mice: Role for Astrocytes in Retroviral Neuropathogenesis

PubMed Central

Peterson, Karin E.; Errett, John S.; Wei, Tao; Dimcheff, Derek E.; Ransohoff, Richard; Kuziel, William A.; Evans, Leonard; Chesebro, Bruce

2004-01-01

Virus infection of the central nervous system (CNS) often results in chemokine upregulation. Although often associated with lymphocyte recruitment, increased chemokine expression is also associated with non-lymphocyte-mediated CNS disease. In these instances, the effect of chemokine upregulation on neurological disease is unclear. In vitro, several chemokines including monocyte chemotactic protein 1 (MCP-1) protect neurons from apoptosis. Therefore, in vivo, chemokine upregulation may be a protective host response to CNS damage. Alternatively, chemokines may contribute to pathogenesis by stimulating intrinsic brain cells or recruiting macrophages to the brain. To investigate these possibilities, we studied a neurovirulent retrovirus, Fr98, that induces severe non-lymphocyte-mediated neurological disease and causes the upregulation of several chemokines that bind to chemokine receptors CCR2 and CCR5. Knockout mice deficient in CCR2 had reduced susceptibility to Fr98 pathogenesis, with significantly fewer mice developing clinical disease than did wild-type controls. In contrast, no reduction in Fr98-induced disease was observed in CCR5 knockout mice. Thus, signaling through CCR2, but not CCR5, plays an important role in Fr98-mediated pathogenesis. Three ligands for CCR2 (MCP-1, MCP-3, and MCP-5) were upregulated during Fr98 infection of the brain. Antibody-blocking experiments demonstrated that MCP-1 was important for retrovirus-induced neurological disease. In situ hybridization analysis revealed that MCP-1 was expressed by glial fibrillary acidic protein-positive astrocytes. Thus, astrocytes, previously not thought to play an effector role in the disease process were found to contribute to pathogenesis through the production of MCP-1. This study also demonstrates that chemokines can mediate pathogenesis in the CNS in the absence of lymphocytic infiltrate and gives credence to the hypothesis that chemokine upregulation is a mechanism by which retroviruses such as

Human Endogenous Retrovirus HERV-Fc1 Association with Multiple Sclerosis Susceptibility: A Meta-Analysis

PubMed Central

García-Montojo, Marta; Alcina, Antonio; Fedetz, María; Alloza, Iraide; Astobiza, Ianire; Leyva, Laura; Fernández, Oscar; Izquierdo, Guillermo; Antigüedad, Alfredo; Arroyo, Rafael; Álvarez-Lafuente, Roberto; Vandenbroeck, Koen; Matesanz, Fuencisla; Urcelay, Elena

2014-01-01

Background Human endogenous retroviruses (HERVs) are repetitive sequences derived from ancestral germ-line infections by exogenous retroviruses and different HERV families have been integrated in the genome. HERV-Fc1 in chromosome X has been previously associated with multiple sclerosis (MS) in Northern European populations. Additionally, HERV-Fc1 RNA levels of expression have been found increased in plasma of MS patients with active disease. Considering the North-South latitude gradient in MS prevalence, we aimed to evaluate the role of HERV-Fc1on MS risk in three independent Spanish cohorts. Methods A single nucleotide polymorphism near HERV-Fc1, rs391745, was genotyped by Taqman chemistry in a total of 2473 MS patients and 3031 ethnically matched controls, consecutively recruited from: Northern (569 patients and 980 controls), Central (883 patients and 692 controls) and Southern (1021 patients and 1359 controls) Spain. Our results were pooled in a meta-analysis with previously published data. Results Significant associations of the HERV-Fc1 polymorphism with MS were observed in two Spanish cohorts and the combined meta-analysis with previous data yielded a significant association [rs391745 C-allele carriers: pM-H = 0.0005; ORM-H (95% CI) = 1.27 (1.11–1.45)]. Concordantly to previous findings, when the analysis was restricted to relapsing remitting and secondary progressive MS samples, a slight enhancement in the strength of the association was observed [pM-H = 0.0003, ORM-H (95% CI) = 1.32 (1.14–1.53)]. Conclusion Association of the HERV-Fc1 polymorphism rs391745 with bout-onset MS susceptibility was confirmed in Southern European cohorts. PMID:24594754

The Env-like open reading frame of the baculovirus-integrated retrotransposon TED encodes a retrovirus-like envelope protein.

PubMed

Ozers, M S; Friesen, P D

1996-12-15

TED is a 7.5-kbp member of the gypsy family of retrotransposons that was first identified by its integration within the baculovirus DNA genome. This lepidopteran (moth) transposon contains three retrovirus-like genes, including functional gag and pol that yield reverse transcriptase-containing virus-like particles. To identify and characterize the product(s) of the third env-like open reading frame, TED ORF3 was expressed in homologous lepidopteran cells by using a baculovirus vector, vENV. Immunoblots and immunoprecipitations with antiserum raised against a bacterial ORF3-fusion protein detected two ORF3-encoded proteins, p68env and gp75env. On the basis of selective incorporation of [3H]mannose and inhibition of modification by tunicamycin which blocks N-linked glycosylation, gp75env is a glycoprotein derived from core precursor p68env. As predicted by the presence of a transmembrane domain near the carboxyl terminus, both p68env and gp75env were associated with heavy membranes of vENV-infected cells. Thus, TED ORF3 encodes a membrane glycoprotein with properties characteristic of retroviral env proteins. These data are consistent with the hypothesis that TED is an invertebrate retrovirus. Moreover, TED integration within the baculovirus genome provides an example of retroelement-mediated acquisition of host genes that may contribute to virus evolution.

A survey of endogenous retrovirus (ERV) sequences in the vicinity of multiple sclerosis (MS)-associated single nucleotide polymorphisms (SNPs).

PubMed

Brütting, Christine; Emmer, Alexander; Kornhuber, Malte; Staege, Martin S

2016-08-01

Although multiple sclerosis (MS) is one of the most common central nervous system diseases in young adults, little is known about its etiology. Several human endogenous retroviruses (ERVs) are considered to play a role in MS. We are interested in which ERVs can be identified in the vicinity of MS associated genetic marker to find potential initiators of MS. We analysed the chromosomal regions surrounding 58 single nucleotide polymorphisms (SNPs) that are associated with MS identified in one of the last major genome wide association studies. We scanned these regions for putative endogenous retrovirus sequences with large open reading frames (ORFs). We observed that more retrovirus-related putative ORFs exist in the relatively close vicinity of SNP marker indices in multiple sclerosis compared to control SNPs. We found very high homologies to HERV-K, HCML-ARV, XMRV, Galidia ERV, HERV-H/env62 and XMRV-like mouse endogenous retrovirus mERV-XL. The associated genes (CYP27B1, CD6, CD58, MPV17L2, IL12RB1, CXCR5, PTGER4, TAGAP, TYK2, ICAM3, CD86, GALC, GPR65 as well as the HLA DRB1*1501) are mainly involved in the immune system, but also in vitamin D regulation. The most frequently detected ERV sequences are related to the multiple sclerosis-associated retrovirus, the human immunodeficiency virus 1, HERV-K, and the Simian foamy virus. Our data shows that there is a relation between MS associated SNPs and the number of retroviral elements compared to control. Our data identifies new ERV sequences that have not been associated with MS, so far.

Regulation of the Human Endogenous Retrovirus K (HML-2) Transcriptome by the HIV-1 Tat Protein

PubMed Central

Gonzalez-Hernandez, Marta J.; Cavalcoli, James D.; Sartor, Maureen A.; Contreras-Galindo, Rafael; Meng, Fan; Dai, Manhong; Dube, Derek; Saha, Anjan K.; Gitlin, Scott D.; Omenn, Gilbert S.; Kaplan, Mark H.

2014-01-01

ABSTRACT Approximately 8% of the human genome is made up of endogenous retroviral sequences. As the HIV-1 Tat protein activates the overall expression of the human endogenous retrovirus type K (HERV-K) (HML-2), we used next-generation sequencing to determine which of the 91 currently annotated HERV-K (HML-2) proviruses are regulated by Tat. Transcriptome sequencing of total RNA isolated from Tat- and vehicle-treated peripheral blood lymphocytes from a healthy donor showed that Tat significantly activates expression of 26 unique HERV-K (HML-2) proviruses, silences 12, and does not significantly alter the expression of the remaining proviruses. Quantitative reverse transcription-PCR validation of the sequencing data was performed on Tat-treated PBLs of seven donors using provirus-specific primers and corroborated the results with a substantial degree of quantitative similarity. IMPORTANCE The expression of HERV-K (HML-2) is tightly regulated but becomes markedly increased following infection with HIV-1, in part due to the HIV-1 Tat protein. The findings reported here demonstrate the complexity of the genome-wide regulation of HERV-K (HML-2) expression by Tat. This work also demonstrates that although HERV-K (HML-2) proviruses in the human genome are highly similar in terms of DNA sequence, modulation of the expression of specific proviruses in a given biological situation can be ascertained using next-generation sequencing and bioinformatics analysis. PMID:24872592

Mutational definition of functional domains within the Rev homolog encoded by human endogenous retrovirus K.

PubMed

Bogerd, H P; Wiegand, H L; Yang, J; Cullen, B R

2000-10-01

Nuclear export of the incompletely spliced mRNAs encoded by several complex retroviruses, including human immunodeficiency virus type 1 (HIV-1), is dependent on a virally encoded adapter protein, termed Rev in HIV-1, that directly binds both to a cis-acting viral RNA target site and to the cellular Crm1 export factor. Human endogenous retrovirus K, a family of ancient endogenous retroviruses that is not related to the exogenous retrovirus HIV-1, was recently shown to also encode a Crm1-dependent nuclear RNA export factor, termed K-Rev. Although HIV-1 Rev and K-Rev display little sequence identity, they share the ability not only to bind to Crm1 and to RNA but also to form homomultimers and shuttle between nucleus and cytoplasm. We have used mutational analysis to identify sequences in the 105-amino-acid K-Rev protein required for each of these distinct biological activities. While mutations in K-Rev that inactivate any one of these properties also blocked K-Rev-dependent nuclear RNA export, several K-Rev mutants were comparable to wild type when assayed for any of these individual activities yet nevertheless defective for RNA export. Although several nonfunctional K-Rev mutants acted as dominant negative inhibitors of K-Rev-, but not HIV-1 Rev-, dependent RNA export, these were not defined by their inability to bind to Crm1, as is seen with HIV-1 Rev. In total, this analysis suggests a functional architecture for K-Rev that is similar to, but distinct from, that described for HIV-1 Rev and raises the possibility that viral RNA export mediated by the approximately 25 million-year-old K-Rev protein may require an additional cellular cofactor that is not required for HIV-1 Rev function.

Mechanisms employed by retroviruses to exploit host factors for translational control of a complicated proteome

PubMed Central

Bolinger, Cheryl; Boris-Lawrie, Kathleen

2009-01-01

Retroviruses have evolved multiple strategies to direct the synthesis of a complex proteome from a single primary transcript. Their mechanisms are modulated by a breadth of virus-host interactions, which are of significant fundamental interest because they ultimately affect the efficiency of virus replication and disease pathogenesis. Motifs located within the untranslated region (UTR) of the retroviral RNA have established roles in transcriptional trans-activation, RNA packaging, and genome reverse transcription; and a growing literature has revealed a necessary role of the UTR in modulating the efficiency of viral protein synthesis. Examples include a 5' UTR post-transcriptional control element (PCE), present in at least eight retroviruses, that interacts with cellular RNA helicase A to facilitate cap-dependent polyribosome association; and 3' UTR constitutive transport element (CTE) of Mason-Pfizer monkey virus that interacts with Tap/NXF1 and SR protein 9G8 to facilitate RNA export and translational utilization. By contrast, nuclear protein hnRNP E1 negatively modulates HIV-1 Gag, Env, and Rev protein synthesis. Alternative initiation strategies by ribosomal frameshifting and leaky scanning enable polycistronic translation of the cap-dependent viral transcript. Other studies posit cap-independent translation initiation by internal ribosome entry at structural features of the 5' UTR of selected retroviruses. The retroviral armamentarium also commands mechanisms to counter cellular post-transcriptional innate defenses, including protein kinase R, 2',5'-oligoadenylate synthetase and the small RNA pathway. This review will discuss recent and historically-recognized insights into retrovirus translational control. The expanding knowledge of retroviral post-transcriptional control is vital to understanding the biology of the retroviral proteome. In a broad perspective, each new insight offers a prospective target for antiviral therapy and strategic improvement of gene

Gene therapy using retrovirus vectors: vector development and biosafety at clinical trials.

PubMed

Doi, Knayo; Takeuchi, Yasuhiro

2015-01-01

Retrovirus vectors (gammaretroviral and lentiviral vectors) have been considered as promising tools to transfer therapeutic genes into patient cells because they can permanently integrate into host cellular genome. To treat monogenic, inherited diseases, retroviral vectors have been used to add correct genes into patient cells. Conventional gammaretroviral vectors achieved successful results in clinical trials: treated patients had therapeutic gene expression in target cells and had improved symptoms of diseases. However, serious side-effects of leukemia occurred, caused by retroviral insertional mutagenesis (IM). These incidences stressed the importance of monitoring vector integration sites in patient cells as well as of re-consideration on safer vectors. More recently lentiviral vectors which can deliver genes into non-dividing cells started to be used in clinical trials including neurological disorders, showing their efficacy. Vector integration site analysis revealed that lentiviruses integrate less likely to near promoter regions of oncogenes than gammaretroviruses and no adverse events have been reported in lentiviral vector-mediated gene therapy clinical trials. Therefore lentiviral vectors have promises to be applied to a wide range of common diseases in near future. For example, T cells from cancer patients were transduced to express chimeric T cell receptors recognizing their tumour cells enhancing patients' anti-cancer immunity.

HTLV-3/4 and simian foamy retroviruses in humans: discovery, epidemiology, cross-species transmission and molecular virology.

PubMed

Gessain, Antoine; Rua, Réjane; Betsem, Edouard; Turpin, Jocelyn; Mahieux, Renaud

2013-01-05

Non-human primates are considered to be likely sources of viruses that can infect humans and thus pose a significant threat to human population. This is well illustrated by some retroviruses, as the simian immunodeficiency viruses and the simian T lymphotropic viruses, which have the ability to cross-species, adapt to a new host and sometimes spread. This leads to a pandemic situation for HIV-1 or an endemic one for HTLV-1. Here, we present the available data on the discovery, epidemiology, cross-species transmission and molecular virology of the recently discovered HTLV-3 and HTLV-4 deltaretroviruses, as well as the simian foamy retroviruses present in different human populations at risk, especially in central African hunters. We discuss also the natural history in humans of these retroviruses of zoonotic origin (magnitude and geographical distribution, possible inter-human transmission). In Central Africa, the increase of the bushmeat trade during the last decades has opened new possibilities for retroviral emergence in humans, especially in immuno-compromised persons. Copyright © 2012 Elsevier Inc. All rights reserved.

Ancestral Mutations Acquired in Refrex-1, a Restriction Factor against Feline Retroviruses, during its Cooption and Domestication

PubMed Central

Ito, Jumpei; Baba, Takuya; Kawasaki, Junna

2015-01-01

ABSTRACT Endogenous retroviruses (ERVs) are remnants of ancestral retroviral infections of germ cells. Retroviral endogenization is an adaptation process for the host genome, and ERVs are gradually attenuated or inactivated by mutation. However, some ERVs that have been “domesticated” by their hosts eventually gain physiological functions, such as placentation or viral resistance. We previously reported the discovery of Refrex-1, a soluble antiretroviral factor in domestic cats that specifically inhibits infection by feline leukemia virus subgroup D (FeLV-D), a chimeric virus of FeLV, and a feline ERV, ERV-DC. Refrex-1 is a truncated envelope protein (Env) encoded by both ERV-DC7 and ERV-DC16 proviral loci. Here, we reconstituted ancestral and functional Env from ERV-DC7 and ERV-DC16 envelope genes (env) by inducing reverse mutations. Unexpectedly, ERV-DC7 and ERV-DC16 full-length Env (ERV-DC7 fl and ERV-DC16 fl), reconstructed by removing stop codons, did not produce infectious viral particles. ERV-DC7 fl and ERV-DC16 fl were highly expressed in cells but were not cleaved into surface subunits (SU) and transmembrane subunits, nor were they incorporated into virions. G407R/N427I-A429T and Y431D substitutions within the SU C-terminal domain of ERV-DC7 fl and ERV-DC16 fl, respectively, caused these dysfunctions. The residues glycine 407 and tyrosine 431 are relatively conserved among infectious gammaretroviruses, and their substitution causes the same dysfunctions as the tested retroviruses. Our results reveal that specific mutations within the SU C-terminal domain suppressed Env cleavage and incorporation into virions and indicate that these mutations contributed to the domestication of Refrex-1 through multistep events that occurred in the postintegration period. IMPORTANCE Domestic cats are colonized with various exogenous retroviruses (exRVs), such as feline leukemia virus (FeLV), and their genomes contain numerous ERVs, some of which are replication

The discovery of HTLV-1, the first pathogenic human retrovirus.

PubMed

Coffin, John M

2015-12-22

After the discovery of retroviral reverse transcriptase in 1970, there was a flurry of activity, sparked by the "War on Cancer," to identify human cancer retroviruses. After many false claims resulting from various artifacts, most scientists abandoned the search, but the Gallo laboratory carried on, developing both specific assays and new cell culture methods that enabled them to report, in the accompanying 1980 PNAS paper, identification and partial characterization of human T-cell leukemia virus (HTLV; now known as HTLV-1) produced by a T-cell line from a lymphoma patient. Follow-up studies, including collaboration with the group that first identified a cluster of adult T-cell leukemia (ATL) cases in Japan, provided conclusive evidence that HTLV was the cause of this disease. HTLV-1 is now known to infect at least 4-10 million people worldwide, about 5% of whom will develop ATL. Despite intensive research, knowledge of the viral etiology has not led to improvement in treatment or outcome of ATL. However, the technology for discovery of HTLV and acknowledgment of the existence of pathogenic human retroviruses laid the technical and intellectual foundation for the discovery of the cause of AIDS soon afterward. Without this advance, our ability to diagnose and treat HIV infection most likely would have been long delayed.

Host Species Barriers to Jaagsiekte Sheep Retrovirus Replication and Carcinogenesis

PubMed Central

Martineau, Henny; De las Heras, Marcelo; Murgia, Claudio; Huang, Robert; Centorame, Patrizia; Di Francesco, Gabriella; Di Gialleonardo, Luigina; Spencer, Thomas E.; Griffiths, David J.; Palmarini, Massimo

2013-01-01

Understanding the factors governing host species barriers to virus transmission has added significantly to our appreciation of virus pathogenesis. Jaagsiekte sheep retrovirus (JSRV) is the causative agent of ovine pulmonary adenocarcinoma (OPA), a transmissible lung cancer of sheep that has rarely been found in goats. In this study, in order to further clarify the pathogenesis of OPA, we investigated whether goats are resistant to JSRV replication and carcinogenesis. We found that JSRV induces lung tumors in goats with macroscopic and histopathological features that dramatically differ from those in sheep. However, the origins of the tumor cells in the two species are identical. Interestingly, in experimentally infected lambs and goat kids, we revealed major differences in the number of virus-infected cells at early stages of infection. These differences were not related to the number of available target cells for virus infection and cell transformation or the presence of a host-specific immune response toward JSRV. Indeed, we also found that goats possess transcriptionally active endogenous retroviruses (enJSRVs) that likely influence the host immune response toward the exogenous JSRV. Overall, these results suggest that goat cells, or at least those cells targeted for viral carcinogenesis, are not permissive to virus replication but can be transformed by JSRV. PMID:23903827

Human endogenous retrovirus rec interferes with germ cell development in mice and may cause carcinoma in situ, the predecessor lesion of germ cell tumors.

PubMed

Galli, Uwe M; Sauter, Marlies; Lecher, Bernd; Maurer, Simone; Herbst, Hermann; Roemer, Klaus; Mueller-Lantzsch, Nikolaus

2005-04-28

Germ cell tumors (GCTs) are among the most common malignancies in young men. We have previously documented that patients with GCT frequently produce serum antibodies directed against proteins encoded by human endogenous retrovirus (HERV) type K sequences. Transcripts originating from the env gene of HERV-K, including the rec-relative of human immunodeficiency virus rev, are highly expressed in GCTs. We report here that mice that inducibly express HERV-K rec show a disturbed germ cell development and may exhibit, by 19 months of age, changes reminiscent of carcinoma in situ, the predecessor lesion of classic seminoma in humans. This provides the first direct evidence that the expression of a human endogenous retroviral gene previously established as a marker in human germ cell tumors may contribute to organ-specific tumorigenesis in a transgenic mouse model.

Gene expression analysis of murine cells producing amphotropic mouse leukaemia virus at a cultivation temperature of 32 and 37 degrees C.

PubMed

Beer, Christiane; Buhr, Petra; Hahn, Heidi; Laubner, Daniela; Wirth, Manfred

2003-07-01

Cultivation of retrovirus packaging cells at 32 degrees C represents a common procedure to achieve high titres in mouse retrovirus production. Gene expression profiling of mouse NIH 3T3 cells producing amphotropic mouse leukaemia virus 4070A revealed that 10 % of the 1176 cellular genes investigated were regulated by temperature shift (37/32 degrees C), while 5 % were affected by retrovirus infection. Strikingly, retrovirus production at 32 degrees C activated the cholesterol biosynthesis/transport pathway and caused an increase in plasma membrane cholesterol levels. Furthermore, these conditions resulted in transcriptional activation of smoothened (smo), patched (ptc) and gli-1; Smo, Ptc and Gli-1, as well as cholesterol, are components of the Sonic hedgehog (Shh) signalling pathway, which directs pattern formation, diversification and tumourigenesis in mammalian cells. These findings suggest a link between cultivation at 32 degrees C, production of MLV-A and the Shh signalling pathway.

Postinhibitory rebound neurons and networks are disrupted in retrovirus-induced spongiform neurodegeneration

PubMed Central

Li, Ying; Davey, Robert A.; Lynch, William P.

2014-01-01

Certain retroviruses induce progressive spongiform motor neuron disease with features resembling prion diseases and amyotrophic lateral sclerosis. With the neurovirulent murine leukemia virus (MLV) FrCasE, Env protein expression within glia leads to postsynaptic vacuolation, cellular effacement, and neuronal loss in the absence of neuroinflammation. To understand the physiological changes associated with MLV-induced spongiosis, and its neuronal specificity, we employed patch-clamp recordings and voltage-sensitive dye imaging in brain slices of the mouse inferior colliculus (IC), a midbrain nucleus that undergoes extensive spongiosis. IC neurons characterized by postinhibitory rebound firing (PIR) were selectively affected in FrCasE-infected mice. Coincident with Env expression in microglia and in glia characterized by NG2 proteoglycan expression (NG2 cells), rebound neurons (RNs) lost PIR, became hyperexcitable, and were reduced in number. PIR loss and hyperexcitability were reversed by raising internal calcium buffer concentrations in RNs. PIR-initiated rhythmic circuits were disrupted, and spontaneous synchronized bursting and prolonged depolarizations were widespread. Other IC neuron cell types and circuits within the same degenerative environment were unaffected. Antagonists of NMDA and/or AMPA receptors reduced burst firing in the IC but did not affect prolonged depolarizations. Antagonists of L-type calcium channels abolished both bursts and slow depolarizations. IC infection by the nonneurovirulent isogenic virus Friend 57E (Fr57E), whose Env protein is structurally similar to FrCasE, showed no RN hyperactivity or cell loss; however, PIR latency increased. These findings suggest that spongiform neurodegeneration arises from the unique excitability of RNs, their local regulation by glia, and the disruption of this relationship by glial expression of abnormal protein. PMID:25252336

The human retrovirus XMRV in prostate cancer and chronic fatigue syndrome.

PubMed

Silverman, Robert H; Nguyen, Carvell; Weight, Christopher J; Klein, Eric A

2010-07-01

Xenotropic murine leukemia virus-related virus (XMRV) is an authentic, newly recognized human retrovirus first identified in prostate cancer tissues from men with a deficiency in the innate immunity gene RNASEL. At present, studies have detected XMRV at widely different rates in prostate cancer cases (0-27%) and in patients with chronic fatigue syndrome (CFS; 0-67%). Indirect or direct modes of carcinogenesis by XMRV have been suggested depending on whether the virus was found in stroma or malignant epithelium. Viral replication in the prostate might be affected by androgens, which stimulate XMRV through a transcriptional enhancer site in viral DNA. By contrast, host restriction factors, such as APOBEC3 and tetherin, inhibit virus replication. Immune dysfunction mediated by XMRV has been suggested as a possible factor in CFS. Recent studies show that some existing antiretroviral drugs suppress XMRV infections and diagnostic assays are under development. Although other retroviruses of the same genus as XMRV (gammaretroviruses) cause cancer and neurological disease in animals, whether XMRV is a cause of either prostate cancer or CFS remains unknown. Emerging science surrounding XMRV is contributing to our knowledge of retroviral infections while focusing intense interest on two major human diseases.

Retinoic acid-induced differentiation of retrovirus-infected HL-60 cells is associated with enhanced transcription from the viral long terminal repeat

DOE Office of Scientific and Technical Information (OSTI.GOV)

Collins, S.J.

1988-11-01

The author infected different human leukemic cell lines with an amphotropic retrovirus vector (designated PA317/N2) which confers G418 resistance and contains the Moloney murine leukemia virus long terminal repeat. In retrovirus-infected G418-resistant HL-60 cells, induction of granulocyte differentiation by retinoic acid was invariably accompanied by a marked increase (5- to 10-fold) in the transcriptional activity of the integrated retroviral long terminal repeat.

Characterization of retroviral infectivity and superinfection resistance during retrovirus-mediated transduction of mammalian cells.

PubMed

Liao, J; Wei, Q; Fan, J; Zou, Y; Song, D; Liu, J; Liu, F; Ma, C; Hu, X; Li, L; Yu, Y; Qu, X; Chen, L; Yu, X; Zhang, Z; Zhao, C; Zeng, Z; Zhang, R; Yan, S; Wu, T; Wu, X; Shu, Y; Lei, J; Li, Y; Zhang, W; Wang, J; Reid, R R; Lee, M J; Huang, W; Wolf, J M; He, T-C; Wang, J

2017-06-01

Retroviral vectors including lentiviral vectors are commonly used tools to stably express transgenes or RNA molecules in mammalian cells. Their utilities are roughly divided into two categories, stable overexpression of transgenes and RNA molecules, which requires maximal transduction efficiency, or functional selection with retrovirus (RV)-based libraries, which takes advantage of retroviral superinfection resistance. However, the dynamic features of RV-mediated transduction are not well characterized. Here, we engineered two murine stem cell virus-based retroviral vectors expressing dual fluorescence proteins and antibiotic markers, and analyzed virion production efficiency and virion stability, dynamic infectivity and superinfection resistance in different cell types, and strategies to improve transduction efficiency. We found that the highest virion production occurred between 60 and 72 h after transfection. The stability of the collected virion supernatant decreased by >60% after 3 days in storage. We found that RV infectivity varied drastically in the tested human cancer lines, while low transduction efficiency was partially overcome with increased virus titer, prolonged infection duration and/or repeated infections. Furthermore, we demonstrated that RV receptors PIT1 and PIT2 were lowly expressed in the analyzed cells, and that PIT1 and/or PIT2 overexpression significantly improved transduction efficiency in certain cell lines. Thus, our findings provide resourceful information for the optimal conditions of retroviral-mediated gene delivery.

75 FR 79006 - Submission for OMB Review; Comment Request; Transfusion-Transmitted Retrovirus and Hepatitis...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-12-17

...- control study design. (3) Determine nationally-representative infectious disease marker prevalence and... control number. Proposed Collection: Title: Transfusion-transmitted retrovirus and hepatitis virus rates... factors in blood donors as assessed using analytical study designs is largely unavailable in the U.S...

Phylogenetic Diversity of Koala Retrovirus within a Wild Koala Population.

PubMed

Chappell, K J; Brealey, J C; Amarilla, A A; Watterson, D; Hulse, L; Palmieri, C; Johnston, S D; Holmes, E C; Meers, J; Young, P R

2017-02-01

Koala populations are in serious decline across many areas of mainland Australia, with infectious disease a contributing factor. Koala retrovirus (KoRV) is a gammaretrovirus present in most wild koala populations and captive colonies. Five subtypes of KoRV (A to E) have been identified based on amino acid sequence divergence in a hypervariable region of the receptor binding domain of the envelope protein. However, analysis of viral genetic diversity has been conducted primarily on KoRV in captive koalas housed in zoos in Japan, the United States, and Germany. Wild koalas within Australia have not been comparably assessed. Here we report a detailed analysis of KoRV genetic diversity in samples collected from 18 wild koalas from southeast Queensland. By employing deep sequencing we identified 108 novel KoRV envelope sequences and determined their phylogenetic diversity. Genetic diversity in KoRV was abundant and fell into three major groups; two comprised the previously identified subtypes A and B, while the third contained the remaining hypervariable region subtypes (C, D, and E) as well as four hypervariable region subtypes that we newly define here (F, G, H, and I). In addition to the ubiquitous presence of KoRV-A, which may represent an exclusively endogenous variant, subtypes B, D, and F were found to be at high prevalence, while subtypes G, H, and I were present in a smaller number of animals. Koala retrovirus (KoRV) is thought to be a significant contributor to koala disease and population decline across mainland Australia. This study is the first to determine KoRV subtype prevalence among a wild koala population, and it significantly expands the total number of KoRV sequences available, providing a more precise picture of genetic diversity. This understanding of KoRV subtype prevalence and genetic diversity will be important for conservation efforts attempting to limit the spread of KoRV. Furthermore, KoRV is one of the only retroviruses shown to exist in

Neuropeptide Y Has a Protective Role during Murine Retrovirus-Induced Neurological Disease▿

PubMed Central

Du, Min; Butchi, Niranjan B.; Woods, Tyson; Morgan, Timothy W.; Peterson, Karin E.

2010-01-01

Viral infections in the central nervous system (CNS) can lead to neurological disease either directly by infection of neurons or indirectly through activation of glial cells and production of neurotoxic molecules. Understanding the effects of virus-mediated insults on neuronal responses and neurotrophic support is important in elucidating the underlying mechanisms of viral diseases of the CNS. In the current study, we examined the expression of neurotrophin- and neurotransmitter-related genes during infection of mice with neurovirulent polytropic retrovirus. In this model, virus-induced neuropathogenesis is indirect, as the virus predominantly infects macrophages and microglia and does not productively infect neurons or astrocytes. Virus infection is associated with glial cell activation and the production of proinflammatory cytokines in the CNS. In the current study, we identified increased expression of neuropeptide Y (NPY), a pleiotropic growth factor which can regulate both immune cells and neuronal cells, as a correlate with neurovirulent virus infection. Increased levels of Npy mRNA were consistently associated with neurological disease in multiple strains of mice and were induced only by neurovirulent, not avirulent, virus infection. NPY protein expression was primarily detected in neurons near areas of virus-infected cells. Interestingly, mice deficient in NPY developed neurological disease at a faster rate than wild-type mice, indicating a protective role for NPY. Analysis of NPY-deficient mice indicated that NPY may have multiple mechanisms by which it influences virus-induced neurological disease, including regulating the entry of virus-infected cells into the CNS. PMID:20702619

Modulation of cGMP by human HO-1 retrovirus gene transfer in pulmonary microvessel endothelial cells.

PubMed

Abraham, Nader G; Quan, Shuo; Mieyal, Paul A; Yang, Liming; Burke-Wolin, Theresa; Mingone, Christopher J; Goodman, Alvin I; Nasjletti, Alberto; Wolin, Michael S

2002-11-01

Carbon monoxide (CO) stimulates guanylate cyclase (GC) and increases guanosine 3',5'-cyclic monophosphate (cGMP) levels. We transfected rat-lung pulmonary endothelial cells with a retrovirus-mediated human heme oxygenase (hHO)-1 gene. Pulmonary cells that expressed hHO-1 exhibited a fourfold increase in HO activity associated with decreases in the steady-state levels of heme and cGMP without changes in soluble GC (sGC) and endothelial nitric oxide synthase (NOS) proteins or basal nitrite production. Heme elicited significant increases in CO production and intracellular cGMP levels in both pulmonary endothelial and pulmonary hHO-1-expressing cells. N(omega)-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NOS, significantly decreased cGMP levels in heme-treated pulmonary endothelial cells but not heme-treated hHO-1-expressing cells. In the presence of exogenous heme, CO and cGMP levels in hHO-1-expressing cells exceeded the corresponding levels in pulmonary endothelial cells. Acute exposure of endothelial cells to SnCl2, which is an inducer of HO-1, increased cGMP levels, whereas chronic exposure decreased heme and cGMP levels. These results indicate that prolonged overexpression of HO-1 ultimately decreases sGC activity by limiting the availability of cellular heme. Heme activates sGC and enhances cGMP levels via a mechanism that is largely insensitive to NOS inhibition.

Multiple invasions of an infectious retrovirus in cat genomes

PubMed Central

Shimode, Sayumi; Nakagawa, So; Miyazawa, Takayuki

2015-01-01

Endogenous retroviruses (ERVs) are remnants of ancient retroviral infections of host germ-line cells. While most ERVs are defective, some are active and express viral proteins. The RD-114 virus is a replication-competent feline ERV, and several feline cell lines produce infectious RD-114 viral particles. All domestic cats are considered to have an ERV locus encoding a replication-competent RD-114 virus in their genomes; however, the locus has not been identified. In this study, we investigated RD-114 virus-related proviral loci in genomes of domestic cats, and found that none were capable of producing infectious viruses. We also found that all domestic cats have an RD-114 virus-related sequence on chromosome C2, termed RDRS C2a, but populations of the other RDRSs are different depending on the regions where cats live or breed. Our results indicate that RDRS C2a, the oldest RD-114-related provirus, entered the host genome before an ancestor of domestic cats started diverging and the other new RDRSs might have integrated into migrating cats in Europe. We also show that infectious RD-114 virus can be resurrected by the recombination between two non-infectious RDRSs. From these data, we conclude that cats do not harbor infectious RD-114 viral loci in their genomes and RD-114-related viruses invaded cat genomes multiple times. PMID:25641657

The activation of human endogenous retrovirus K (HERV-K) is implicated in melanoma cell malignant transformation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Serafino, A.; Balestrieri, E.; Pierimarchi, P.

2009-03-10

Melanoma development is a multi-step process arising from a series of genetic and epigenetic events. Although the sequential stages involved in progression from melanocytes to malignant melanoma are clearly defined, our current understanding of the mechanisms leading to melanoma onset is still incomplete. Growing evidence show that the activation of endogenous retroviral sequences might be involved in transformation of melanocytes as well as in the increased ability of melanoma cells to escape immune surveillance. Here we show that human melanoma cells in vitro undergo a transition from adherent to a more malignant, non-adherent phenotype when exposed to stress conditions. Melanoma-derivedmore » non-adherent cells are characterized by an increased proliferative potential and a decreased expression of both HLA class I molecules and Melan-A/MART-1 antigen, similarly to highly malignant cells. These phenotypic and functional modifications are accompanied by the activation of human endogenous retrovirus K expression (HERV-K) and massive production of viral-like particles. Down-regulation of HERV-K expression by RNA interference prevents the transition from the adherent to the non-adherent growth phenotype in low serum. These results implicate HERV-K in at least some critical steps of melanoma progression.« less

Murine Stem Cell-Based Retrovirus Production for Marking Primary Mouse Mammary Cells for Metastasis Studies.

PubMed

Beverly, Levi J; Podsypanina, Katrina

2016-02-01

Since the introduction of retroviral vector technology, permanent genetic marking of cells has considerably contributed to the understanding of different physiological and disease processes in vivo. Recent marking strategies aim to elucidate the contribution of cells on the clonal level, and the advent of fluorescent proteins has opened new avenues for the in vivo analysis of gene-marked cells. Gene-modified cells are easily identifiable (e.g., via the introduced fluorescent protein) within whole organ structures, allowing one to measure the contribution of transduced cells to malignant outgrowth. In our laboratory, we use the tetracycline-inducible system to study oncogene cooperation in metastatic progression. We use bicistronic retroviruses expressing the tetracycline transactivator (tTA) and the candidate gene (MIT-gene) or the tTA alone (MIT-Rx) to infect primary mammary cells from mice harboring tetracycline-inducible transgenes. This allows for constitutive expression of the candidate gene and tTA-dependent expression of the inducible oncogene. We also use MIG-based vectors, which allow for constitutive expression of the candidate gene and a green fluorescent protein. Here we describe how to produce retroviral particles carrying both MIT- and MIG-based vectors. Because of the fragility of the retroviral envelope, we do not attempt to concentrate the virus, and we directly use packaging cell media to infect primary epithelial cells (either normal or tumor). Infected cells can be transplanted into recipient mice to investigate metastatic colonization. © 2016 Cold Spring Harbor Laboratory Press.

Noninfectious virus-like particles produced by Moloney murine leukemia virus-based retrovirus packaging cells deficient in viral envelope become infectious in the presence of lipofection reagents

PubMed Central

Sharma, Sanjai; Murai, Fukashi; Miyanohara, Atsushi; Friedmann, Theodore

1997-01-01

Retrovirus packaging cell lines expressing the Moloney murine leukemia virus gag and pol genes but lacking virus envelope genes produce virus-like particles constitutively, whether or not they express a transcript from an integrated retroviral provirus. In the absence of a proviral transcript, the assembled particles contain processed gag and reverse transcriptase, and particles made by cells expressing an integrated lacZ provirus also contain viral RNA. The virus-like particles from both cell types are enveloped and are secreted/budded into the extracellular space but are noninfectious. Their physicochemical properties are similar to those of mature retroviral particles. The noninfectious gag pol RNA particles can readily be made infectious by the addition of lipofection reagents to produce preparations with titers of up to 105 colony-forming units per ml. PMID:9380714

Human endogenous retroviruses: nature, occurrence, and clinical implications in human disease.

PubMed Central

Urnovitz, H B; Murphy, W H

1996-01-01

Retroviral diagnostics have become standard in human laboratory medicine. While current emphasis is placed on the human exogenous viruses (human immunodeficiency virus and human T-cell leukemia virus), evidence implicating human endogenous retroviruses (HERVs) in various human disease entities continues to mount. Literature on the occurrence of HERVs in human tissues and cells was analyzed. Substantial evidence documents that retrovirus particles were clearly demonstrable in various tissues and cells in both health and disease and were abundant in the placenta and that their occurrence could be implicated in some of the reproductive diseases. The characteristics of HERVs are summarized, mechanisms of replication and regulation are outlined, and the consistent hormonal responsiveness of HERVs is noted. Clear evidence implicating HERV gene products as participants in glomerulonephritis in some cases of systemic lupus erythematosus is adduced. Data implicating HERVs as etiologic factors in reproductive diseases, in some of the autoimmune diseases, in some forms of rheumatoid arthritis and connective tissue disease, in psoriasis, and in some of the inflammatory neurologic diseases are reviewed. The current major needs are to improve methods for HERV detection, to identify the most appropriate HERV prototypes, and to develop diagnostic reagents so that the putative biologic and pathologic roles of HERVs can be better evaluated. PMID:8665478

Characterization of resistance to rhabdovirus and retrovirus infection in a human myeloid cell line.

PubMed

Boso, Guney; Somia, Nikunj V

2015-01-01

Viruses interact with various permissive and restrictive factors in host cells throughout their replication cycle. Cell lines that are non-permissive to viral infection have been particularly useful in discovering host cell proteins involved in viral life cycles. Here we describe the characterization of a human myeloid leukemia cell line, KG-1, that is resistant to infection by retroviruses and a Rhabdovirus. We show that KG-1 cells are resistant to infection by Vesicular Stomatits Virus as well as VSV Glycoprotein (VSVG) pseudotyped retroviruses due to a defect in binding. Moreover our results indicate that entry by xenotropic retroviral envelope glycoprotein RD114 is impaired in KG-1 cells. Finally we characterize a post- entry block in the early phase of the retroviral life cycle in KG-1 cells that renders the cell line refractory to infection. This cell line will have utility in discovering proteins involved in infection by VSV and HIV-1.

Hybridization capture reveals evolution and conservation across the entire Koala retrovirus genome.

PubMed

Tsangaras, Kyriakos; Siracusa, Matthew C; Nikolaidis, Nikolas; Ishida, Yasuko; Cui, Pin; Vielgrader, Hanna; Helgen, Kristofer M; Roca, Alfred L; Greenwood, Alex D

2014-01-01

The koala retrovirus (KoRV) is the only retrovirus known to be in the midst of invading the germ line of its host species. Hybridization capture and next generation sequencing were used on modern and museum DNA samples of koala (Phascolarctos cinereus) to examine ca. 130 years of evolution across the full KoRV genome. Overall, the entire proviral genome appeared to be conserved across time in sequence, protein structure and transcriptional binding sites. A total of 138 polymorphisms were detected, of which 72 were found in more than one individual. At every polymorphic site in the museum koalas, one of the character states matched that of modern KoRV. Among non-synonymous polymorphisms, radical substitutions involving large physiochemical differences between amino acids were elevated in env, potentially reflecting anti-viral immune pressure or avoidance of receptor interference. Polymorphisms were not detected within two functional regions believed to affect infectivity. Host sequences flanking proviral integration sites were also captured; with few proviral loci shared among koalas. Recently described variants of KoRV, designated KoRV-B and KoRV-J, were not detected in museum samples, suggesting that these variants may be of recent origin.

Hybridization Capture Reveals Evolution and Conservation across the Entire Koala Retrovirus Genome

PubMed Central

Ishida, Yasuko; Cui, Pin; Vielgrader, Hanna; Helgen, Kristofer M.; Roca, Alfred L.; Greenwood, Alex D.

2014-01-01

The koala retrovirus (KoRV) is the only retrovirus known to be in the midst of invading the germ line of its host species. Hybridization capture and next generation sequencing were used on modern and museum DNA samples of koala (Phascolarctos cinereus) to examine ca. 130 years of evolution across the full KoRV genome. Overall, the entire proviral genome appeared to be conserved across time in sequence, protein structure and transcriptional binding sites. A total of 138 polymorphisms were detected, of which 72 were found in more than one individual. At every polymorphic site in the museum koalas, one of the character states matched that of modern KoRV. Among non-synonymous polymorphisms, radical substitutions involving large physiochemical differences between amino acids were elevated in env, potentially reflecting anti-viral immune pressure or avoidance of receptor interference. Polymorphisms were not detected within two functional regions believed to affect infectivity. Host sequences flanking proviral integration sites were also captured; with few proviral loci shared among koalas. Recently described variants of KoRV, designated KoRV-B and KoRV-J, were not detected in museum samples, suggesting that these variants may be of recent origin. PMID:24752422

Intact EAV-HP Endogenous Retrovirus in Sonnerat's Jungle Fowl

PubMed Central

Sacco, M. A.; Howes, K.; Venugopal, K.

2001-01-01

The EAV-HP group of chicken endogenous retrovirus elements was previously shown to be defective, with large deletions of the pol gene. In this report, we demonstrate that genomes of other Gallus species also maintain EAV-HP elements with similar deletions. The chicken EAV-HP1 locus was detected in both red (Gallus gallus gallus) and Sonnerat's (Gallus sonneratii) jungle fowl with identical integration sites, indicating that these elements had integrated before separation of the Gallus species. Furthermore, we demonstrate for the first time that the G. sonneratii genome carries EAV-HP elements with intact pol regions. PMID:11160706

On the general theory of the origins of retroviruses

PubMed Central

2010-01-01

Background The order retroviridae comprises viruses based on ribonucleic acids (RNA). Some, such as HIV and HTLV, are human pathogens. Newly emerged human retroviruses have zoonotic origins. As far as has been established, both repeated infections (themselves possibly responsible for the evolution of viral mutations (Vm) and host adaptability (Ha)); along with interplay between inhibitors and promoters of cell tropism, are needed to effect retroviral cross-species transmissions. However, the exact modus operadi of intertwine between these factors at molecular level remains to be established. Knowledge of such intertwine could lead to a better understanding of retrovirology and possibly other infectious processes. This study was conducted to derive the mathematical equation of a general theory of the origins of retroviruses. Methods and results On the basis of an arbitrarily non-Euclidian geometrical "thought experiment" involving the cross-species transmission of simian foamy virus (sfv) from a non-primate species Xy to Homo sapiens (Hs), initially excluding all social factors, the following was derived. At the port of exit from Xy (where the species barrier, SB, is defined by the Index of Origin, IO), sfv shedding is (1) enhanced by two transmitting tensors (Tt), (i) virus-specific immunity (VSI) and (ii) evolutionary defenses such as APOBEC, RNA interference pathways, and (when present) expedited therapeutics (denoted e2D); and (2) opposed by the five accepting scalars (At): (a) genomic integration hot spots, gIHS, (b) nuclear envelope transit (NMt) vectors, (c) virus-specific cellular biochemistry, VSCB, (d) virus-specific cellular receptor repertoire, VSCR, and (e) pH-mediated cell membrane transit, (↓pH CMat). Assuming As and Tt to be independent variables, IO = Tt/As. The same forces acting in an opposing manner determine SB at the port of sfv entry (defined here by the Index of Entry, IE = As/Tt). Overall, If sfv encounters no unforeseen effects on transit

A Simple Model for Immature Retrovirus Capsid Assembly

NASA Astrophysics Data System (ADS)

Paquay, Stefan; van der Schoot, Paul; Dragnea, Bogdan

In this talk I will present simulations of a simple model for capsomeres in immature virus capsids, consisting of only point particles with a tunable range of attraction constrained to a spherical surface. We find that, at sufficiently low density, a short interaction range is sufficient for the suppression of five-fold defects in the packing and causes instead larger tears and scars in the capsid. These findings agree both qualitatively and quantitatively with experiments on immature retrovirus capsids, implying that the structure of the retroviral protein lattice can, for a large part, be explained simply by the effective interaction between the capsomeres. We thank the HFSP for funding under Grant RGP0017/2012.

Expression of Rous sarcoma virus-derived retroviral vectors in the avian blastoderm: Potential as stable genetic markers

DOE Office of Scientific and Technical Information (OSTI.GOV)

Reddy, S.T.; Stoker, A.W.; Bissell, M.J.

1991-12-01

Retroviruses are valuable tools in studies of embryonic development, both as gene expression vectors and as cell lineage markers. In this study early chicken blastoderm cells are shown to be permissive for infection by Rous sarcoma virus and derivative replication-defective by Rous sarcoma virus and derivative replication-defective vectors, and, in contrast to previously published data, these cells will readily express viral genes. In cultured blastoderm cells, Rous sarcoma virus stably integrates and is transcribed efficiently, producing infectious virus particles. Using replication-defective vectors encoding the bacterial lacZ gene, the authors further show that blastoderms can be infected in culture and inmore » ovo. In ovo, lacZ expression is seen within 24 hours of virus inoculation, and by 96 hours stably expressing clones of cells are observed in diverse tissues throughout the embryo, including epidermis, somites, and heart, as well as in extraembryonic membranes. Given the rapid onset of vector expression and the broad range of permissive cell types, it should be feasible to use Rous sarcoma virus-derived retroviruses as early lineage markers and expression vectors beginning at the blastoderm stage of avian embryogenesis.« less

Genetic Engineering of T Cells to Target HERV-K, an Ancient Retrovirus on Melanoma.

PubMed

Krishnamurthy, Janani; Rabinovich, Brian A; Mi, Tiejuan; Switzer, Kirsten C; Olivares, Simon; Maiti, Sourindra N; Plummer, Joshua B; Singh, Harjeet; Kumaresan, Pappanaicken R; Huls, Helen M; Wang-Johanning, Feng; Cooper, Laurence J N

2015-07-15

The human endogenous retrovirus (HERV-K) envelope (env) protein is a tumor-associated antigen (TAA) expressed on melanoma but not normal cells. This study was designed to engineer a chimeric antigen receptor (CAR) on T-cell surface, such that they target tumors in advanced stages of melanoma. Expression of HERV-K protein was analyzed in 220 melanoma samples (with various stages of disease) and 139 normal organ donor tissues using immunohistochemical (IHC) analysis. HERV-K env-specific CAR derived from mouse monoclonal antibody was introduced into T cells using the transposon-based Sleeping Beauty (SB) system. HERV-K env-specific CAR(+) T cells were expanded ex vivo on activating and propagating cells (AaPC) and characterized for CAR expression and specificity. This includes evaluating the HERV-K-specific CAR(+) T cells for their ability to kill A375-SM metastasized tumors in a mouse xenograft model. We detected HERV-K env protein on melanoma but not in normal tissues. After electroporation of T cells and selection on HERV-K(+) AaPC, more than 95% of genetically modified T cells expressed the CAR with an effector memory phenotype and lysed HERV-K env(+) tumor targets in an antigen-specific manner. Even though there is apparent shedding of this TAA from tumor cells that can be recognized by HERV-K env-specific CAR(+) T cells, we observed a significant antitumor effect. Adoptive cellular immunotherapy with HERV-K env-specific CAR(+) T cells represents a clinically appealing treatment strategy for advanced-stage melanoma and provides an approach for targeting this TAA on other solid tumors. ©2015 American Association for Cancer Research.

Small finger protein of avian and murine retroviruses has nucleic acid annealing activity and positions the replication primer tRNA onto genomic RNA.

PubMed Central

Prats, A C; Sarih, L; Gabus, C; Litvak, S; Keith, G; Darlix, J L

1988-01-01

Retrovirus virions carry a diploid genome associated with a large number of small viral finger protein molecules which are required for encapsidation. Our present results show that finger protein p12 of Rous sarcoma virus (RSV) and p10 of murine leukaemia virus (MuLV) positions replication primer tRNA on the replication initiation site (PBS) at the 5' end of the RNA genome. An RSV mutant with a Val-Pro insertion in the finger motif of p12 is able to partially encapsidate genomic RNA but is not infectious because mutated p12 is incapable of positioning the replication primer, tRNATrp. Since all known replication competent retroviruses, and the plant virus CaMV, code for finger proteins analogous to RSV p12 or MuLV p10, the initial stage of reverse transcription in avian, mammalian and human retroviruses and in CaMV is probably controlled in an analogous way. Images PMID:2458920

Small finger protein of avian and murine retroviruses has nucleic acid annealing activity and positions the replication primer tRNA onto genomic RNA.

PubMed

Prats, A C; Sarih, L; Gabus, C; Litvak, S; Keith, G; Darlix, J L

1988-06-01

Retrovirus virions carry a diploid genome associated with a large number of small viral finger protein molecules which are required for encapsidation. Our present results show that finger protein p12 of Rous sarcoma virus (RSV) and p10 of murine leukaemia virus (MuLV) positions replication primer tRNA on the replication initiation site (PBS) at the 5' end of the RNA genome. An RSV mutant with a Val-Pro insertion in the finger motif of p12 is able to partially encapsidate genomic RNA but is not infectious because mutated p12 is incapable of positioning the replication primer, tRNATrp. Since all known replication competent retroviruses, and the plant virus CaMV, code for finger proteins analogous to RSV p12 or MuLV p10, the initial stage of reverse transcription in avian, mammalian and human retroviruses and in CaMV is probably controlled in an analogous way.

Seroprevalence of xenotropic murine leukemia virus-related virus in normal and retrovirus-infected blood donors.

PubMed

Qiu, Xiaoxing; Swanson, Priscilla; Tang, Ning; Leckie, Gregor W; Devare, Sushil G; Schochetman, Gerald; Hackett, John

2012-02-01

Xenotropic murine leukemia virus-related virus (XMRV) has been reported in patients with prostate cancer and chronic fatigue syndrome. Although results have been conflicting, the potential of XMRV as an infectious human retrovirus has raised concerns about transfusion safety. To address this issue, normal and retrovirus-infected blood donors were screened for evidence of XMRV infection. Plasma from 1000 US, 100 human immunodeficiency virus Type 1-infected Cameroonian, and 642 human T-lymphotropic virus Type I (HTLV-I)-infected or uninfected Japanese blood donors as well as 311 sexually transmitted disease diagnostic specimens were screened for antibodies to XMRV gp70 and p15E using chemiluminescent immunoassays (CMIAs). CMIA-reactive samples were evaluated by p30 CMIA, Western blot, and real-time reverse transcriptase polymerase chain reaction. XMRV seroreactivity was low (0%-0.6%) with the exception of the HTLV-I-infected donors (4.9%). Antibody was detected against only a single XMRV protein (p15E or gp70); none of the seroreactive samples had detectable XMRV pol or env sequences. The elevated seroreactivity in HTLV-I-infected donors was due to an increased p15E seroreactive rate (4.1%). Inspection of XMRV and HTLV sequences revealed a high level of conservation within the immunodominant region (IDR) of the transmembrane protein. In some cases, HTLV IDR peptide competitively reduced the XMRV p15E signal. Based on the low prevalence of seroreactivity, detection of antibody to only a single XMRV protein and the absence of XMRV sequences, this study finds no compelling evidence of XMRV in normal or retrovirus-infected blood donors. The increased p15E seroreactivity observed in HTLV infection is likely due to cross-reactive antibodies. © 2012 American Association of Blood Banks.

Upregulation of Human Endogenous Retrovirus-K Is Linked to Immunity and Inflammation in Pulmonary Arterial Hypertension.

PubMed

Saito, Toshie; Miyagawa, Kazuya; Chen, Shih-Yu; Tamosiuniene, Rasa; Wang, Lingli; Sharpe, Orr; Samayoa, Erik; Harada, Daisuke; Moonen, Jan-Renier A J; Cao, Aiqin; Chen, Pin-I; Hennigs, Jan K; Gu, Mingxia; Li, Caiyun G; Leib, Ryan D; Li, Dan; Adams, Christopher M; Del Rosario, Patricia A; Bill, Matthew; Haddad, Francois; Montoya, Jose G; Robinson, William H; Fantl, Wendy J; Nolan, Garry P; Zamanian, Roham T; Nicolls, Mark R; Chiu, Charles Y; Ariza, Maria E; Rabinovitch, Marlene

2017-11-14

Immune dysregulation has been linked to occlusive vascular remodeling in pulmonary arterial hypertension (PAH) that is hereditary, idiopathic, or associated with other conditions. Circulating autoantibodies, lung perivascular lymphoid tissue, and elevated cytokines have been related to PAH pathogenesis but without a clear understanding of how these abnormalities are initiated, perpetuated, and connected in the progression of disease. We therefore set out to identify specific target antigens in PAH lung immune complexes as a starting point toward resolving these issues to better inform future application of immunomodulatory therapies. Lung immune complexes were isolated and PAH target antigens were identified by liquid chromatography tandem mass spectrometry, confirmed by enzyme-linked immunosorbent assay, and localized by confocal microscopy. One PAH antigen linked to immunity and inflammation was pursued and a link to PAH pathophysiology was investigated by next-generation sequencing, functional studies in cultured monocytes and endothelial cells, and hemodynamic and lung studies in a rat. SAM domain and HD domain-containing protein 1 (SAMHD1), an innate immune factor that suppresses HIV replication, was identified and confirmed as highly expressed in immune complexes from 16 hereditary and idiopathic PAH versus 12 control lungs. Elevated SAMHD1 was localized to endothelial cells, perivascular dendritic cells, and macrophages, and SAMHD1 antibodies were prevalent in tertiary lymphoid tissue. An unbiased screen using metagenomic sequencing related SAMHD1 to increased expression of human endogenous retrovirus K (HERV-K) in PAH versus control lungs (n=4). HERV-K envelope and deoxyuridine triphosphate nucleotidohydrolase mRNAs were elevated in PAH versus control lungs (n=10), and proteins were localized to macrophages. HERV-K deoxyuridine triphosphate nucleotidohydrolase induced SAMHD1 and proinflammatory cytokines (eg, interleukin 6, interleukin 1β, and tumor

Conference Highlights of the 16th International Conference on Human Retrovirology: HTLV and Related Retroviruses, 26–30 June 2013, Montreal, Canada

PubMed Central

2014-01-01

The 16th International Conference on Human Retrovirology: HTLV and Related Retroviruses was held in Montreal, Québec from June 26th to June 30th, 2013 and was therefore hosted by a Canadian city for the first time. The major topic of the meeting was human T-lymphotropic viruses (HTLVs) and was covered through distinct oral and poster presentation sessions: clinical research, animal models, immunology, molecular and cellular biology, human endogenous and emerging exogenous retroviruses and virology. In this review, highlights of the meeting are provided by different experts for each of these research areas. PMID:24558960

Utility of next-generation RNA-sequencing in identifying chimeric transcription involving human endogenous retroviruses.

PubMed

Sokol, Martin; Jessen, Karen Margrethe; Pedersen, Finn Skou

2016-01-01

Several studies have shown that human endogenous retroviruses and endogenous retrovirus-like repeats (here collectively HERVs) impose direct regulation on human genes through enhancer and promoter motifs present in their long terminal repeats (LTRs). Although chimeric transcription in which novel gene isoforms containing retroviral and human sequence are transcribed from viral promoters are commonly associated with disease, regulation by HERVs is beneficial in other settings; for example, in human testis chimeric isoforms of TP63 induced by an ERV9 LTR protect the male germ line upon DNA damage by inducing apoptosis, whereas in the human globin locus the γ- and β-globin switch during normal hematopoiesis is mediated by complex interactions of an ERV9 LTR and surrounding human sequence. The advent of deep sequencing or next-generation sequencing (NGS) has revolutionized the way researchers solve important scientific questions and develop novel hypotheses in relation to human genome regulation. We recently applied next-generation paired-end RNA-sequencing (RNA-seq) together with chromatin immunoprecipitation with sequencing (ChIP-seq) to examine ERV9 chimeric transcription in human reference cell lines from Encyclopedia of DNA Elements (ENCODE). This led to the discovery of advanced regulation mechanisms by ERV9s and other HERVs across numerous human loci including transcription of large gene-unannotated genomic regions, as well as cooperative regulation by multiple HERVs and non-LTR repeats such as Alu elements. In this article, well-established examples of human gene regulation by HERVs are reviewed followed by a description of paired-end RNA-seq, and its application in identifying chimeric transcription genome-widely. Based on integrative analyses of RNA-seq and ChIP-seq, data we then present novel examples of regulation by ERV9s of tumor suppressor genes CADM2 and SEMA3A, as well as transcription of an unannotated region. Taken together, this article highlights

Specific in vivo labeling with GFP retroviruses, lentiviruses, and adenoviruses for imaging

NASA Astrophysics Data System (ADS)

Hoffman, Robert M.; Kishimoto, Hiroyuki; Fujiwara, Toshiyoshi

2008-02-01

Fluorescent proteins have revolutionized the field of imaging. Our laboratory pioneered in vivo imaging with fluorescent proteins. Fluorescent proteins have enabled imaging at the subcellular level in mice. We review here the use of different vectors carrying fluorescent proteins to selectively label normal and tumor tissue in vivo. We show that a GFP retrovirus and telomerase-driven GFP adenovirus can selectively label tumors in mice. We also show that a GFP lentivirus can selectively label the liver in mice. The practical application of these results are discussed.

Reticuloendotheliosis virus: detection of immunological relationship to mammalian type C retroviruses.

PubMed Central

Charman, H P; Gilden, R V; Oroszlan, S

1979-01-01

Reticuloendotheliosis virus (REV) p30 shares cross-reactive determinants and a common NH2-terminal tripeptide with mammalian type C viral p30's. An interspecies competition radioimmunoassay was developed, using iodinated REV p30 and a broadly reactive antiserum to mammalian virus p30's. The avian leukosis-sarcoma viruses and mammalian non-type C retroviruses did not compete in this assay. Previous data indicating that the REV group is not represented completely in normal avian cell DNA lead us to speculate that this may be the first example of interclass transmission, albeit in the remote past, among the Retroviridae. PMID:87519

Tracking interspecies transmission and long-term evolution of an ancient retrovirus using the genomes of modern mammals

PubMed Central

Diehl, William E; Patel, Nirali; Halm, Kate; Johnson, Welkin E

2016-01-01

Mammalian genomes typically contain hundreds of thousands of endogenous retroviruses (ERVs), derived from ancient retroviral infections. Using this molecular 'fossil' record, we reconstructed the natural history of a specific retrovirus lineage (ERV-Fc) that disseminated widely between ~33 and ~15 million years ago, corresponding to the Oligocene and early Miocene epochs. Intercontinental viral spread, numerous instances of interspecies transmission and emergence in hosts representing at least 11 mammalian orders, and a significant role for recombination in diversification of this viral lineage were also revealed. By reconstructing the canonical retroviral genes, we identified patterns of adaptation consistent with selection to maintain essential viral protein functions. Our results demonstrate the unique potential of the ERV fossil record for studying the processes of viral spread and emergence as they play out across macro-evolutionary timescales, such that looking back in time may prove insightful for predicting the long-term consequences of newly emerging viral infections. DOI: http://dx.doi.org/10.7554/eLife.12704.001 PMID:26952212

Tracking interspecies transmission and long-term evolution of an ancient retrovirus using the genomes of modern mammals.

PubMed

Diehl, William E; Patel, Nirali; Halm, Kate; Johnson, Welkin E

2016-03-08

Mammalian genomes typically contain hundreds of thousands of endogenous retroviruses (ERVs), derived from ancient retroviral infections. Using this molecular 'fossil' record, we reconstructed the natural history of a specific retrovirus lineage (ERV-Fc) that disseminated widely between ~33 and ~15 million years ago, corresponding to the Oligocene and early Miocene epochs. Intercontinental viral spread, numerous instances of interspecies transmission and emergence in hosts representing at least 11 mammalian orders, and a significant role for recombination in diversification of this viral lineage were also revealed. By reconstructing the canonical retroviral genes, we identified patterns of adaptation consistent with selection to maintain essential viral protein functions. Our results demonstrate the unique potential of the ERV fossil record for studying the processes of viral spread and emergence as they play out across macro-evolutionary timescales, such that looking back in time may prove insightful for predicting the long-term consequences of newly emerging viral infections.

Characterization of a nucleocapsid-like region and of two distinct primer tRNALys,2 binding sites in the endogenous retrovirus Gypsy.

PubMed

Gabus, Caroline; Ivanyi-Nagy, Roland; Depollier, Julien; Bucheton, Alain; Pelisson, Alain; Darlix, Jean-Luc

2006-01-01

Mobile LTR-retroelements comprising retroviruses and LTR-retrotransposons form a large part of eukaryotic genomes. Their mode of replication and abundance favour the notion that they are major actors in eukaryote evolution. The Gypsy retroelement can spread in the germ line of the fruit fly Drosophila melanogaster via both env-independent and env-dependent processes. Thus, Gypsy is both an active retrotransposon and an infectious retrovirus resembling the gammaretrovirus MuLV. However, unlike gammaretroviruses, the Gypsy Gag structural precursor is not processed into Matrix, Capsid and Nucleocapsid (NC) proteins. In contrast, it has features in common with Gag of the ancient yeast TY1 retroelement. These characteristics of Gypsy make it a very interesting model to study replication of a retroelement at the frontier between ancient retrotransposons and retroviruses. We investigated Gypsy replication using an in vitro model system and transfection of insect cells. Results show that an unstructured domain of Gypsy Gag has all the properties of a retroviral NC. This NC-like peptide forms ribonucleoparticle-like complexes upon binding Gypsy RNA and directs the annealing of primer tRNA(Lys,2) to two distinct primer binding sites (PBS) at the genome 5' and 3' ends. Only the 5' PBS is indispensable for cDNA synthesis in vitro and in Drosophila cells.

Sequence variation of koala retrovirus transmembrane protein p15E among koalas from different geographic regions.

PubMed

Ishida, Yasuko; McCallister, Chelsea; Nikolaidis, Nikolas; Tsangaras, Kyriakos; Helgen, Kristofer M; Greenwood, Alex D; Roca, Alfred L

2015-01-15

The koala retrovirus (KoRV), which is transitioning from an exogenous to an endogenous form, has been associated with high mortality in koalas. For other retroviruses, the envelope protein p15E has been considered a candidate for vaccine development. We therefore examined proviral sequence variation of KoRV p15E in a captive Queensland and three wild southern Australian koalas. We generated 163 sequences with intact open reading frames, which grouped into 39 distinct haplotypes. Sixteen distinct haplotypes comprising 139 of the sequences (85%) coded for the same polypeptide. Among the remaining 23 haplotypes, 22 were detected only once among the sequences, and each had 1 or 2 non-synonymous differences from the majority sequence. Several analyses suggested that p15E was under purifying selection. Important epitopes and domains were highly conserved across the p15E sequences and in previously reported exogenous KoRVs. Overall, these results support the potential use of p15E for KoRV vaccine development. Copyright © 2014 Elsevier Inc. All rights reserved.

Multiple sclerosis-associated retrovirus, Epstein-Barr virus, and vitamin D status in patients with relapsing remitting multiple sclerosis.

PubMed

Mostafa, Aliehossadat; Jalilvand, Somayeh; Shoja, Zabihollah; Nejati, Ahmad; Shahmahmoodi, Shohreh; Sahraian, Mohammad Ali; Marashi, Sayed Mahdi

2017-07-01

The relationship between infections and autoimmune diseases is complex and there are several reports highlighting the role of human endogenous retroviruses (HERVs) in these patients. The levels of multiple sclerosis-associated retrovirus (MSRV)-type DNA of Env gene was measured in peripheral blood mononuclear cells from 52 patients with relapsing-remitting multiple sclerosis (RRMS) and 40 healthy controls using specific quantitative PCR (qPCR) analysis. Furthermore, we analyzed the status of HERV-W/MSRV in these patients with regards to both EBV (DNA load and anti-EBNA1 IgG antibody) and vitamin D concentration. MSRV DNA copy number were significantly higher in RRMS patients than healthy controls (P < 0.0001). Interestingly, an inverse correlation was found between MSRV DNA copy number and serum vitamin D concentration (P < 0.01), but not for EBV load or anti-EBNA-1 IgG antibody. © 2017 Wiley Periodicals, Inc.

Orchestrating the Selection and Packaging of Genomic RNA by Retroviruses: An Ensemble of Viral and Host Factors

PubMed Central

Kaddis Maldonado, Rebecca J.; Parent, Leslie J.

2016-01-01

Infectious retrovirus particles contain two copies of unspliced viral RNA that serve as the viral genome. Unspliced retroviral RNA is transcribed in the nucleus by the host RNA polymerase II and has three potential fates: (1) it can be spliced into subgenomic messenger RNAs (mRNAs) for the translation of viral proteins; or it can remain unspliced to serve as either (2) the mRNA for the translation of Gag and Gag–Pol; or (3) the genomic RNA (gRNA) that is packaged into virions. The Gag structural protein recognizes and binds the unspliced viral RNA to select it as a genome, which is selected in preference to spliced viral RNAs and cellular RNAs. In this review, we summarize the current state of understanding about how retroviral packaging is orchestrated within the cell and explore potential new mechanisms based on recent discoveries in the field. We discuss the cis-acting elements in the unspliced viral RNA and the properties of the Gag protein that are required for their interaction. In addition, we discuss the role of host factors in influencing the fate of the newly transcribed viral RNA, current models for how retroviruses distinguish unspliced viral mRNA from viral genomic RNA, and the possible subcellular sites of genomic RNA dimerization and selection by Gag. Although this review centers primarily on the wealth of data available for the alpharetrovirus Rous sarcoma virus, in which a discrete RNA packaging sequence has been identified, we have also summarized the cis- and trans-acting factors as well as the mechanisms governing gRNA packaging of other retroviruses for comparison. PMID:27657110

Profound Amplification of Pathogenic Murine Polytropic Retrovirus Release from Coinfected Cells

PubMed Central

Rosenke, Kyle; Lavignon, Marc; Malik, Frank; Kolokithas, Angelo; Hendrick, Duncan; Virtaneva, Kimmo; Peterson, Karin

2012-01-01

Previous studies indicate that mice infected with mixtures of mouse retroviruses (murine leukemia viruses [MuLVs]) exhibit dramatically altered pathology compared to mice infected with individual viruses of the mixture. Coinoculation of the ecotropic virus Friend MuLV (F-MuLV) with Fr98, a polytropic MuLV, induced a rapidly fatal neurological disease that was not observed in infections with either virus alone. The polytropic virus load in coinoculated mice was markedly enhanced, while the ecotropic F-MuLV load was unchanged. Furthermore, pseudotyping of the polytropic MuLV genome within ecotropic virions was nearly complete in coinoculated mice. In an effort to better understand these phenomena, we examined mixed retrovirus infections by utilizing in vitro cell lines. Similar to in vivo mixed infections, the polytropic MuLV genome was extensively pseudotyped within ecotropic virions; polytropic virus release was profoundly elevated in coinfected cells, and the ecotropic virus release was unchanged. A reduced level of polytropic SU protein on the surfaces of coinfected cells was observed and correlated with a reduced level of nonpseudotyped polytropic virion release. Marked amplification and pseudotyping of the polytropic MuLV were also observed in mixed Fr98–F-MuLV infections of cell lines derived from the central nervous system (CNS), the target for Fr98 pathogenesis. Additional experiments indicated that pseudotyping contributed to the elevated polytropic virus titer by increasing the efficiency of packaging and release of the polytropic genomes within ecotropic virions. Mixed infections are the rule rather than the exception in retroviral infection, and the ability to examine them in vitro should facilitate a more thorough understanding of retroviral interactions in general. PMID:22514353

Vaccination directed against the human endogenous retrovirus-K envelope protein inhibits tumor growth in a murine model system.

PubMed

Kraus, Benjamin; Fischer, Katrin; Büchner, Sarah M; Wels, Winfried S; Löwer, Roswitha; Sliva, Katja; Schnierle, Barbara S

2013-01-01

Human endogenous retrovirus (HERV) genomes are chromosomally integrated in all cells of an individual. They are normally transcriptionally silenced and transmitted only vertically. Enhanced expression of HERV-K accompanied by the emergence of anti-HERV-K-directed immune responses has been observed in tumor patients and HIV-infected individuals. As HERV-K is usually not expressed and immunological tolerance development is unlikely, it is an appropriate target for the development of immunotherapies. We generated a recombinant vaccinia virus (MVA-HKenv) expressing the HERV-K envelope glycoprotein (ENV), based on the modified vaccinia virus Ankara (MVA), and established an animal model to test its vaccination efficacy. Murine renal carcinoma cells (Renca) were genetically altered to express E. coli beta-galactosidase (RLZ cells) or the HERV-K ENV gene (RLZ-HKenv cells). Intravenous injection of RLZ-HKenv cells into syngenic BALB/c mice led to the formation of pulmonary metastases, which were detectable by X-gal staining. A single vaccination of tumor-bearing mice with MVA-HKenv drastically reduced the number of pulmonary RLZ-HKenv tumor nodules compared to vaccination with wild-type MVA. Prophylactic vaccination of mice with MVA-HKenv precluded the formation of RLZ-HKenv tumor nodules, whereas wild-type MVA-vaccinated animals succumbed to metastasis. Protection from tumor formation correlated with enhanced HERV-K ENV-specific killing activity of splenocytes. These data demonstrate for the first time that HERV-K ENV is a useful target for vaccine development and might offer new treatment opportunities for diverse types of cancer.

RNAi and retroviruses: are they in RISC?

PubMed

Vasselon, Thierry; Bouttier, Manuella; Saumet, Anne; Lecellier, Charles-Henri

2013-02-01

RNA interference (RNAi) is a potent cellular system against viruses in various organisms. Although common traits are observed in plants, insects, and nematodes, the situation observed in mammals appears more complex. In mammalian somatic cells, RNAi is implicated in endonucleolytic cleavage mediated by artificially delivered small interfering RNAs (siRNAs) as well as in translation repression mediated by microRNAs (miRNAs). Because siRNAs and miRNAs recognize viral mRNAs, RNAi inherently limits virus production and participates in antiviral defense. However, several observations made in the cases of hepatitis C virus and retroviruses (including the human immunodeficiency virus and the primate foamy virus) bring evidence that this relationship is much more complex and that certain components of the RNAi effector complex [called the RNA-induced silencing complex (RISC)], such as AGO2, are also required for viral replication. Here, we summarize recent discoveries that have revealed this dual implication in virus biology. We further discuss their potential implications for the functions of RNAi-related proteins, with special emphasis on retrotransposition and genome stability.

Identification of an Envelope Protein from the FRD Family of Human Endogenous Retroviruses (HERV-FRD) Conferring Infectivity and Functional Conservation among Simians

PubMed Central

Blaise, Sandra; Ruggieri, Alessia; Dewannieux, Marie; Cosset, François-Loic; Heidmann, Thierry

2004-01-01

A member of the HERV-W family of human endogenous retroviruses (HERV) had previously been demonstrated to encode a functional envelope which can form pseudotypes with human immunodeficiency virus type 1 virions and confer infectivity on the resulting retrovirus particles. Here we show that a second envelope protein sorted out by a systematic search for fusogenic proteins that we made among all the HERV coding envelope genes and belonging to the HERV-FRD family can also make pseudotypes and confer infectivity. We further show that the orthologous envelope genes that were isolated from simians—from New World monkeys to humans—are also functional in the infectivity assay, with one singular exception for the gibbon HERV-FRD gene, which is found to be fusogenic in a cell-cell fusion assay, as observed for the other simian envelopes, but which is not infectious. Sequence comparison of the FRD envelopes revealed a limited number of mutations among simians, and one point mutation—located in the TM subunit—was shown to be responsible for the loss of infectivity of the gibbon envelope. The functional characterization of the identified envelopes is strongly indicative of an ancestral retrovirus infection and endogenization, with some of the envelope functions subsequently retained in evolution. PMID:14694139

Identification of an envelope protein from the FRD family of human endogenous retroviruses (HERV-FRD) conferring infectivity and functional conservation among simians.

PubMed

Blaise, Sandra; Ruggieri, Alessia; Dewannieux, Marie; Cosset, François-Loic; Heidmann, Thierry

2004-01-01

A member of the HERV-W family of human endogenous retroviruses (HERV) had previously been demonstrated to encode a functional envelope which can form pseudotypes with human immunodeficiency virus type 1 virions and confer infectivity on the resulting retrovirus particles. Here we show that a second envelope protein sorted out by a systematic search for fusogenic proteins that we made among all the HERV coding envelope genes and belonging to the HERV-FRD family can also make pseudotypes and confer infectivity. We further show that the orthologous envelope genes that were isolated from simians-from New World monkeys to humans-are also functional in the infectivity assay, with one singular exception for the gibbon HERV-FRD gene, which is found to be fusogenic in a cell-cell fusion assay, as observed for the other simian envelopes, but which is not infectious. Sequence comparison of the FRD envelopes revealed a limited number of mutations among simians, and one point mutation-located in the TM subunit-was shown to be responsible for the loss of infectivity of the gibbon envelope. The functional characterization of the identified envelopes is strongly indicative of an ancestral retrovirus infection and endogenization, with some of the envelope functions subsequently retained in evolution.

Characterization of a nucleocapsid-like region and of two distinct primer tRNALys,2 binding sites in the endogenous retrovirus Gypsy

PubMed Central

Gabus, Caroline; Ivanyi-Nagy, Roland; Depollier, Julien; Bucheton, Alain; Pelisson, Alain; Darlix, Jean-Luc

2006-01-01

Mobile LTR-retroelements comprising retroviruses and LTR-retrotransposons form a large part of eukaryotic genomes. Their mode of replication and abundance favour the notion that they are major actors in eukaryote evolution. The Gypsy retroelement can spread in the germ line of the fruit fly Drosophila melanogaster via both env-independent and env-dependent processes. Thus, Gypsy is both an active retrotransposon and an infectious retrovirus resembling the gammaretrovirus MuLV. However, unlike gammaretroviruses, the Gypsy Gag structural precursor is not processed into Matrix, Capsid and Nucleocapsid (NC) proteins. In contrast, it has features in common with Gag of the ancient yeast TY1 retroelement. These characteristics of Gypsy make it a very interesting model to study replication of a retroelement at the frontier between ancient retrotransposons and retroviruses. We investigated Gypsy replication using an in vitro model system and transfection of insect cells. Results show that an unstructured domain of Gypsy Gag has all the properties of a retroviral NC. This NC-like peptide forms ribonucleoparticle-like complexes upon binding Gypsy RNA and directs the annealing of primer tRNALys,2 to two distinct primer binding sites (PBS) at the genome 5′ and 3′ ends. Only the 5′ PBS is indispensable for cDNA synthesis in vitro and in Drosophila cells. PMID:17040893

Immunotherapy of murine retrovirus-induced acquired immunodeficiency by CD4 T regulatory cell depletion and PD-1 blockade.

PubMed

Li, Wen; Green, William R

2011-12-01

LP-BM5 retrovirus induces a complex disease featuring an acquired immunodeficiency syndrome termed murine AIDS (MAIDS) in susceptible strains of mice, such as C57BL/6 (B6). CD4 T helper effector cells are required for MAIDS induction and progression of viral pathogenesis. CD8 T cells are not needed for viral pathogenesis, but rather, are essential for protection from disease in resistant strains, such as BALB/c. We have discovered an immunodominant cytolytic T lymphocyte (CTL) epitope encoded in a previously unrecognized LP-BM5 retroviral alternative (+1 nucleotide [nt]) gag translational open reading frame. CTLs specific for this cryptic gag epitope are the basis of protection from LP-BM5-induced immunodeficiency in BALB/c mice, and the inability of B6 mice to mount an anti-gag CTL response appears critical to the initiation and progression of LP-BM5-induced MAIDS. However, uninfected B6 mice primed by LP-BM5-induced tumors can generate CTL responses to an LP-BM5 retrovirus infection-associated epitope(s) that is especially prevalent on such MAIDS tumor cells, indicating the potential to mount a protective CD8 T-cell response. Here, we utilized this LP-BM5 retrovirus-induced disease system to test whether modulation of normal immune down-regulatory mechanisms can alter retroviral pathogenesis. Thus, following in vivo depletion of CD4 T regulatory (Treg) cells and/or selective interruption of PD-1 negative signaling in the CD8 T-cell compartment, retroviral pathogenesis was significantly decreased, with the combined treatment of CD4 Treg cell depletion and PD-1 blockade working in a synergistic fashion to substantially reduce the induction of MAIDS.

Chromosomal Distribution of Endogenous Jaagsiekte Sheep Retrovirus Proviral Sequences in the Sheep Genome

PubMed Central

Carlson, Jonathan; Lyon, Monique; Bishop, Jeanette; Vaiman, Anne; Cribiu, Edmond; Mornex, Jean-François; Brown, Susan; Knudson, Dennis; DeMartini, James; Leroux, Caroline

2003-01-01

A family of endogenous retroviruses (enJSRV) closely related to Jaagsiekte sheep retrovirus (JSRV) is ubiquitous in domestic and wild sheep and goats. Southern blot hybridization studies indicate that there is little active replication or movement of the enJSRV proviruses in these species. Two approaches were used to investigate the distribution of proviral loci in the sheep genome. Fluorescence in situ hybridization (FISH) to metaphase chromosome spreads using viral DNA probes was used to detect loci on chromosomes. Hybridization signals were reproducibly detected on seven sheep chromosomes and eight goat chromosomes in seven cell lines. In addition, a panel of 30 sheep-hamster hybrid cell lines, each of which carries one or more sheep chromosomes and which collectively contain the whole sheep genome, was examined for enJSRV sequences. DNA from each of the lines was used as a template for PCR with JSRV gag-specific primers. A PCR product was amplified from 27 of the hybrid lines, indicating that JSRV gag sequences are found on at least 15 of the 28 sheep chromosomes, including those identified by FISH. Thus, enJSRV proviruses are essentially randomly distributed among the chromosomes of sheep and goats. FISH and/or Southern blot hybridization on DNA from several of the sheep-hamster hybrid cell lines suggests that loci containing multiple copies of enJSRV are present on chromosomes 6 and 9. The origin and functional significance of these arrays is not known. PMID:12915578

Transduction of Schistosoma mansoni by vesicular stomatitis virus glycoprotein-pseudotyped Moloney murine leukemia retrovirus.

PubMed

Kines, Kristine J; Mann, Victoria H; Morales, Maria E; Shelby, Bryan D; Kalinna, Bernd H; Gobert, Geoffrey N; Chirgwin, Sharon R; Brindley, Paul J

2006-04-01

Retroviral transduction of cultured schistosomes offers a potential means to establish transgenic lines of schistosomes and thereby to facilitate the elucidation of schistosome gene function and expression. The Moloney murine leukemia retroviral (MMLV) vector pLNHX was modified to incorporate EGFP or luciferase reporter genes under control of schistosome endogenous gene promoters from the spliced leader RNA and HSP70 genes. These constructs and a plasmid encoding vesicular stomatitis virus glycoprotein (VSVG) were utilized along with GP2-293 cells to produce replication incompetent retrovirus particles pseudotyped with the VSVG envelope. Exposure of several developmental stages, including sporocysts, of Schistosoma mansoni to these virions was facilitated by incubation with polybrene and/or by centrifugation. The early stages of binding and uptake of virus to the parasite tegument were demonstrated by the immunofluorescence colocalization of VSVG envelope and retroviral capsid proteins. Southern hybridization analysis indicated the integration of proviral forms of the MMLV constructs in genomic DNA isolated from the virus exposed schistosomes. Furthermore, analysis of RNA isolated from virus treated parasites demonstrated the presence of transcripts encoding reporter transgenes. Together these results indicated productive transduction by VSVG pseudotyped MMLV of cultured schistosomes, and suggest a tractable route forward towards heritable schistosome transgenesis.

Crystallization and preliminary X-ray studies of dUTPase from Mason–Pfizer monkey retrovirus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Barabás, Orsolya; Németh, Veronika; Vértessy, Beáta G., E-mail: vertessy@enzim.hu

2006-04-01

Deoxyuridine 5′-triphosphate nucleotidohydrolase from Mason–Pfizer monkey retrovirus (M-PMV dUTPase) is a betaretroviral member of the dUTPase enzyme family. The nucleocapsid-free dUTPase (48426 Da) was co-crystallized with a dUTP substrate analogue using the hanging-drop vapour-diffusion method. Deoxyuridine 5′-triphosphate nucleotidohydrolase from Mason–Pfizer monkey retrovirus (M-PMV dUTPase) is a betaretroviral member of the dUTPase enzyme family. In the mature M-PMV virion, this enzyme is present as the C-terminal domain of the fusion protein nucleocapsid-dUTPase. The homotrimeric organization characteristic of dUTPases is retained in this bifunctional fusion protein. The fusion protein supposedly plays a role in adequate localization of dUTPase activity in the vicinitymore » of nucleic acids during reverse transcription and integration. Here, the nucleocapsid-free dUTPase (48 426 Da) was cocrystallized with a dUTP substrate analogue using the hanging-drop vapour-diffusion method. The obtained crystals belong to the primitive hexagonal space group P6{sub 3}, with unit-cell parameters a = 60.6, b = 60.6, c = 63.6 Å, α = 90, β = 90, γ = 120°. Native and PtCl{sub 4}-derivative data sets were collected using synchrotron radiation to 1.75 and 2.3 Å, respectively. Phasing was successfully performed by isomorphous replacement combined with anomalous scattering.« less

Messenger RNA- versus retrovirus-based induced pluripotent stem cell reprogramming strategies: analysis of genomic integrity.

PubMed

Steichen, Clara; Luce, Eléanor; Maluenda, Jérôme; Tosca, Lucie; Moreno-Gimeno, Inmaculada; Desterke, Christophe; Dianat, Noushin; Goulinet-Mainot, Sylvie; Awan-Toor, Sarah; Burks, Deborah; Marie, Joëlle; Weber, Anne; Tachdjian, Gérard; Melki, Judith; Dubart-Kupperschmitt, Anne

2014-06-01

The use of synthetic messenger RNAs to generate human induced pluripotent stem cells (iPSCs) is particularly appealing for potential regenerative medicine applications, because it overcomes the common drawbacks of DNA-based or virus-based reprogramming strategies, including transgene integration in particular. We compared the genomic integrity of mRNA-derived iPSCs with that of retrovirus-derived iPSCs generated in strictly comparable conditions, by single-nucleotide polymorphism (SNP) and copy number variation (CNV) analyses. We showed that mRNA-derived iPSCs do not differ significantly from the parental fibroblasts in SNP analysis, whereas retrovirus-derived iPSCs do. We found that the number of CNVs seemed independent of the reprogramming method, instead appearing to be clone-dependent. Furthermore, differentiation studies indicated that mRNA-derived iPSCs differentiated efficiently into hepatoblasts and that these cells did not load additional CNVs during differentiation. The integration-free hepatoblasts that were generated constitute a new tool for the study of diseased hepatocytes derived from patients' iPSCs and their use in the context of stem cell-derived hepatocyte transplantation. Our findings also highlight the need to conduct careful studies on genome integrity for the selection of iPSC lines before using them for further applications. ©AlphaMed Press.

Role of the human endogenous retrovirus HERV-K18 in autoimmune disease susceptibility: study in the Spanish population and meta-analysis.

PubMed

de la Hera, Belén; Varadé, Jezabel; García-Montojo, Marta; Lamas, José Ramón; de la Encarnación, Ana; Arroyo, Rafael; Fernández-Gutiérrez, Benjamín; Alvarez-Lafuente, Roberto; Urcelay, Elena

2013-01-01

Human endogenous retroviruses (HERVs) are genomic sequences that resulted from ancestral germ-line infections by exogenous retroviruses and therefore are transmitted in a Mendelian fashion. Increased HERV expression and antibodies to HERV antigens have been found in various autoimmune diseases. HERV-K18 in chromosome 1 was previously associated with type one diabetes and multiple sclerosis (MS). The etiology of these complex conditions has not been completely elucidated even after the powerful genome wide association studies (GWAS) performed. Nonetheless, this approach does not scrutinize the repetitive sequences within the genome, and part of the missing heritability could lie behind these sequences. We aimed at evaluating the role of HERV-K18 in chromosome 1 on autoimmune disease susceptibility. Two HERV-K18 SNPs (97Y/C and 154W/Stop substitutions) conforming three haplotypes were genotyped in Spanish cohorts of multiple sclerosis (n = 942), rheumatoid arthritis (n = 462) and ethnically matched controls (n = 601). Our findings were pooled in a meta-analysis including 5312 autoimmune patients and 4032 controls. Significant associations of both HERV-K18 polymorphisms in chromosome 1 with MS patients stratified by HLA-DRB1*15:01 were observed [97Y/C p = 0.02; OR (95% CI) = 1.5 (1.04-2.17) and 154W/Stop: p = 0.001; OR (95% CI) = 1.6 (1.19-2.16)]. Combined meta-analysis of the previously published association studies of HERV-K18 with different autoimmune diseases, together with data derived from Spanish cohorts, yielded a significant association of the HERV-K18.3 haplotype [97Y-154W: p(M-H) = 0.0008; OR(M-H) (95% CI) = 1.22 (1.09-1.38)]. Association of the HERV-K18.3 haplotype in chromosome 1 with autoimmune-disease susceptibility was confirmed through meta-analysis.

Differentially-Expressed Pseudogenes in HIV-1 Infection.

PubMed

Gupta, Aditi; Brown, C Titus; Zheng, Yong-Hui; Adami, Christoph

2015-09-29

Not all pseudogenes are transcriptionally silent as previously thought. Pseudogene transcripts, although not translated, contribute to the non-coding RNA pool of the cell that regulates the expression of other genes. Pseudogene transcripts can also directly compete with the parent gene transcripts for mRNA stability and other cell factors, modulating their expression levels. Tissue-specific and cancer-specific differential expression of these "functional" pseudogenes has been reported. To ascertain potential pseudogene:gene interactions in HIV-1 infection, we analyzed transcriptomes from infected and uninfected T-cells and found that 21 pseudogenes are differentially expressed in HIV-1 infection. This is interesting because parent genes of one-third of these differentially-expressed pseudogenes are implicated in HIV-1 life cycle, and parent genes of half of these pseudogenes are involved in different viral infections. Our bioinformatics analysis identifies candidate pseudogene:gene interactions that may be of significance in HIV-1 infection. Experimental validation of these interactions would establish that retroviruses exploit this newly-discovered layer of host gene expression regulation for their own benefit.

Differentially-Expressed Pseudogenes in HIV-1 Infection

PubMed Central

Gupta, Aditi; Brown, C. Titus; Zheng, Yong-Hui; Adami, Christoph

2015-01-01

Not all pseudogenes are transcriptionally silent as previously thought. Pseudogene transcripts, although not translated, contribute to the non-coding RNA pool of the cell that regulates the expression of other genes. Pseudogene transcripts can also directly compete with the parent gene transcripts for mRNA stability and other cell factors, modulating their expression levels. Tissue-specific and cancer-specific differential expression of these “functional” pseudogenes has been reported. To ascertain potential pseudogene:gene interactions in HIV-1 infection, we analyzed transcriptomes from infected and uninfected T-cells and found that 21 pseudogenes are differentially expressed in HIV-1 infection. This is interesting because parent genes of one-third of these differentially-expressed pseudogenes are implicated in HIV-1 life cycle, and parent genes of half of these pseudogenes are involved in different viral infections. Our bioinformatics analysis identifies candidate pseudogene:gene interactions that may be of significance in HIV-1 infection. Experimental validation of these interactions would establish that retroviruses exploit this newly-discovered layer of host gene expression regulation for their own benefit. PMID:26426037

Sequence and Transcriptional Analyses of the Fish Retroviruses Walleye Epidermal Hyperplasia Virus Types 1 and 2: Evidence for a Gene Duplication

PubMed Central

LaPierre, Lorie A.; Holzschu, Donald L.; Bowser, Paul R.; Casey, James W.

1999-01-01

Walleye epidermal hyperplasia virus types 1 and 2 (WEHV1 and WEHV2, respectively) are associated with a hyperproliferative skin lesion on walleyes that appears and regresses seasonally. We have determined the complete nucleotide sequences and transcriptional profiles of these viruses. WEHV1 and WEHV2 are large, complex retroviruses of 12,999 and 13,125 kb in length, respectively, that are closely related to one another and to walleye dermal sarcoma virus (WDSV). These walleye retroviruses contain three open reading frames, orfA, orfB, and orfC, in addition to gag, pol, and env. orfA and orfB are adjacent to one another and located downstream of env. The OrfA proteins were previously identified as cyclin D homologs that may contribute to the induction of cell proliferation leading to epidermal hyperplasia and dermal sarcoma. The sequence analysis of WEHV1 and WEHV2 revealed that the OrfB proteins are distantly related to the OrfA proteins, suggesting that orfB arose by gene duplication. Presuming that the precursor of orfA and orfB was derived from a cellular cyclin, these genes are the first accessory genes of complex retroviruses that can be traced to a cellular origin. WEHV1, WEHV2, and WDSV are the only retroviruses that have an open reading frame, orfC, of considerable size (ca. 130 amino acids) in the leader region preceding gag. While we were unable to predict a function for the OrfC proteins, they are more conserved than OrfA and OrfB, suggesting that they may be biologically important to the viruses. The transcriptional profiles of WEHV1 and WEHV2 were also similar to that of WDSV; Northern blot analyses detected only low levels of the orfA transcripts in developing lesions, whereas abundant levels of genomic, env, orfA, and orfB transcripts were detected in regressing lesions. The splice donors and acceptors of individual transcripts were identified by reverse transcriptase PCR. The similarities of WEHV1, WEHV2, and WDSV suggest that these viruses use

Jaagsiekte Sheep Retrovirus Envelope Efficiently Pseudotypes Human Immunodeficiency Virus Type 1-Based Lentiviral Vectors

PubMed Central

Liu, Shan-Lu; Halbert, Christine L.; Miller, A. Dusty

2004-01-01

Jaagsiekte sheep retrovirus (JSRV) infects lung epithelial cells in sheep, and oncoretroviral vectors bearing JSRV Env can mediate transduction of human cells, suggesting that such vectors might be useful for lung-directed gene therapy. Here we show that JSRV Env can also efficiently pseudotype a human immunodeficiency virus type 1-based lentiviral vector, a more suitable vector for transduction of slowly dividing lung epithelial cells. We created several chimeric Env proteins that, unlike the parental Env, do not transform rodent fibroblasts but are still capable of pseudotyping lentiviral and oncoretroviral vectors. PMID:14963173

Type W Human Endogenous Retrovirus (HERV-W) Integrations and Their Mobilization by L1 Machinery: Contribution to the Human Transcriptome and Impact on the Host Physiopathology.

PubMed

Grandi, Nicole; Tramontano, Enzo

2017-06-27

Human Endogenous Retroviruses (HERVs) are ancient infection relics constituting ~8% of our DNA. While HERVs' genomic characterization is still ongoing, impressive amounts of data have been obtained regarding their general expression across tissues. Among HERVs, one of the most studied is the W group, which is the sole HERV group specifically mobilized by the long interspersed element-1 (LINE-1) machinery, providing a source of novel insertions by retrotransposition of HERV-W processed pseudogenes, and comprising a member encoding a functional envelope protein coopted for human placentation. The HERV-W group has been intensively investigated for its putative role in several diseases, such as cancer, inflammation, and autoimmunity. Despite major interest in the link between HERV-W expression and human pathogenesis, no conclusive correlation has been demonstrated so far. In general, (i) the absence of a proper identification of the specific HERV-W sequences expressed in a given condition, and (ii) the lack of studies attempting to connect the various observations in the same experimental conditions are the major problems preventing the definitive assessment of the HERV-W impact on human physiopathology. In this review, we summarize the current knowledge on the HERV-W group presence within the human genome and its expression in physiological tissues as well as in the main pathological contexts.

A transcriptome resource for the koala (Phascolarctos cinereus): insights into koala retrovirus transcription and sequence diversity.

PubMed

Hobbs, Matthew; Pavasovic, Ana; King, Andrew G; Prentis, Peter J; Eldridge, Mark D B; Chen, Zhiliang; Colgan, Donald J; Polkinghorne, Adam; Wilkins, Marc R; Flanagan, Cheyne; Gillett, Amber; Hanger, Jon; Johnson, Rebecca N; Timms, Peter

2014-09-11

The koala, Phascolarctos cinereus, is a biologically unique and evolutionarily distinct Australian arboreal marsupial. The goal of this study was to sequence the transcriptome from several tissues of two geographically separate koalas, and to create the first comprehensive catalog of annotated transcripts for this species, enabling detailed analysis of the unique attributes of this threatened native marsupial, including infection by the koala retrovirus. RNA-Seq data was generated from a range of tissues from one male and one female koala and assembled de novo into transcripts using Velvet-Oases. Transcript abundance in each tissue was estimated. Transcripts were searched for likely protein-coding regions and a non-redundant set of 117,563 putative protein sequences was produced. In similarity searches there were 84,907 (72%) sequences that aligned to at least one sequence in the NCBI nr protein database. The best alignments were to sequences from other marsupials. After applying a reciprocal best hit requirement of koala sequences to those from tammar wallaby, Tasmanian devil and the gray short-tailed opossum, we estimate that our transcriptome dataset represents approximately 15,000 koala genes. The marsupial alignment information was used to look for potential gene duplications and we report evidence for copy number expansion of the alpha amylase gene, and of an aldehyde reductase gene.Koala retrovirus (KoRV) transcripts were detected in the transcriptomes. These were analysed in detail and the structure of the spliced envelope gene transcript was determined. There was appreciable sequence diversity within KoRV, with 233 sites in the KoRV genome showing small insertions/deletions or single nucleotide polymorphisms. Both koalas had sequences from the KoRV-A subtype, but the male koala transcriptome has, in addition, sequences more closely related to the KoRV-B subtype. This is the first report of a KoRV-B-like sequence in a wild population. This transcriptomic

Evolutionary Relationships among Extinct and Extant Sloths: The Evidence of Mitogenomes and Retroviruses

PubMed Central

Slater, Graham J.; Cui, Pin; Forasiepi, Analía M.; Lenz, Dorina; Tsangaras, Kyriakos; Voirin, Bryson; de Moraes-Barros, Nadia; MacPhee, Ross D. E.; Greenwood, Alex D.

2016-01-01

Macroevolutionary trends exhibited by retroviruses are complex and not entirely understood. The sloth endogenized foamy-like retrovirus (SloEFV), which demonstrates incongruence in virus–host evolution among extant sloths (Order Folivora), has not been investigated heretofore in any extinct sloth lineages and its premodern history within folivorans is therefore unknown. Determining retroviral coevolutionary trends requires a robust phylogeny of the viral host, but the highly reduced modern sloth fauna (6 species in 2 genera) does not adequately represent what was once a highly diversified clade (∼100 genera) of placental mammals. At present, the amount of molecular data available for extinct sloth taxa is limited, and analytical results based on these data tend to conflict with phylogenetic inferences made on the basis of morphological studies. To augment the molecular data set, we applied hybridization capture and next-generation Illumina sequencing to two extinct and three extant sloth species to retrieve full mitochondrial genomes (mitogenomes) from the hosts and the polymerase gene of SloEFV. The results produced a fully resolved and well-supported phylogeny that supports dividing crown families into two major clades: 1) The three-toed sloth, Bradypus, and Nothrotheriidae and 2) Megalonychidae, including the two-toed sloth, Choloepus, and Mylodontidae. Our calibrated time tree indicates that the Miocene epoch (23.5 Ma), particularly its earlier part, was an important interval for folivoran diversification. Both extant and extinct sloths demonstrate multiple complex invasions of SloEFV into the ancestral sloth germline followed by subsequent introgressions across different sloth lineages. Thus, sloth mitogenome and SloEFV evolution occurred separately and in parallel among sloths. PMID:26878870

Human retroviruses and AIDS 1997

DOE Office of Scientific and Technical Information (OSTI.GOV)

Korber, B.; Foley, B.; Leitner, T.

1997-12-01

This compendium is the result of an effort to compile, organize, and rapidly publish as much relevant molecular data concerning the human immunodeficiency viruses (HIV) and related retroviruses as possible. The scope of the compendium and database is best summarized by the four parts that it comprises: (1) Nucleic Acid Alignments, (2) Amino Acid Alignments, (3) Reviews and Analyses, and (4) Related Sequences. Information within all the parts is updated throughout the year on the Web site, http://hiv-web.lanl.gov. This year we are not including floppy diskettes as the entire compendium is available both at our Web site and at ourmore » ftp site. If you need floppy diskettes please contact either Bette Korber (btk@t10.lanl.gov) or Kersti Rock (karm@t10.lanl.gov) by email or fax ((505) 665-4453). While this publication could take the form of a review or sequence monograph, it is not so conceived. Instead, the literature from which the database is derived has simply been summarized and some elementary computational analyses have been performed upon the data. Interpretation and commentary have been avoided insofar as possible so that the reader can form his or her own judgments concerning the complex information. The exception to this are reviews submitted by experts in areas deemed of particular and basic importance to research involving AIDS viral sequence information. These are included in Part III, and are contributed by scientists with particular expertise in the area of interest. In addition to the general descriptions below of the parts of the compendium, the user should read the individual introductions for each part.« less

Immunotherapeutic Potential of Anti-Human Endogenous Retrovirus-K Envelope Protein Antibodies in Targeting Breast Tumors

PubMed Central

Rycaj, Kiera; Plummer, Joshua B.; Li, Ming; Yin, Bingnan; Frerich, Katherine; Garza, Jeremy G.; Shen, Jianjun; Lin, Kevin; Yan, Peisha; Glynn, Sharon A.; Dorsey, Tiffany H.; Hunt, Kelly K.; Ambs, Stefan; Johanning, Gary L.

2012-01-01

Background The envelope (env) protein of the human endogenous retrovirus type K (HERV-K) family is commonly expressed on the surface of breast cancer cells. We assessed whether HERV-K env is a potential target for antibody-based immunotherapy of breast cancer. Methods We examined the expression of HERV-K env protein in various malignant (MDA-MB-231, MCF-7, SKBR3, MDA-MB-453, T47D, and ZR-75-1) and nonmalignant (MCF-10A and MCF-10AT) human breast cell lines by immunoblot, enzyme-linked immunosorbent assay, immunofluorescence staining, and flow cytometry. Anti-HERV-K env monoclonal antibodies (mAbs; 6H5, 4D1, 4E11, 6E11, and 4E6) were used to target expression of HERV-K, and antitumor effects were assessed by quantifying growth and apoptosis of breast cancer cells in vitro, and tumor growth in vivo in mice (n = 5 per group) bearing xenograft tumors. The mechanisms responsible for 6H5 mAb–mediated effects were investigated by microarray assays, flow cytometry, immunoblot, and immunofluorescence staining. The expression of HERV-K env protein was assessed in primary breast tumors (n = 223) by immunohistochemistry. All statistical tests were two-sided. Results The expression of HERV-K env protein in malignant breast cancer cell lines was substantially higher than nonmalignant breast cells. Anti–HERV-K-specific mAbs inhibited growth and induced apoptosis of breast cancer cells in vitro. Mice treated with 6H5 mAb showed statistically significantly reduced growth of xenograft tumors compared with mice treated with control immunoglobulin (control [mIgG] vs 6H5 mAb, for tumors originating from MDA-MB-231 cells, mean size = 1448.33 vs 475.44 mm3; difference = 972.89 mm3, 95% CI = 470.17 to 1475.61 mm3; P < .001). Several proteins involved in the apoptotic signaling pathways were overexpressed in vitro in 6H5 mAb–treated malignant breast cells compared with mIgG-treated control. HERV-K expression was detected in 148 (66%) of 223 primary breast tumors, and a higher rate of

Distribution of a macaque immunosuppressive type D retrovirus in neural, lymphoid, and salivary tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lackner, A.A.; Rodriguez, M.H.; Bush, C.E.

1988-06-01

Simian acquired immune deficiency syndrome (SAIDS) in rhesus macaques (Macaca mulatta) at the California Primate Research Center is caused by a type D retrovirus designated SAIDS retrovirus serotype 1 (SRV-1). This syndrome is characterized by profound immunosuppression and death associated with opportunistic infections. Neurologic signs and lesions have not been described as part of this syndrome. The distribution of SRV-1 in the salivary glands, lymph nodes, spleens, thymuses, and brains of eight virus-infected rhesus macaques was examined by immunohistochemistry. Electron microscopy, in situ RNA hybridization, and Southern blot hybridization were also performed on selected tissues to detect viral particles, RNA,more » and DNA, respectively. In seven of eight SRV-1-infected animals, the transmembrane envelope glycoprotein (gp20) of SRV-1 was present in three or more tissues, but never in the brain. In the remaining animal, no viral antigen was detected in any tissue. In this same group of animals, viral nucleic acid was detected in the lymph nodes of six of six animals by Southern blot hybridization, in the salivary glands of two of five animals by both Southern blot and in situ hybridizations, and, surprisingly, in the brains of three of three animals by Southern blot and of three of five animals by in situ hybridization, including the one animal in which viral gp20 was undetectable. None of these animals had neurologic signs or lesions. The detection of viral nucleic acid in the absence of viral antigen in the brain suggests latent SRV-1 infection of the central nervous system.« less

A murine retrovirus co-Opts YB-1, a translational regulator and stress granule-associated protein, to facilitate virus assembly.

PubMed

Bann, Darrin V; Beyer, Andrea R; Parent, Leslie J

2014-04-01

The Gag protein of the murine retrovirus mouse mammary tumor virus (MMTV) orchestrates the assembly of immature virus particles in the cytoplasm which are subsequently transported to the plasma membrane for release from the cell. The morphogenetic pathway of MMTV assembly is similar to that of Saccharomyces cerevisiae retrotransposons Ty1 and Ty3, which assemble virus-like particles (VLPs) in intracytoplasmic ribonucleoprotein (RNP) complexes. Assembly of Ty1 and Ty3 VLPs depends upon cellular mRNA processing factors, prompting us to examine whether MMTV utilizes a similar set of host proteins to facilitate viral capsid assembly. Our data revealed that MMTV Gag colocalized with YB-1, a translational regulator found in stress granules and P bodies, in intracytoplasmic foci. The association of MMTV Gag and YB-1 in cytoplasmic granules was not disrupted by cycloheximide treatment, suggesting that these sites were not typical stress granules. However, the association of MMTV Gag and YB-1 was RNA dependent, and an MMTV RNA reporter construct colocalized with Gag and YB-1 in cytoplasmic RNP complexes. Knockdown of YB-1 resulted in a significant decrease in MMTV particle production, indicating that YB-1 plays a role in MMTV capsid formation. Analysis by live-cell imaging with fluorescence recovery after photobleaching (FRAP) revealed that the population of Gag proteins localized within YB-1 complexes was relatively immobile, suggesting that Gag forms stable complexes in association with YB-1. Together, our data imply that the formation of intracytoplasmic Gag-RNA complexes is facilitated by YB-1, which promotes MMTV virus assembly. Cellular mRNA processing factors regulate the posttranscriptional fates of mRNAs, affecting localization and utilization of mRNAs under normal conditions and in response to stress. RNA viruses such as retroviruses interact with cellular mRNA processing factors that accumulate in ribonucleoprotein complexes known as P bodies and stress granules

Epigenetic Control of Prostate Cancer Metastasis: Role of Runx2 Phosphorylation

DTIC Science & Technology

2014-04-01

prostate cancer cells. In the third budget year, we achieved the following: a. Generation of retrovirus and lentivirus vectors expressing WT RUNX2 and S301A... retrovirus vectors will be developed that express β-galactosidase (negative control), wild type Runx2, S301A/S319A (non-phosphorylated) or S301E/S310E...constitutively active) Runx2 mutants. As described last year, retrovirus and lentivirus vectors were constructed to stably introduce wild type and mutant

Electron microscope detection of an endogenous infection of retrovirus-like particles in L20B cells.

PubMed

Roberts, Jason A; Thorley, Bruce R; Bruggink, Leesa D; Marshall, John A

2013-08-01

L20B cells are a cell line commonly used for the isolation of poliovirus. The current study indicates that L20B cells are chronically infected with a retrovirus-like particle that replicates in the cytoplasm and buds through the plasma membrane. The findings indicate that care is needed in the use of L20B cells for certain virus isolation studies and emphasize the importance of electron microscope studies as an adjunct to the development of diagnostic virology protocols.

Retrovirus XMRV Is Inhibited by Host Proteins and Anti-HIV Drugs AZT, Tenofovir, and Raltegravir | Center for Cancer Research

Cancer.gov

A newly discovered retrovirus, XMRV, isolated from prostate cancer tissues for the first time in 2006, has recently been reported in patients with this cancer, as well as in patients with chronic fatigue syndrome (CFS). However, five subsequent studies could not validate these reports. Since XMRV was isolated from the T and B cells of CFS patients, Vinay Pathak and his

Retrovirus-mediated transfer of a hygromycin phosphotransferase-thymidine kinase fusion gene into human CD34+ bone marrow cells.

PubMed

Akatsuka, Y; Emi, N; Kato, H; Abe, A; Tanimoto, M; Lupton, S D; Saito, H

1994-12-01

Retrovirus-mediated gene transfer into human hematopoietic stem cells has been proposed as a means of therapy for various inherited diseases and as a method of gene marking. The transduction efficiency of an amphotropic retroviral vector (PA317/HyTK) containing a hygromycin phosphotransferase-thymidine kinase fusion gene was examined with human CD34+ bone marrow cells in the presence of interleukin-3 (IL-3), interleukin-6 (IL-6), and stem cell factor. Transduction efficiencies determined from the ability of transduced granulocyte-macrophage colony forming units (CFU-GM) to grow in hygromycin B and from polymerase chain reaction analysis of individual transduced CFU-GM growing in the presence of hygromycin B were 0.3-3.0% (mean +/- S.D., 1.1 +/- 0.9%) and 0.1-1.2% (mean +/- S.D., 0.5 +/- 0.4%), respectively. Ganciclovir at a dose of approximately 1 microM reduced the number of CFU-GM derived from vector-infected CD34+ cells by 50%. These findings demonstrate that human hematopoietic stem cells infected with this retroviral vector are susceptible to ganciclovir, offering the potential to control transduced gene expression in vivo.

Human endogenous retrovirus K(HML-2) Gag and Env specific T-cell responses are not detected in HTLV-I-infected subjects using standard peptide screening methods.

PubMed

Jones, R Brad; Leal, Fabio E; Hasenkrug, Aaron M; Segurado, Aluisio C; Nixon, Douglas F; Ostrowski, Mario A; Kallas, Esper G

2013-01-10

An estimated 10-20 million individuals are infected with the retrovirus human T-cell leukemia virus type 1 (HTLV-1). While the majority of these individuals remain asymptomatic, 0.3-4% develop a neurodegenerative inflammatory disease, termed HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). HAM/TSP results in the progressive demyelination of the central nervous system and is a differential diagnosis of multiple sclerosis (MS). The etiology of HAM/TSP is unclear, but evidence points to a role for CNS-inflitrating T-cells in pathogenesis. Recently, the HTLV-1-Tax protein has been shown to induce transcription of the human endogenous retrovirus (HERV) families W, H and K. Intriguingly, numerous studies have implicated these same HERV families in MS, though this association remains controversial. Here, we explore the hypothesis that HTLV-1-infection results in the induction of HERV antigen expression and the elicitation of HERV-specific T-cells responses which, in turn, may be reactive against neurons and other tissues. PBMC from 15 HTLV-1-infected subjects, 5 of whom presented with HAM/TSP, were comprehensively screened for T-cell responses to overlapping peptides spanning HERV-K(HML-2) Gag and Env. In addition, we screened for responses to peptides derived from diverse HERV families, selected based on predicted binding to predicted optimal epitopes. We observed a lack of responses to each of these peptide sets. Thus, although the limited scope of our screening prevents us from conclusively disproving our hypothesis, the current study does not provide data supporting a role for HERV-specific T-cell responses in HTLV-1 associated immunopathology.

Type W Human Endogenous Retrovirus (HERV-W) Integrations and Their Mobilization by L1 Machinery: Contribution to the Human Transcriptome and Impact on the Host Physiopathology

PubMed Central

2017-01-01

Human Endogenous Retroviruses (HERVs) are ancient infection relics constituting ~8% of our DNA. While HERVs’ genomic characterization is still ongoing, impressive amounts of data have been obtained regarding their general expression across tissues. Among HERVs, one of the most studied is the W group, which is the sole HERV group specifically mobilized by the long interspersed element-1 (LINE-1) machinery, providing a source of novel insertions by retrotransposition of HERV-W processed pseudogenes, and comprising a member encoding a functional envelope protein coopted for human placentation. The HERV-W group has been intensively investigated for its putative role in several diseases, such as cancer, inflammation, and autoimmunity. Despite major interest in the link between HERV-W expression and human pathogenesis, no conclusive correlation has been demonstrated so far. In general, (i) the absence of a proper identification of the specific HERV-W sequences expressed in a given condition; and (ii) the lack of studies attempting to connect the various observations in the same experimental conditions are the major problems preventing the definitive assessment of the HERV-W impact on human physiopathology. In this review, we summarize the current knowledge on the HERV-W group presence within the human genome and its expression in physiological tissues as well as in the main pathological contexts. PMID:28653997

An avian, oncogenic retrovirus replicates in vivo in more than 50% of CD4+ and CD8+ T lymphocytes from an endangered grouse

USDA-ARS?s Scientific Manuscript database

Reoccurring infection of reticuloendotheliosis virus (REV), an avian oncogenic retrovirus, has been a major obstacle in attempts to breed and release an endangered grouse, the Attwater's prairie chicken (Tympanicus cupido attwateri). REV infection of these birds in breeding facilities was found to r...

Molecular Mechanisms of Neurodegenerative Diseases Induced by Human Retroviruses: A Review

PubMed Central

Irish, Bryan P.; Khan, Zafar K.; Jain, Pooja; Nonnemacher, Michael R.; Pirrone, Vanessa; Rahman, Saifur; Rajagopalan, Nirmala; Suchitra, Joyce B.; Mostoller, Kate; Wigdahl, Brian

2010-01-01

Problem statement Infection with retroviruses such as human immunodeficiency virus type 1 (HIV-1) and human T cell leukemia virus type 1 (HTLV-1) have been shown to lead to neurodegenerative diseases such as HIV-associated dementia (HAD) or neuroAIDS and HTLV-1-Associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP), respectively. Approach HIV-1-induced neurologic disease is associated with an influx of HIV-infected monocytic cells across the blood-brain barrier. Following neuroinvasion, HIV-1 and viral proteins, in addition to cellular mediators released from infected and uninfected cells participate in astrocytic and neuronal dysregulation, leading to mild to severe neurocognitive disorders. Results The molecular architecture of viral regulatory components including the Long Terminal Repeat (LTR), genes encoding the viral proteins Tat, Vpr and Nef as well as the envelope gene encoding gp120 and gp41 have been implicated in ‘indirect’ mechanisms of neuronal injury, mechanisms which are likely responsible for the majority of CNS damage induced by HIV-1 infection. The neuropathogenesis of HAM/TSP is linked, in part, with both intra-and extracellular effectors functions of the viral transactivator protein Tax and likely other viral proteins. Tax is traditionally known to localize in the nucleus of infected cells serving as a regulator of both viral and cellular gene expression. Conclusion/Recommendations However, recent evidence has suggested that Tax may also accumulate in the cytoplasm and be released from the infected cell through regulated cellular secretion processes. Once in the extracellular environment, Tax may cause functional alterations in cells of the peripheral blood, lymphoid organs and the central nervous system. These extracellular biological activities of Tax are likely very relevant to the neuropathogenesis of HTLV-1 and represent attractive targets for therapeutic intervention. PMID:20352020

The ability of multimerized cyclophilin A to restrict retrovirus infection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Javanbakht, Hassan; Diaz-Griffero, Felipe; Yuan Wen

2007-10-10

In owl monkeys, the typical retroviral restriction factor of primates, TRIM5{alpha}, is replaced by TRIMCyp. TRIMCyp consists of the TRIM5 RING, B-box 2 and coiled-coil domains, as well as the intervening linker regions, fused with cyclophilin A. TRIMCyp restricts infection of retroviruses, such as human immunodeficiency virus (HIV-1) and feline immunodeficiency virus (FIV), with capsids that can bind cyclophilin A. The TRIM5 coiled coil promotes the trimerization of TRIMCyp. Here we show that cyclophilin A that is oligomeric as a result of fusion with a heterologous multimer exhibits substantial antiretroviral activity. The addition of the TRIM5 RING, B-box 2 andmore » Linker 2 to oligomeric cyclophilin A generated a protein with antiretroviral activity approaching that of wild-type TRIMCyp. Multimerization increased the binding of cyclophilin A to the HIV-1 capsid, promoting accelerated uncoating of the capsid and restriction of infection.« less

Evidence of simian retrovirus type D by polymerase chain reaction.

PubMed

Hwa, Christian Z R; Tsai, Sheung Pun; Yee, JoAnn L; Van Rompay, Koen K; Roberts, Jeffrey A

2017-06-01

Over the past few years, there have been reports of finding Simian retrovirus type D (SRV) in macaque colonies where some animals were characterized as antibody positive but virus negative raising questions about how SRV was transmitted or whether there is a variant strain detected by antibody but not polymerase chain reaction (PCR) in current use. We developed a three-round nested PCR assay using degenerate primers targeting the pol gene to detect for SRV serotypes 1-5 and applied this newly validated PCR assay to test macaque DNA samples collected in China from 2010 to 2015. Using the nested PCR assay validated in this study, we found 0.15% of the samples archived on FTA ® cards were positive. The source of SRV infection identified within domestic colonies might have originated from imported macaques. The multiplex nested PCR assay developed here may supplement the current assays for SRV. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Protective effects of granulocyte colony-stimulating factor on endotoxin shock in mice with retrovirus-induced immunodeficiency syndrome.

PubMed

Toki, S; Hiromatsu, K; Aoki, Y; Makino, M; Yoshikai, Y

1997-10-01

Mice with retrovirus-induced murine acquired immunodeficiency syndrome (MAIDS) were hypersensitive to lipopolysaccharide (LPS)-induced lethal shock accompanied by marked elevations of systematic interleukin 1beta (IL-beta) and interferon gamma (IFN-gamma) after LPS challenge. Pretreatment with 10 microg of recombinant human granulocyte colony-stimulating factor (rhG-CSF) protected MAIDS mice from hypersensitivity to LPS-induced lethal shock and this protection was concomitant with suppression of IFN-gamma production. Copyright 1997 Academic Press Limited.

The xenotropic murine leukemia virus-related retrovirus debate continues at first international workshop.

PubMed

Stoye, Jonathan P; Silverman, Robert H; Boucher, Charles A; Le Grice, Stuart F J

2010-12-22

The 1st International Workshop on Xenotropic Murine Leukemia Virus-Related Retrovirus (XMRV), co-sponsored by the National Institutes of Health, The Department of Health and Human Services and Abbott Diagnostics, was convened on September 7/8, 2010 on the NIH campus, Bethesda, MD. Attracting an international audience of over 200 participants, the 2-day event combined a series of plenary talks with updates on different aspects of XMRV research, addressing basic gammaretrovirus biology, host response, association of XMRV with chronic fatigue syndrome and prostate cancer, assay development and epidemiology. The current status of XMRV research, concerns among the scientific community and suggestions for future actions are summarized in this meeting report.

Cellular Prion Protein Combined with Galectin-3 and -6 Affects the Infectivity Titer of an Endogenous Retrovirus Assayed in Hippocampal Neuronal Cells

PubMed Central

Shin, Hae-Young; Goto, Joy J.; Carp, Richard I.; Choi, Eun-Kyoung; Kim, Yong-Sun

2016-01-01

Prion diseases are infectious and fatal neurodegenerative diseases which require the cellular prion protein, PrPC, for development of diseases. The current study shows that the PrPC augments infectivity and plaque formation of a mouse endogenous retrovirus, MuLV. We have established four neuronal cell lines expressing mouse PrPC, PrP+/+; two express wild type PrPC (MoPrPwild) and the other two express mutant PrPC (MoPrPmut). Infection of neuronal cells from various PrP+/+ and PrP-/- (MoPrPKO) lines with MuLV yielded at least three times as many plaques in PrP+/+ than in PrP-/-. Furthermore, among the four PrP+/+ lines, one mutant line, P101L, had at least 2.5 times as many plaques as the other three PrP+/+ lines. Plaques in P101L were four times larger than those in other PrP+/+ lines. Colocalization of PrP and CAgag was seen in MuLV-infected PrP+/+ cells. In the PrP-MuLV interaction, the involvement of galectin-3 and -6 was observed by immunoprecipitation with antibody to PrPC. These results suggest that PrPC combined with galectin-3 and -6 can act as a receptor for MuLV. P101L, the disease form of mutant PrPC results suggest the genetic mutant form of PrPC may be more susceptible to viral infection. PMID:27936017

Multiple UBXN family members inhibit retrovirus and lentivirus production and canonical NFκΒ signaling by stabilizing IκBα.

PubMed

Hu, Yani; O'Boyle, Kaitlin; Auer, Jim; Raju, Sagar; You, Fuping; Wang, Penghua; Fikrig, Erol; Sutton, Richard E

2017-02-01

UBXN proteins likely participate in the global regulation of protein turnover, and we have shown that UBXN1 interferes with RIG-I-like receptor (RLR) signaling by interacting with MAVS and impeding its downstream effector functions. Here we demonstrate that over-expression of multiple UBXN family members decreased lentivirus and retrovirus production by several orders-of-magnitude in single cycle assays, at the level of long terminal repeat-driven transcription, and three family members, UBXN1, N9, and N11 blocked the canonical NFκB pathway by binding to Cullin1 (Cul1), inhibiting IκBα degradation. Multiple regions of UBXN1, including its UBA domain, were critical for its activity. Elimination of UBXN1 resulted in early murine embryonic lethality. shRNA-mediated knockdown of UBXN1 enhanced human immunodeficiency virus type 1 (HIV) production up to 10-fold in single cycle assays. In primary human fibroblasts, knockdown of UBXN1 caused prolonged degradation of IκBα and enhanced NFκB signaling, which was also observed after CRISPR-mediated knockout of UBXN1 in mouse embryo fibroblasts. Knockout of UBXN1 significantly up- and down-regulated hundreds of genes, notably those of several cell adhesion and immune signaling pathways. Reduction in UBXN1 gene expression in Jurkat T cells latently infected with HIV resulted in enhanced HIV gene expression, consistent with the role of UBXN1 in modulating the NFκB pathway. Based upon co-immunoprecipitation studies with host factors known to bind Cul1, models are presented as to how UBXN1 could be inhibiting Cul1 activity. The ability of UBXN1 and other family members to negatively regulate the NFκB pathway may be important for dampening the host immune response in disease processes and also re-activating quiescent HIV from latent viral reservoirs in chronically infected individuals.

Special coverage: 9th Conference on Retroviruses. TB, pneumonia deaths down, but heart attacks up.

PubMed

2002-05-01

The profile of AIDS deaths has changed since HIV-infected patients began to receive highly active antiretroviral therapy in 1996. While fewer people are now dying overall, more are dying from non-AIDS-related diseases. Research from the Centers for Disease Control and Prevention and other sources is showing an increase in non-AIDS-defining illnesses. One CDC study presented at the recent retroviruses conference showed that while there have been declines in the proportions of deaths from some AIDS-related causes, there is an increase in deaths from non-AIDS-related causes, including liver and kidney disease and possibly ischemic heart disease.

Viral causes of feline lymphoma: retroviruses and beyond.

PubMed

Beatty, Julia

2014-08-01

The most widely recognised cause of feline lymphoma is the gammaretrovirus feline leukaemia virus (FeLV). Research into the mechanisms of cellular transformation employed by FeLV and other oncogenic retroviruses has provided as much information on the regulation of eukaryotic cell growth and differentiation as it has about cancer. The recognition that a cancer has a viral cause opens up the possibility of novel treatments that spare the host from cytotoxic side-effects by specifically targeting the virus, or the host's immune response to it. The ultimate prize for viral-associated cancers is their prevention. Vaccination and changes in management practices have seen the global prevalence of FeLV infection fall and, with it, the incidence of FeLV-related cancers. Remarkably, in the face of this success, the prevalence of feline lymphoma remains high. At least one other virus, the lentivirus feline immunodeficiency virus (FIV), accounts for some of these cases. Transformation by FIV involves incompletely understood mechanisms that are distinct from those employed by FeLV. This review will focus on the current understanding of FeLV-associated and FIV-associated lymphoma and consider whether yet more viral aetiologies could be waiting to be discovered. Copyright © 2014 Elsevier Ltd. All rights reserved.

Evolution of Foamy Viruses: The Most Ancient of All Retroviruses †

PubMed Central

Rethwilm, Axel; Bodem, Jochen

2013-01-01

Recent evidence indicates that foamy viruses (FVs) are the oldest retroviruses (RVs) that we know and coevolved with their hosts for several hundred million years. This coevolution may have contributed to the non-pathogenicity of FVs, an important factor in development of foamy viral vectors in gene therapy. However, various questions on the molecular evolution of FVs remain still unanswered. The analysis of the spectrum of animal species infected by exogenous FVs or harboring endogenous FV elements in their genome is pivotal. Furthermore, animal studies might reveal important issues, such as the identification of the FV in vivo target cells, which than require a detailed characterization, to resolve the molecular basis of the accuracy with which FVs copy their genome. The issues of the extent of FV viremia and of the nature of the virion genome (RNA vs. DNA) also need to be experimentally addressed. PMID:24072062

Tumor-targeted inhibition by a novel strategy - mimoretrovirus expressing siRNA targeting the Pokemon gene.

PubMed

Tian, Zhiqiang; Wang, Huaizhi; Jia, Zhengcai; Shi, Jinglei; Tang, Jun; Mao, Liwei; Liu, Hongli; Deng, Yijing; He, Yangdong; Ruan, Zhihua; Li, Jintao; Wu, Yuzhang; Ni, Bing

2010-12-01

Pokemon gene has crucial but versatile functions in cell differentiation, proliferation and tumorigenesis. It is a master regulator of the ARF-HDM2-p53 and Rb-E2F pathways. The facts that the expression of Pokemon is essential for tumor formation and many kinds of tumors over-express the Pokemon gene make it an attractive target for therapeutic intervention for cancer treatment. In this study, we used an RNAi strategy to silence the Pokemon gene in a cervical cancer model. To address the issues involving tumor specific delivery and durable expression of siRNA, we applied the Arg-Gly-Asp (RGD) peptide ligand and polylysine (K(18)) fusion peptide to encapsulate a recombinant retrovirus plasmid expressing a siRNA targeting the Pokemon gene and produced the 'mimoretrovirus'. At charge ratio 2.0 of fusion peptide/plasmid, the mimoretrovirus formed stable and homogenous nanoparticles, and provided complete DNase I protection and complete gel retardation. This nanoparticle inhibited SiHa cell proliferation and invasion, while it promoted SiHa cell apoptosis. The binding of the nanoparticle to SiHa cells was mediated via the RGD-integrin α(v)β(3) interaction, as evidenced by the finding that unconjugated RGD peptide inhibited this binding significantly. This tumor-targeting mimoretrovirus exhibited excellent anti-tumor capacity in vivo in a nude mouse model. Moreover, the mimoretrovirus inhibited tumor growth with a much higher efficiency than recombinant retrovirus expressing siRNA or the K(18)/P4 nanoparticle lacking the RGD peptide. Results suggest that the RNAi/RGD-based mimoretrovirus developed in this study represents a novel anti-tumor strategy that may be applicable to most research involving cancer therapy and, thus, has promising potential as a cervical cancer treatment.

CMV induces HERV-K and HERV-W expression in kidney transplant recipients.

PubMed

Bergallo, Massimiliano; Galliano, Ilaria; Montanari, Paola; Gambarino, Stefano; Mareschi, Katia; Ferro, Francesca; Fagioli, Franca; Tovo, Pier-Angelo; Ravanini, Paolo

2015-07-01

Human endogenous retrovirus (HERVs) constitute approximately 8% of the human genome. Induction of HERV transcription is possible under certain circumstances, and may have a possible role in some pathological conditions. The aim of this study was to evaluate HERV-K and -W pol gene expression in kidney transplant recipients and to investigate the possible relationship between HERVs gene expression and CMV infection in these patients. Thirty-three samples of kidney transplant patients and twenty healthy blood donors were used to analyze, HERV-K and -W pol gene RNA expression by relative quantitative relative Real-Time PCR. We demonstrated that HERVs pol gene expression levels were higher in kidney transplant recipients than in healthy subjects. Moreover, HERV-K and -W pol gene expression was significantly higher in the group of kidney transplant recipients with high CMV viral load than in the groups with no or moderate CMV viral load. Our data suggest that CMV may facilitate in vivo HERV activation. Published by Elsevier B.V.

Characterization of a protein that binds multiple sequences in mammalian type C retrovirus enhancers.

PubMed Central

Sun, W; O'Connell, M; Speck, N A

1993-01-01

Mammalian type C retrovirus enhancer factor 1 (MCREF-1) is a nuclear protein that binds several directly repeated sequences (CNGGN6CNGG) in the Moloney and Friend murine leukemia virus (MLV) enhancers (N. R. Manley, M. O'Connell, W. Sun, N. A. Speck, and N. Hopkins, J. Virol. 67:1967-1975, 1993). In this paper, we describe the partial purification of MCREF-1 from calf thymus nuclei and further characterize the binding properties of MCREF-1. MCREF-1 binds four sites in the Moloney MLV enhancer and three sites in the Friend MLV enhancer. Ethylation interference analysis suggests that the MCREF-1 binding site spans two adjacent minor grooves of DNA. Images PMID:8445719

Special coverage: 9th Conference on Retroviruses. New drugs, new data hold promise for next decade of HIV treatment.

PubMed

2002-05-01

Antiretroviral research presented recently at the 9th Conference on Retroviruses and Opportunistic Infections demonstrates that investigators and pharmaceutical companies continue to strive for the next highly potent and easily tolerated anti-HIV drug. Among the new approaches are entry inhibitor drug and second-generation non-nucleoside reverse transcriptase inhibitors. New studies also looked into potency against multidrug-resistant virus and medication regimens that are simpler to take and have fewer side effects.

Decreased expression of endogenous feline leukemia virus in cat lymphomas: a case control study.

PubMed

Krunic, Milica; Ertl, Reinhard; Hagen, Benedikt; Sedlazeck, Fritz J; Hofmann-Lehmann, Regina; von Haeseler, Arndt; Klein, Dieter

2015-04-10

Cats infected with exogenous feline leukemia virus (exFeLV) have a higher chance of lymphoma development than uninfected cats. Furthermore, an increased exFeLV transcription has been detected in lymphomas compared to non-malignant tissues. The possible mechanisms of lymphoma development by exFeLV are insertional mutagenesis or persistent stimulation of host immune cells by viral antigens, bringing them at risk for malignant transformation. Vaccination of cats against exFeLV has in recent years decreased the overall infection rate in most countries. Nevertheless, an increasing number of lymphomas have been diagnosed among exFeLV-negative cats. Endogenous feline leukemia virus (enFeLV) is another retrovirus for which transcription has been observed in cat lymphomas. EnFeLV provirus elements are present in the germline of various cat species and share a high sequence similarity with exFeLV but, due to mutations, are incapable of producing infectious viral particles. However, recombination between exFeLV and enFeLV could produce infectious particles. We examined the FeLV expression in cats that have developed malignant lymphomas and discussed the possible mechanisms that could have induced malignant transformation. For expression analysis we used next-generation RNA-sequencing (RNA-Seq) and for validation reverse transcription quantitative PCR (RT-qPCR). First, we showed that there was no expression of exFeLV in all samples, which eliminates the possibility of recombination between exFeLV and enFeLV. Next, we analyzed the difference in expression of three enFeLV genes between control and lymphoma samples. Our analysis showed an average of 3.40-fold decreased viral expression for the three genes in lymphoma compared to control samples. The results were confirmed by RT-qPCR. There is a decreased expression of enFeLV genes in lymphomas versus control samples, which contradicts previous observations for the exFeLV. Our results suggest that a persistent stimulation of host

Distribution of endogenous type B and type D sheep retrovirus sequences in ungulates and other mammals.

PubMed Central

Hecht, S J; Stedman, K E; Carlson, J O; DeMartini, J C

1996-01-01

The jaagsiekte sheep retrovirus (JSRV), which appears to be a type B/D retrovirus chimera, has been incriminated as the cause of ovine pulmonary carcinoma. Recent studies suggest that the sequences related to this virus are found in the genomes of normal sheep and goats. To learn whether there are breeds of sheep that lack the endogenous viral sequences and to study their distribution among other groups of mammals, we surveyed several domestic sheep and goat breeds, other ungulates, and various mammal groups for sequences related to JSRV. Probes prepared from the envelope (SU) region of JSRV and the capsid (CA) region of a Peruvian type D virus related to JSRV were used in Southern blot hybridization with genomic DNA followed by low- and high-stringency washes. Fifteen to 20 CA and SU bands were found in all members of the 13 breeds of domestic sheep and 6 breeds of goats tested. There were similar findings in 6 wild Ovis and Capra genera. Within 22 other genera of Bovidae including domestic cattle, and 7 other families of Artiodactyla including Cervidae, there were usually a few CA or SU bands at low stringency and rare bands at high stringency. Among 16 phylogenetically distant genera, there were generally fewer bands hybridizing with either probe. These results reveal wide-spread phylogenetic distribution of endogenous type B and type D retroviral sequences related to JSRV among mammals and argue for further investigation of their potential role in disease. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:8622932

Genetic tagging of tumor cells with retrovirus vectors: Clonal analysis of tumor growth and metastasis in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Korczak; Robson, I.B.; Lamarche, C.

1988-08-01

Retrovirus vector infection was used to introduce large numbers of unique genetic markers into tumor cell populations for the purpose of analyzing comparative changes in the clonal composition of metastatic versus that of nonmetastatic tumors during their progressive growth in vivo. The cell lines were SP1, a nonmetastatic, aneuploid mouse mammary adenocarcinoma, and SP1HU9L, a metastatic variant of SP1. Cells were infected with ..delta..e..delta..rhoMoTn, a replication-defective retrovirus vector which possesses the dominant selectable neo gene and crippled long terminal repeats. G418/sup r/ colonies were obtained at a frequency of 4 x 10/sup -3/. Southern blot analysis of a number ofmore » clones provided evidence of random and heritable integration of one or two copies of the proviral DNA. Clonal equation of primary tumor growth and the nature of lineage relationships among spontaneous metastases and primary tumors were analyzed by subcutaneously injecting 10/sup 5/ cells from a pooled mixture of 3.6 x 10/sup 2/ G418/sup r/ SP1HU9L or 10/sup 4/ G418/sup r/ SP1 colonies into syngeneic CBA/J mice. The most striking finding was the relative clonal homogeneity of advanced primary tumors; they invariably consisted of a small number (less than 10) of distinct clones despite the fact that hundreds of thousands of uniquely marked clones had been injected.« less

Human endogenous retrovirus K and cancer: Innocent bystander or tumorigenic accomplice?

PubMed

Downey, Ronan F; Sullivan, Francis J; Wang-Johanning, Feng; Ambs, Stefan; Giles, Francis J; Glynn, Sharon A

2015-09-15

Harbored as relics of ancient germline infections, human endogenous retroviruses (HERVs) now constitute up to 8% of our genome. A proportion of this sequence has been co-opted for molecular and cellular processes, beneficial to human physiology, such as the fusogenic activity of the envelope protein, a vital component of placentogenesis. However, the discovery of high levels of HERV-K mRNA and protein and even virions in a wide array of cancers has revealed that HERV-K may be playing a more sinister role-a role as an etiological agent in cancer itself. Whether the presence of this retroviral material is simply an epiphenomenon, or an actual causative factor, is a hotly debated topic. This review will summarize the current state of knowledge regarding HERV-K and cancer and attempt to outline the potential mechanisms by which HERV-K could be involved in the onset and promotion of carcinogenesis. © 2014 The Authors. Published by Wiley Periodicals, Inc. on behalf of UICC.

Characterization of an endogenous retrovirus class in elephants and their relatives

PubMed Central

Greenwood, Alex D; Englbrecht, Claudia C; MacPhee, Ross DE

2004-01-01

Background Endogenous retrovirus-like elements (ERV-Ls, primed with tRNA leucine) are a diverse group of reiterated sequences related to foamy viruses and widely distributed among mammals. As shown in previous investigations, in many primates and rodents this class of elements has remained transpositionally active, as reflected by increased copy number and high sequence diversity within and among taxa. Results Here we examine whether proviral-like sequences may be suitable molecular probes for investigating the phylogeny of groups known to have high element diversity. As a test we characterized ERV-Ls occurring in a sample of extant members of superorder Uranotheria (Asian and African elephants, manatees, and hyraxes). The ERV-L complement in this group is even more diverse than previously suspected, and there is sequence evidence for active expansion, particularly in elephantids. Many of the elements characterized have protein coding potential suggestive of activity. Conclusions In general, the evidence supports the hypothesis that the complement had a single origin within basal Uranotheria. PMID:15476555

Down-regulation of human endogenous retrovirus type K (HERV-K) viral env RNA in pancreatic cancer cells decreases cell proliferation and tumor growth

PubMed Central

Li, Ming; Radvanyi, Laszlo; Yin, Bingnan; Li, Jia; Chivukula, Raghavender; Lin, Kevin; Lu, Yue; Shen, JianJun; Chang, David Z.; Li, Donghui; Johanning, Gary L.; Wang-Johanning, Feng

2017-01-01

Purpose We investigated the role of the human endogenous retrovirus type K (HERV-K) envelope (env) gene in pancreatic cancer (PC). Experimental Design shRNA was employed to knockdown (KD) the expression of HERV-K in PC cells. Results HERV-K env expression was detected in seven PC cell lines and in 80% of PC patient biopsies, but not in two normal pancreatic cell lines or uninvolved normal tissues. A new HERV-K splice variant was discovered in several PC cell lines. RT activity and virus-like particles were observed in culture media supernatant obtained from Panc-1 and Panc-2 cells. HERV-K viral RNA levels and anti-HERV-K antibody titers were significantly higher in PC patient sera (N=106) than in normal donor sera (N=40). Importantly, the in vitro and in vivo growth rates of three PC cell lines were significantly reduced after HERV-K KD by shRNA targeting HERV-K env, and there was reduced metastasis to lung after treatment. RNA-seq results revealed changes in gene expression after HERV-K env KD, including RAS and TP53. Furthermore, downregulation of HERV-K Env protein expression by shRNA also resulted in decreased expression of RAS, p-ERK, p-RSK, and p-AKT in several PC cells or tumors. Conclusion These results demonstrate that HERV-K influences signal transduction via the RAS-ERK-RSK pathway in PC. Our data highlight the potentially important role of HERV-K in tumorigenesis and progression of PC, and indicate that HERV-K viral proteins may be attractive biomarkers and/or tumor-associated antigens, as well as potentially useful targets for detection, diagnosis and immunotherapy of PC. PMID:28679769

Expression of endogenous retroviruses is negatively regulated by the pluripotency marker Rex1/Zfp42

PubMed Central

Guallar, D.; Pérez-Palacios, R.; Climent, M.; Martínez-Abadía, I.; Larraga, A.; Fernández-Juan, M.; Vallejo, C.; Muniesa, P.; Schoorlemmer, J.

2012-01-01

Rex1/Zfp42 is a Yy1-related zinc-finger protein whose expression is frequently used to identify pluripotent stem cells. We show that depletion of Rex1 levels notably affected self-renewal of mouse embryonic stem (ES) cells in clonal assays, in the absence of evident differences in expression of marker genes for pluripotency or differentiation. By contrast, marked differences in expression of several endogenous retroviral elements (ERVs) were evident upon Rex1 depletion. We demonstrate association of REX1 to specific elements in chromatin-immunoprecipitation assays, most strongly to muERV-L and to a lower extent to IAP and musD elements. Rex1 regulates muERV-L expression in vivo, as we show altered levels upon transient gain-and-loss of Rex1 function in pre-implantation embryos. We also find REX1 can associate with the lysine-demethylase LSD1/KDM1A, suggesting they act in concert. Similar to REX1 binding to retrotransposable elements (REs) in ES cells, we also detected binding of the REX1 related proteins YY1 and YY2 to REs, although the binding preferences of the two proteins were slightly different. Altogether, we show that Rex1 regulates ERV expression in mouse ES cells and during pre-implantation development and suggest that Rex1 and its relatives have evolved as regulators of endogenous retroviral transcription. PMID:22844087

Identification and characterization of avian retroviruses in chicken embryo-derived yellow fever vaccines: investigation of transmission to vaccine recipients.

PubMed

Hussain, Althaf I; Johnson, Jeffrey A; Da Silva Freire, Marcos; Heneine, Walid

2003-01-01

All currently licensed yellow fever (YF) vaccines are propagated in chicken embryos. Recent studies of chick cell-derived measles and mumps vaccines show evidence of two types of retrovirus particles, the endogenous avian retrovirus (EAV) and the endogenous avian leukosis virus (ALV-E), which originate from the chicken embryonic fibroblast substrates. In this study, we investigated substrate-derived avian retrovirus contamination in YF vaccines currently produced by three manufacturers (YF-vax [Connaught Laboratories], Stamaril [Aventis], and YF-FIOCRUZ [FIOCRUZ-Bio-Manguinhos]). Testing for reverse transcriptase (RT) activity was not possible because of assay inhibition. However, Western blot analysis of virus pellets with anti-ALV RT antiserum detected three distinct RT proteins in all vaccines, indicating that more than one source is responsible for the RTs present in the vaccines. PCR analysis of both chicken substrate DNA and particle-associated RNA from the YF vaccines showed no evidence of the long terminal repeat sequences of exogenous ALV subgroups A to D in any of the vaccines. In contrast, both ALV-E and EAV particle-associated RNA were detected at equivalent titers in each vaccine by RT-PCR. Quantitative real-time RT-PCR revealed 61,600, 348,000, and 1,665,000 ALV-E RNA copies per dose of Stamaril, YF-FIOCRUZ, and YF-vax vaccines, respectively. ev locus-specific PCR testing of the vaccine-associated chicken substrate DNA was positive both for the nondefective ev-12 locus in two vaccines and for the defective ev-1 locus in all three vaccines. Both intact and ev-1 pol sequences were also identified in the particle-associated RNA. To investigate the risks of transmission, serum samples from 43 YF vaccine recipients were studied. None of the samples were seropositive by an ALV-E-based Western blot assay or had detectable EAV or ALV-E RNA sequences by RT-PCR. YF vaccines produced by the three manufacturers all have particles containing EAV genomes and

Restriction by APOBEC3 proteins of endogenous retroviruses with an extracellular life cycle: ex vivo effects and in vivo "traces" on the murine IAPE and human HERV-K elements

PubMed Central

Esnault, Cécile; Priet, Stéphane; Ribet, David; Heidmann, Odile; Heidmann, Thierry

2008-01-01

Background APOBEC3 cytosine deaminases have been demonstrated to restrict infectivity of a series of retroviruses, with different efficiencies depending on the retrovirus. In addition, APOBEC3 proteins can severely restrict the intracellular transposition of a series of retroelements with a strictly intracellular life cycle, including the murine IAP and MusD LTR-retrotransposons. Results Here we show that the IAPE element, which is the infectious progenitor of the strictly intracellular IAP elements, and the infectious human endogenous retrovirus HERV-K are restricted by both murine and human APOBEC3 proteins in an ex vivo assay for infectivity, with evidence in most cases of strand-specific G-to-A editing of the proviruses, with the expected signatures. In silico analysis of the naturally occurring genomic copies of the corresponding endogenous elements performed on the mouse and human genomes discloses "traces" of APOBEC3-editing, with the specific signature of the murine APOBEC3 and human APOBEC3G enzymes, respectively, and to a variable extent depending on the family member. Conclusion These results indicate that the IAPE and HERV-K elements, which can only replicate via an extracellular infection cycle, have been restricted at the time of their entry, amplification and integration into their target host genomes by definite APOBEC3 proteins, most probably acting in evolution to limit the mutagenic effect of these endogenized extracellular parasites. PMID:18702815

Retroviral expression screening of oncogenes in natural killer cell leukemia.

PubMed

Choi, Young Lim; Moriuchi, Ryozo; Osawa, Mitsujiro; Iwama, Atsushi; Makishima, Hideki; Wada, Tomoaki; Kisanuki, Hiroyuki; Kaneda, Ruri; Ota, Jun; Koinuma, Koji; Ishikawa, Madoka; Takada, Shuji; Yamashita, Yoshihiro; Oshimi, Kazuo; Mano, Hiroyuki

2005-08-01

Aggressive natural killer cell leukemia (ANKL) is an intractable malignancy that is characterized by the outgrowth of NK cells. To identify transforming genes in ANKL, we constructed a retroviral cDNA expression library from an ANKL cell line KHYG-1. Infection of 3T3 cells with recombinant retroviruses yielded 33 transformed foci. Nucleotide sequencing of the DNA inserts recovered from these foci revealed that 31 of them encoded KRAS2 with a glycine-to-alanine mutation at codon 12. Mutation-specific PCR analysis indicated that the KRAS mutation was present only in KHYG-1 cells, not in another ANKL cell line or in clinical specimens (n=8).

Endogenous Retrovirus Insertion in the KIT Oncogene Determines White and White spotting in Domestic Cats

PubMed Central

David, Victor A.; Menotti-Raymond, Marilyn; Wallace, Andrea Coots; Roelke, Melody; Kehler, James; Leighty, Robert; Eizirik, Eduardo; Hannah, Steven S.; Nelson, George; Schäffer, Alejandro A.; Connelly, Catherine J.; O’Brien, Stephen J.; Ryugo, David K.

2014-01-01

The Dominant White locus (W) in the domestic cat demonstrates pleiotropic effects exhibiting complete penetrance for absence of coat pigmentation and incomplete penetrance for deafness and iris hypopigmentation. We performed linkage analysis using a pedigree segregating White to identify KIT (Chr. B1) as the feline W locus. Segregation and sequence analysis of the KIT gene in two pedigrees (P1 and P2) revealed the remarkable retrotransposition and evolution of a feline endogenous retrovirus (FERV1) as responsible for two distinct phenotypes of the W locus, Dominant White, and white spotting. A full-length (7125 bp) FERV1 element is associated with white spotting, whereas a FERV1 long terminal repeat (LTR) is associated with all Dominant White individuals. For purposes of statistical analysis, the alternatives of wild-type sequence, FERV1 element, and LTR-only define a triallelic marker. Taking into account pedigree relationships, deafness is genetically linked and associated with this marker; estimated P values for association are in the range of 0.007 to 0.10. The retrotransposition interrupts a DNAase I hypersensitive site in KIT intron 1 that is highly conserved across mammals and was previously demonstrated to regulate temporal and tissue-specific expression of KIT in murine hematopoietic and melanocytic cells. A large-population genetic survey of cats (n = 270), representing 30 cat breeds, supports our findings and demonstrates statistical significance of the FERV1 LTR and full-length element with Dominant White/blue iris (P < 0.0001) and white spotting (P < 0.0001), respectively. PMID:25085922

Endogenous retrovirus insertion in the KIT oncogene determines white and white spotting in domestic cats.

PubMed

David, Victor A; Menotti-Raymond, Marilyn; Wallace, Andrea Coots; Roelke, Melody; Kehler, James; Leighty, Robert; Eizirik, Eduardo; Hannah, Steven S; Nelson, George; Schäffer, Alejandro A; Connelly, Catherine J; O'Brien, Stephen J; Ryugo, David K

2014-08-01

The Dominant White locus (W) in the domestic cat demonstrates pleiotropic effects exhibiting complete penetrance for absence of coat pigmentation and incomplete penetrance for deafness and iris hypopigmentation. We performed linkage analysis using a pedigree segregating White to identify KIT (Chr. B1) as the feline W locus. Segregation and sequence analysis of the KIT gene in two pedigrees (P1 and P2) revealed the remarkable retrotransposition and evolution of a feline endogenous retrovirus (FERV1) as responsible for two distinct phenotypes of the W locus, Dominant White, and white spotting. A full-length (7125 bp) FERV1 element is associated with white spotting, whereas a FERV1 long terminal repeat (LTR) is associated with all Dominant White individuals. For purposes of statistical analysis, the alternatives of wild-type sequence, FERV1 element, and LTR-only define a triallelic marker. Taking into account pedigree relationships, deafness is genetically linked and associated with this marker; estimated P values for association are in the range of 0.007 to 0.10. The retrotransposition interrupts a DNAase I hypersensitive site in KIT intron 1 that is highly conserved across mammals and was previously demonstrated to regulate temporal and tissue-specific expression of KIT in murine hematopoietic and melanocytic cells. A large-population genetic survey of cats (n = 270), representing 30 cat breeds, supports our findings and demonstrates statistical significance of the FERV1 LTR and full-length element with Dominant White/blue iris (P < 0.0001) and white spotting (P < 0.0001), respectively. Copyright © 2014 David et al.

Inhibition effect of small interfering RNA of connective tissue growth factor on the expression of extracellular matrix molecules in cultured human renal proximal tubular cells.

PubMed

Liu, Yuyuan; Li, Weiwei; Liu, Hong; Peng, Youming; Yang, Qiu; Xiao, Li; Liu, Yinghong; Liu, Fuyou

2014-03-01

In this study, we investigated the effect of small interfering RNA (siRNA) of connective tissue growth factor (CTGF) by pRetro-Super (PRS) retrovirus vector on the expression of CTGF and related extracellular matrix molecules in human renal proximal tubular cells (HKCs) induced by high glucose, to provide help for renal tubulointerstitial fibrosis therapy. HKCs were exposed to d-glucose to observe their dose and time effect, while the mannitol as osmotic control. Retrovirus producing CTGF siRNA were constructed from the inverted oligonucleotides and transferred into packaging cell line PT67 with lipofectamine, and the virus supernatant was used to infect HKC. The expression of CTGF, fibronectin (FN) and collagen-type I (col1) were measured by semi-quantitative RT-PCR and Western blot. In response to high glucose, CTGF expression in HKCs was increased in a dose- and time-dependent manner, whereas the increase did not occur in the osmotic control. Introduction of PRS-CTGF-siRNA resulted in the significant reduction of CTGF, FN, col1 mRNA (p < 0.01, respectively) and CTGF, col1 protein (p < 0.05, respectively) expression, while PRS void vector group did not have these effects (p > 0.05). CTGF siRNA therapy can effectively reduce the levels of CTGF, FN and col1 induced by high glucose in cultured HKCs, which suggested that it may be a potential therapeutic strategy to prevent the renal interstitial fibrosis in the future.

Highlights from the 24th Conference on Retroviruses and Opportunistic Infections: 13-16 February 2017, Seattle, Washington, USA.

PubMed

Crowell, Trevor A; Lyall, Hermione; Malatinkova, Eva; Bhagani, Sanjay; Hsu, Denise; Colby, Donn J; Polyak, Christina; Psomas, Christina; Hill, Andrew; Gathogo, Esther N; Trypsteen, Wim; Vandekerckhove, Linos; Kinloch, Sabine

2017-04-01

From the 13th to 16th February 2017, researchers from around the world convened for the 24th annual Conference on Retroviruses and Opportunistic Infections (CROI) at the Washington State Convention Center in Seattle, Washington. The conference was organised by the International Antiviral Society-USA (IAS-USA) in partnership with the CROI Foundation. The conference included over 1000 oral and poster presentations of peer-reviewed original research as well as lectures and symposia featuring insights from leading basic, translational and clinical researchers. Highlighted here are key data presented at the conference.

Transcriptional Profiling of Human Endogenous Retrovirus Group HERV-K(HML-2) Loci in Melanoma

PubMed Central

Schmitt, Katja; Reichrath, Jörg; Roesch, Alexander; Meese, Eckart; Mayer, Jens

2013-01-01

Recent studies suggested a role for the human endogenous retrovirus (HERV) group HERV-K(HML-2) in melanoma because of upregulated transcription and expression of HERV-K(HML-2)-encoded proteins. Very little is known about which HML-2 loci are transcribed in melanoma. We assigned >1,400 HML-2 cDNA sequences generated from various melanoma and related samples to genomic HML-2 loci, identifying a total of 23 loci as transcribed. Transcription profiles of loci differed significantly between samples. One locus was found transcribed only in melanoma-derived samples but not in melanocytes and might represent a marker for melanoma. Several of the transcribed loci harbor ORFs for retroviral Gag and/or Env proteins. Env-encoding loci were transcribed only in melanoma. Specific investigation of rec and np9 transcripts indicated transcription of protein encoding loci in melanoma and melanocytes hinting at the relevance of Rec and Np9 in melanoma. UVB irradiation changed transcription profiles of loci and overall transcript levels decreased in melanoma and melanocytes. We further identified transcribed HML-2 loci formed by reverse transcription of spliced HML-2 transcripts by L1 machinery or in a retroviral fashion, with loci potentially encoding HML-2-like proteins. We reveal complex, sample-specific transcription of HML-2 loci in melanoma and related samples. Identified HML-2 loci and proteins encoded by those loci are particularly relevant for further studying the role of HML-2 in melanoma. Transcription of HERVs appears as a complex mechanism requiring specific studies to elucidate which HERV loci are transcribed and how transcribed HERVs may be involved in disease. PMID:23338945

A deep-branching clade of retrovirus-like retrotransposons in bdelloid rotifers

PubMed Central

Gladyshev, Eugene A.; Meselson, Matthew; Arkhipova, Irina R.

2007-01-01

Rotifers of class Bdelloidea, a group of aquatic invertebrates in which males and meiosis have never been documented, are also unusual in their lack of multicopy LINE-like and gypsy-like retrotransposons, groups inhabiting the genomes of nearly all other metazoans. Bdelloids do contain numerous DNA transposons, both intact and decayed, and domesticated Penelope-like retroelements Athena, concentrated at telomeric regions. Here we describe two LTR retrotransposons, each found at low copy number in a different bdelloid species, which define a clade different from previously known clades of LTR retrotransposons. Like bdelloid DNA transposons and Athena, these elements have been found preferentially in telomeric regions. Unlike bdelloid DNA transposons, many of which are decayed, the newly described elements, named Vesta and Juno, inhabiting the genomes of Philodina roseola and Adineta vaga, respectively, appear to be intact and to represent recent insertions, possibly from an exogenous source. We describe the retrovirus-like structure of the new elements, containing gag, pol, and env-like open reading frames, and discuss their possible origins, transmission, and behavior in bdelloid genomes. PMID:17129685

Retrovirus XMRV Is Inhibited by Host Proteins and Anti-HIV Drugs AZT, Tenofovir, and Raltegravir | Center for Cancer Research

Cancer.gov

A newly discovered retrovirus, XMRV, isolated from prostate cancer tissues for the first time in 2006, has recently been reported in patients with this cancer, as well as in patients with chronic fatigue syndrome (CFS). However, five subsequent studies could not validate these reports. Since XMRV was isolated from the T and B cells of CFS patients, Vinay Pathak and his colleagues in the HIV Drug Resistance Program sought to determine how XMRV was countering intracellular defense mechanisms that inhibit retroviral replication in human cells.

Expression of Endogenous Betaretroviruses in the Ovine Uterus: Effects of Neonatal Age, Estrous Cycle, Pregnancy, and Progesterone

PubMed Central

Palmarini, Massimo; Gray, C. Allison; Carpenter, Karen; Fan, Hung; Bazer, Fuller W.; Spencer, Thomas E.

2001-01-01

The ovine genome contains 15 to 20 copies of endogenous retroviruses (enJSRVs) highly related to the oncogenic jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus. enJSRVs are highly expressed in the endometrial lumenal epithelia (LE) and glandular epithelia (GE) of the ovine uterus. The effects of neonatal age, estrous cycle, pregnancy, and progesterone on expression of enJSRVs in the ovine uterus were determined. Expression of enJSRV RNAs was absent from the uterus of ewes at birth, but enJSRV RNAs were expressed specifically in the LE and developing GE from postnatal day (PND) 7 to PND 56. In adult ewes, enJSRV RNAs were detected only in the epithelia of the uterine endometrium, as well as epithelia of the oviduct, cervix, and vagina. In cyclic ewes, endometrial enJSRV RNA abundance was lowest on day 1, increased 12-fold between days 1 and 13, and then decreased to day 15. In pregnant ewes, levels of endometrial enJSRV RNAs were high on day 11, increased to day 13, and then decreased to day 19. In day 17 and 19 conceptuses, enJSRV RNAs were also detected in binucleate cells of the trophectoderm. Immunoreactive JSRV capsid and envelope proteins were detected in the endometrial LE and GE, as well as in the binucleate cells of the conceptus. In transfection assays utilizing ovine endometrial LE cells, progesterone increased transcriptional activity of several enJSRV long terminal repeats. Collectively, these results indicate that transcription of enJSRVs in the endometrial epithelia of the ovine uterus is increased by progesterone and might support a role for enJSRVs in conceptus-endometrium interactions during the peri-implantation period and early placental morphogenesis. PMID:11689612

Heme oxygenase-1 gene expression modulates angiotensin II-induced increase in blood pressure.

PubMed

Yang, Liming; Quan, Shuo; Nasjletti, Alberto; Laniado-Schwartzman, Michal; Abraham, Nader G

2004-06-01

The heme-heme oxygenase (HO) system has been implicated in the regulation of vascular reactivity and blood pressure. This study examines the notion that overexpression of HO decreases pressor responsiveness to angiotensin II (Ang II). Five-day-old Sprague-Dawley rats received an intraleft ventricular injection of approximately 5x10(9) cfu/mL of retroviruses containing human HO-1 sense (LSN-HHO-1), rat HO-1 antisense (LSN-RHO-1-AS), or control retrovirus (LXSN). Three months later, rats were instrumented with femoral arterial and venous catheters for mean arterial pressure (MAP) determination and Ang II administration, respectively. Rats injected with LSN-HHO-1, but not with LXSN, expressed human HO-1 mRNA and protein in several tissues. BP increased with administration of Ang II in rats expressing and not expressing human HO-1. However, the Ang II-induced pressor response (mm Hg) in LSN-HHO-1 rats (16+/-3, 27+/-3, and 38+/-3 at 0.5, 2, and 10 ng) was surpassed (P<0.05) in LXSN rats (23+/-1, 37+/-2, and 52+/-2 at 0.5, 2, and 10 ng). Importantly, treating LSN-HHO-1 rats with the HO inhibitor tin mesoporphyrin (SnMP) enhanced (P<0.05) the Ang II-induced pressor response to a level not different from that observed in LXSN rats. Rats injected with LSN-RHO-1-AS showed a decrease in renal HO-1 protein expression and HO activity relative to control LXSN rats. Administration of Ang II (0.1 to 2 ng) caused small (4 to 5 mm Hg) but significant increases in MAP in rats injected with LSN-RHO-1-AS (P<0.05) compared with rats injected with LXSN. These data demonstrate that overexpression of HO-1 brings about a reduction in pressor responsiveness to Ang II, which is most likely due to increased generation of an HO-1 product, presumably CO, with the ability to inhibit vascular reactivity to constrictor stimuli.

Induction of cardiomyocyte-like cells in infarct hearts by gene transfer of Gata4, Mef2c, and Tbx5.

PubMed

Inagawa, Kohei; Miyamoto, Kazutaka; Yamakawa, Hiroyuki; Muraoka, Naoto; Sadahiro, Taketaro; Umei, Tomohiko; Wada, Rie; Katsumata, Yoshinori; Kaneda, Ruri; Nakade, Koji; Kurihara, Chitose; Obata, Yuichi; Miyake, Koichi; Fukuda, Keiichi; Ieda, Masaki

2012-10-12

After myocardial infarction (MI), massive cell death in the myocardium initiates fibrosis and scar formation, leading to heart failure. We recently found that a combination of 3 cardiac transcription factors, Gata4, Mef2c, and Tbx5 (GMT), reprograms fibroblasts directly into functional cardiomyocytes in vitro. To investigate whether viral gene transfer of GMT into infarcted hearts induces cardiomyocyte generation. Coronary artery ligation was used to generate MI in the mouse. In vitro transduction of GMT retrovirus converted cardiac fibroblasts from the infarct region into cardiomyocyte-like cells with cardiac-specific gene expression and sarcomeric structures. Injection of the green fluorescent protein (GFP) retrovirus into mouse hearts, immediately after MI, infected only proliferating noncardiomyocytes, mainly fibroblasts, in the infarct region. The GFP expression diminished after 2 weeks in immunocompetent mice but remained stable for 3 months in immunosuppressed mice, in which cardiac induction did not occur. In contrast, injection of GMT retrovirus into α-myosin heavy chain (αMHC)-GFP transgenic mouse hearts induced the expression of αMHC-GFP, a marker of cardiomyocytes, in 3% of virus-infected cells after 1 week. A pooled GMT injection into the immunosuppressed mouse hearts induced cardiac marker expression in retrovirus-infected cells within 2 weeks, although few cells showed striated muscle structures. To transduce GMT efficiently in vivo, we generated a polycistronic retrovirus expressing GMT separated by 2A "self-cleaving" peptides (3F2A). The 3F2A-induced cardiomyocyte-like cells in fibrotic tissue expressed sarcomeric α-actinin and cardiac troponin T and had clear cross striations. Quantitative RT-PCR also demonstrated that FACS-sorted 3F2A-transduced cells expressed cardiac-specific genes. GMT gene transfer induced cardiomyocyte-like cells in infarcted hearts.

Epidermal Growth Factor-Dependent Transformation by a Human EGF Receptor Proto-Oncogene

NASA Astrophysics Data System (ADS)

Velu, Thierry J.; Beguinot, Laura; Vass, William C.; Willingham, Mark C.; Merlino, Glenn T.; Pastan, Ira; Lowy, Douglas R.

1987-12-01

The epidermal growth factor (EGF) receptor gene EGFR has been placed in a retrovirus vector to examine the growth properties of cells that experimentally overproduce a full-length EGF receptor. NIH 3T3 cells transfected with the viral DNA or infected with the corresponding rescued retrovirus developed a fully transformed phenotype in vitro that required both functional EGFR expression and the presence of EGF in the growth medium. Cells expressing 4 × 105 EGF receptors formed tumors in nude mice, while control cells did not. Therefore, the EGFR retrovirus, which had a titer on NIH 3T3 cells that was greater than 107 focus-forming units per milliliter, can efficiently transfer and express this gene, and increased numbers of EGF receptors can contribute to the transformed phenotype.

Efficacy of Antiretroviral Agents against Murine Replication-Competent Retrovirus Infection in Human Cells

PubMed Central

Powell, Sharon K.; Artlip, Moria; Kaloss, Michele; Brazinski, Scott; Lyons, Russette; McGarrity, Gerard J.; Otto, Edward

1999-01-01

Retroviral vectors for gene therapy are designed to minimize the occurrence of replication-competent retrovirus (RCR); nonetheless, it is possible that a vector-derived RCR could establish an infection in a patient. Since the efficacy of antiretroviral agents can be impacted by interactions between virus, host cell, and drug, five commonly used antiretroviral drugs were evaluated for their abilities to inhibit the replication of a murine leukemia virus (MLV)-derived RCR in human cells. The results obtained indicate that the combination of nucleoside analogs zidovudine and dideoxyinosine with the protease inhibitor indinavir effectively inhibits MLV-derived RCR replication in three human cell lines. In addition, MLV-derived RCR was found to be inherently resistant to the nucleoside analogs lamivudine and stavudine, suggesting that mutations conferring resistance to nucleoside analogs in human immunodeficiency virus type 1 have the same effect even in an alternative viral backbone. PMID:10482636

Human Retroviruses and AIDS. A compilation and analysis of nucleic acid and amino acid sequences: I--II; III--V

DOE Office of Scientific and Technical Information (OSTI.GOV)

Myers, G.; Korber, B.; Wain-Hobson, S.

1993-12-31

This compendium and the accompanying floppy diskettes are the result of an effort to compile and rapidly publish all relevant molecular data concerning the human immunodeficiency viruses (HIV) and related retroviruses. The scope of the compendium and database is best summarized by the five parts that it comprises: (I) HIV and SIV Nucleotide Sequences; (II) Amino Acid Sequences; (III) Analyses; (IV) Related Sequences; and (V) Database Communications. Information within all the parts is updated at least twice in each year, which accounts for the modes of binding and pagination in the compendium.

Reporter gene expression in fish following cutaneous infection with pantropic retroviral vectors.

PubMed

Paul, T A; Burns, J C; Shike, H; Getchell, R; Bowser, P R; Whitlock, K E; Casey, J W

2001-06-01

A central issue in gene delivery systems is choosing promoters that will direct defined and sustainable levels of gene expression. Pantropic retroviral vectors provide a means to insert genes into either somatic or germline cells. In this study, we focused on somatic cell infection by evaluating the activity of 3 promoters inserted by vectors into fish cell lines and fish skin using pantropic retroviruses. In bluegill and zebrafish cell lines, the highest levels of luciferase expression were observed from the 5' murine leukemia virus long terminal repeat of the retroviral vector. The Rous sarcoma virus long terminal repeat and cytomegalovirus early promoter, as internal promoters, generated lower levels of luciferase. Luciferase reporter vectors infected zebrafish skin, as measured by the presence of viral DNA, and expressed luciferase. We infected developing walleye dermal sarcomas with retroviral vectors to provide an environment with enhanced cell proliferation, a condition necessary for integration of the provirus into the host genome. We demonstrated a 4-fold to 7-fold increase in luciferase gene expression in tumor tissue over infections in normal walleye skin.

Modelling the role of Tax expression in HTLV-I persistence in vivo.

PubMed

Li, Michael Y; Lim, Aaron G

2011-12-01

Human T-lymphotropic virus type I (HTLV-I) is a persistent human retrovirus characterized by life-long infection and risk of developing HAM/TSP, a progressive neurological and inflammatory disease, and adult T-cell leukemia (ATL). Chronically infected individuals often harbor high proviral loads despite maintaining a persistently activated immune response. Based on a new hypothesis for the persistence of HTLV-I infection, a three-dimensional compartmental model is constructed that describes the dynamic interactions among latently infected target cells, target-cell activation, and immune responses to HTLV-I, with an emphasis on understanding the role of Tax expression in the persistence of HTLV-I.

Processed pseudogenes of human endogenous retroviruses generated by LINEs: their integration, stability, and distribution.

PubMed

Pavlícek, Adam; Paces, Jan; Elleder, Daniel; Hejnar, Jirí

2002-03-01

We report here the presence of numerous processed pseudogenes derived from the W family of endogenous retroviruses in the human genome. These pseudogenes are structurally colinear with the retroviral mRNA followed by a poly(A) tail. Our analysis of insertion sites of HERV-W processed pseudogenes shows a strong preference for the insertion motif of long interspersed nuclear element (LINE) retrotransposons. The genomic distribution, stability during evolution, and frequent truncations at the 5' end resemble those of the pseudogenes generated by LINEs. We therefore suggest that HERV-W processed pseudogenes arose by multiple and independent LINE-mediated retrotransposition of retroviral mRNA. These data document that the majority of HERV-W copies are actually nontranscribed promoterless pseudogenes. The current search for HERV-Ws associated with several human diseases should concentrate on a small subset of transcriptionally competent elements.

Flamenco, a gene controlling the gypsy retrovirus of drosophila melanogaster

DOE Office of Scientific and Technical Information (OSTI.GOV)

Prud`homme, N.; Gans, M.; Masson, M.

1995-02-01

Gypsy is an endogenous retrovirus of Drosophila melanogaster. It is table and does not transpose with detectable frequencies in most Drosophila strains. However, we have characterized unstable strains, known as MG, in which it transposes at high frequency. These stocks contain more copies of gypsy than usual stocks. Transposition results in mutations in several genes such as ovo and cut. They are stable and are due to gypsy insertions. Integrations into the ovo{sup D1} female sterile-dominant mutation result in a null allele of the gene and occurrence of fertile females. This phenomenon, known as the ovo{sup D1} reversion assay, canmore » be used to quantitate gypsy activity. We have shown that the properties of MG strains result from mutation of a host gene that we called flamenco (flam). It has a strict maternal effect on gypsy mobilization: transposition occurs at high frequency only in the germ line of the progeny of females homozygous for mutations of the gene. It is located at position 65.9 (20A1-3) on the X chromosome. The mutant allele present in MG strains is essentially recessive. Flamenco seems to control the infective properties of gypsy. 40 refs., 10 figs., 6 tabs.« less

Lymphoma, Koala Retrovirus Infection and Reproductive Chlamydiosis in a Koala (Phascolarctos cinereus).

PubMed

Fabijan, J; Woolford, L; Lathe, S; Simmons, G; Hemmatzadeh, F; Trott, D J; Speight, N

Koala retrovirus (KoRV) infection, thought to be associated with lymphoid neoplasia, and Chlamydia pecorum-related ocular and urogenital disease are both highly prevalent in eastern Australian koala (Phascolarctos cinereus) populations. However, in South Australian koalas, little is known about KoRV infection and C. pecorum-associated disease. We report the first South Australian case of lymphoma in a KoRV-A-positive female koala also affected by severe reproductive chlamydiosis. The koala was from the Mount Lofty Ranges population and was presented with hindlimb lameness. Clinical examination identified right stifle crepitus, enlarged superficial lymph nodes and paraovarian cysts. Necropsy examination revealed extensive cartilage degeneration and loss over the medial femoral condyle, solid femoral bone marrow, mesenteric and ovarian tumours, paraovarian cysts and purulent metritis. Histopathology confirmed lymphoma in the bone marrow, mesenteric lymph nodes and ovary, with infiltration and parenchymal effacement in the pancreas, adrenal glands and other tissues. Lymphoma, KoRV and chlamydiosis are being investigated further in this population. Copyright © 2017 Elsevier Ltd. All rights reserved.

Inhibition of catecholamine degradation ameliorates while chemical sympathectomy aggravates the severity of acute Friend retrovirus infection in mice.

PubMed

Bloemker, Dominique; Mollerus, Sina; Gibbert, Kathrin; Dittmer, Ulf; Del Rey, Adriana; Schedlowski, Manfred; Engler, Harald

2016-05-01

Several lines of evidence indicate that the sympathetic nervous system (SNS) might be involved in the pathogenesis and progression of retroviral infections. However, experimental data are scarce and findings inconsistent. Here, we investigated the role of the SNS during acute infection with Friend virus (FV), a pathogenic murine retrovirus that causes polyclonal proliferation of erythroid precursor cells and splenomegaly in adult mice. Experimental animals were infected with FV complex, and viral load, spleen weight, and splenic noradrenaline (NA) concentration was analyzed until 25 days post infection. Results show that FV infection caused a massive but transient depletion in splenic NA during the acute phase of the disease. At the peak of the virus-induced splenomegaly, splenic NA concentration was reduced by about 90% compared to naïve uninfected mice. Concurrently, expression of the catecholamine degrading enzymes monoamine oxidase A (MAO-A) and catechol-O-methyltransferase (COMT) was significantly upregulated in immune cells of the spleen. Pharmacological inhibition of MAO-A and COMT by the selective inhibitors clorgyline and 3,5-dinitrocatechol, respectively, efficiently blocked NA degradation and significantly reduced viral load and virus-induced splenomegaly. In contrast, chemical sympathectomy prior to FV inoculation aggravated the acute infection and extended the duration of the disease. Together these findings demonstrate that catecholamine availability at the site of viral replication is an important factor affecting the course of retroviral infections. Copyright © 2016 Elsevier Inc. All rights reserved.

The National Heart, Lung, and Blood Institute retrovirus epidemiology donor studies (Retrovirus Epidemiology Donor Study and Retrovirus Epidemiology Donor Study-II): twenty years of research to advance blood product safety and availability.

PubMed

Kleinman, Steven; King, Melissa R; Busch, Michael P; Murphy, Edward L; Glynn, Simone A

2012-10-01

The Retrovirus Epidemiology Donor Study (REDS), conducted from 1989 to 2001, and the REDS-II, conducted from 2004 to 2012, were National Heart, Lung, and Blood Institute-funded, multicenter programs focused on improving blood safety and availability in the United States. The REDS-II also included international study sites in Brazil and China. The 3 major research domains of REDS/REDS-II have been infectious disease risk evaluation, blood donation availability, and blood donor characterization. Both programs have made significant contributions to transfusion medicine research methodology by the use of mathematical modeling, large-scale donor surveys, innovative methods of repository sample storage, and establishing an infrastructure that responded to potential emerging blood safety threats such as xenotropic murine leukemia virus-related virus. Blood safety studies have included protocols evaluating epidemiologic and/or laboratory aspects of human immunodeficiency virus, human T-lymphotropic virus 1/2, hepatitis C virus, hepatitis B virus, West Nile virus, cytomegalovirus, human herpesvirus 8, parvovirus B19, malaria, Creutzfeldt-Jakob disease, influenza, and Trypanosoma cruzi infections. Other analyses have characterized blood donor demographics, motivations to donate, factors influencing donor return, behavioral risk factors, donors' perception of the blood donation screening process, and aspects of donor deferral. In REDS-II, 2 large-scale blood donor protocols examined iron deficiency in donors and the prevalence of leukocyte antibodies. This review describes the major study results from over 150 peer-reviewed articles published by these 2 REDS programs. In 2011, a new 7-year program, the Recipient Epidemiology and Donor Evaluation Study-III, was launched. The Recipient Epidemiology and Donor Evaluation Study-III expands beyond donor-based research to include studies of blood transfusion recipients in the hospital setting and adds a third country, South Africa

Expression patterns of ERVWE1/Syncytin-1 and other placentally expressed human endogenous retroviruses along the malignant transformation process of hydatidiform moles.

PubMed

Bolze, Pierre-Adrien; Patrier, Sophie; Cheynet, Valérie; Oriol, Guy; Massardier, Jérôme; Hajri, Touria; Guillotte, Michèle; Bossus, Marc; Sanlaville, Damien; Golfier, François; Mallet, François

2016-03-01

Up to 20% of hydatidiform moles are followed by malignant transformation in gestational trophoblastic neoplasia and require chemotherapy. Syncytin-1 is involved in human placental morphogenesis and is also expressed in various cancers. We assessed the predictive value of the expression of Syncytin-1 and its interactants in the malignant transformation process of hydatidiform moles. Syncytin-1 glycoprotein was localized by immunohistochemistry in hydatidiform moles, gestational trophoblastic neoplasia and control placentas. The transcription levels of its locus ERVWE1, its interaction partners (hASCT1, hASCT2, TLR4 and DC-SIGN) and two loci (ERVFRDE1 and ERV3) involved the expression of other placental envelopes were assessed by real-time PCR. Syncytin-1 glycoprotein was expressed in syncytiotrophoblast of hydatidiform moles with an apical enhancement when compared with normal placentas. Moles with further malignant transformation had a higher staining intensity of Syncytin-1 surface unit C-terminus but the transcription level of its locus ERVWE1 was not different from that of moles with further remission and normal placentas. hASCT1 and TLR4, showed lower transcription levels in complete moles when compared to normal placentas. ERVWE1, ERVFRDE1 and ERV3 transcription was down-regulated in hydatidiform moles and gestational trophoblastic neoplasia. Variations of Syncytin-1 protein localization and down-regulation of hASCT1 and TLR4 transcription are likely to reflect altered functions of Syncytin-1 in the premalignant context of complete moles. The reduced transcription in gestational trophoblastic diseases of ERVWE1, ERVFRDE1 and ERV3, which expression during normal pregnancy is differentially regulated by promoter region methylation, suggest a joint dysregulation mechanism in malignant context. Copyright © 2016 Elsevier Ltd. All rights reserved.

An indicator gene to demonstrate intracellular transposition of defective retroviruses.

PubMed Central

Heidmann, T; Heidmann, O; Nicolas, J F

1988-01-01

An indicator gene for detection and quantitation of RNA-mediated transposition was constructed (neoRT). It was inserted into a Moloney murine leukemia provirus (Mo-MLV) deleted for the envelope gene to test for intracellular transposition of defective retroviruses [Mo-MLV(neo)RT]. NeoRT contains the selectable neo gene (which confers resistance to the drug G418), inactivated by a polyadenylylation sequence inserted between the neo promotor and coding sequence. The polyadenylylation sequence is flanked (on the antisense strand of the DNA) by a donor and an acceptor splice site so as to be removed upon passage of the provirus through an RNA intermediate. 3T3 cells transfected with the defective Mo-MLV(neo)RT provirus are sensitive to G418. After trans-complementation with Mo-MLV, viral transcripts confer resistance to G418 upon infection of test cells. In the resistant cells, the polyadenylylation sequence has been removed, as a result in most cases of precise splicing of the intronic domain. Retrotransposition of the defective Mo-MLV(neo)RT provirus was demonstrated by submitting transfected G418-sensitive clones to selection. Between 1 and 10 G418-resistant clones were obtained per 10(7) cells. Several possess additional copies, with evidence for precise removal of the intronic domain. By using target test cells in coculture experiments, extracellular intermediates of retrotransposition could not be detected. Images PMID:2832848

Structural and quantitative expression analyses of HERV gene family in human tissues.

PubMed

Ahn, Kung; Kim, Heui-Soo

2009-08-31

Human endogenous retroviruses (HERVs) have been implicated in the pathogenesis of several human diseases as multi-copy members in the human genome. Their gene expression profiling could provide us with important insights into the pathogenic relationship between HERVs and cancer. In this study, we have evaluated the genomic structure and quantitatively determined the expression patterns in the env gene of a variety of HERV family members located on six specific loci by the RetroTector 10 program, as well as real-time RT-PCR amplification. The env gene transcripts evidenced significant differences in the human tumor/normal adjacent tissues (colon, liver, uterus, lung and testis). As compared to the adjacent normal tissues, high levels of expression were noted in testis tumor tissues for HERV-K, in liver and lung tumor tissues for HERV-R, in liver, lung, and testis tumor tissues for HERV-H, and in colon and liver tumor tissues for HERV-P. These data warrant further studies with larger groups of patients to develop biomarkers for specific human cancers.

Conserved expression of transposon-derived non-coding transcripts in primate stem cells.

PubMed

Ramsay, LeeAnn; Marchetto, Maria C; Caron, Maxime; Chen, Shu-Huang; Busche, Stephan; Kwan, Tony; Pastinen, Tomi; Gage, Fred H; Bourque, Guillaume

2017-02-28

A significant portion of expressed non-coding RNAs in human cells is derived from transposable elements (TEs). Moreover, it has been shown that various long non-coding RNAs (lncRNAs), which come from the human endogenous retrovirus subfamily H (HERVH), are not only expressed but required for pluripotency in human embryonic stem cells (hESCs). To identify additional TE-derived functional non-coding transcripts, we generated RNA-seq data from induced pluripotent stem cells (iPSCs) of four primate species (human, chimpanzee, gorilla, and rhesus) and searched for transcripts whose expression was conserved. We observed that about 30% of TE instances expressed in human iPSCs had orthologous TE instances that were also expressed in chimpanzee and gorilla. Notably, our analysis revealed a number of repeat families with highly conserved expression profiles including HERVH but also MER53, which is known to be the source of a placental-specific family of microRNAs (miRNAs). We also identified a number of repeat families from all classes of TEs, including MLT1-type and Tigger families, that contributed a significant amount of sequence to primate lncRNAs whose expression was conserved. Together, these results describe TE families and TE-derived lncRNAs whose conserved expression patterns can be used to identify what are likely functional TE-derived non-coding transcripts in primate iPSCs.

Low pH inactivation for xenotropic gamma retrovirus in recombinant human TNF-α receptor immunoglobulin G and mechanism of inactivation.

PubMed

Ma, Rong; Cui, Xiaolan

2014-01-01

CHO-derived recombinant proteins for human therapeutic are used commonly. There are noninfectious endogenous retroviruses in CHO cells. Validation study for inactivation process is required. Murine xenotropic gamma retrovirus (X-MulV) is a model virus in validation study. In our previous study, optimum conditions for X-MulV inactivation were sifted. In this study, we performed a further research on low pH inactivation for evaluation of X-MulV clearance in manufacturing of recombinant human TNF-α receptor immunoglobulin G fusion proteins (rhTNF-α) for injection. Cell-based infectivity assay was used for the evaluation of X-MulV clearance. RhTNF-α were spiked with X-MulV and were inactivated at pH 3.60 ∼ 3.90, 25 ± 2 °C, and 0 ∼ 240 min, respectively. Samples incubated at the conditions for 15 ∼ 180 min were not inactivated effectively. For 4 h incubation, log10 reductions were achieved 5.0 log10. Biological activity of rhTNF-α incubated at pH 3.60, 25 °C for 4 h, which was assayed on murine L929 fibroblasts cells, was not affected by low pH. Env gene of X-MulV, which was detected by conventional PCR method for the first time, was not detected after incubation at pH 3.60, and it may be the mechanism of low pH inactivation. Copyright © 2013. Published by Elsevier Ltd.

Retrovirus Integration Database (RID): a public database for retroviral insertion sites into host genomes.

PubMed

Shao, Wei; Shan, Jigui; Kearney, Mary F; Wu, Xiaolin; Maldarelli, Frank; Mellors, John W; Luke, Brian; Coffin, John M; Hughes, Stephen H

2016-07-04

The NCI Retrovirus Integration Database is a MySql-based relational database created for storing and retrieving comprehensive information about retroviral integration sites, primarily, but not exclusively, HIV-1. The database is accessible to the public for submission or extraction of data originating from experiments aimed at collecting information related to retroviral integration sites including: the site of integration into the host genome, the virus family and subtype, the origin of the sample, gene exons/introns associated with integration, and proviral orientation. Information about the references from which the data were collected is also stored in the database. Tools are built into the website that can be used to map the integration sites to UCSC genome browser, to plot the integration site patterns on a chromosome, and to display provirus LTRs in their inserted genome sequence. The website is robust, user friendly, and allows users to query the database and analyze the data dynamically. https://rid.ncifcrf.gov ; or http://home.ncifcrf.gov/hivdrp/resources.htm .

Contribution of type W human endogenous retroviruses to the human genome: characterization of HERV-W proviral insertions and processed pseudogenes.

PubMed

Grandi, Nicole; Cadeddu, Marta; Blomberg, Jonas; Tramontano, Enzo

2016-09-09

Human endogenous retroviruses (HERVs) are ancient sequences integrated in the germ line cells and vertically transmitted through the offspring constituting about 8 % of our genome. In time, HERVs accumulated mutations that compromised their coding capacity. A prominent exception is HERV-W locus 7q21.2, producing a functional Env protein (Syncytin-1) coopted for placental syncytiotrophoblast formation. While expression of HERV-W sequences has been investigated for their correlation to disease, an exhaustive description of the group composition and characteristics is still not available and current HERV-W group information derive from studies published a few years ago that, of course, used the rough assemblies of the human genome available at that time. This hampers the comparison and correlation with current human genome assemblies. In the present work we identified and described in detail the distribution and genetic composition of 213 HERV-W elements. The bioinformatics analysis led to the characterization of several previously unreported features and provided a phylogenetic classification of two main subgroups with different age and structural characteristics. New facts on HERV-W genomic context of insertion and co-localization with sequences putatively involved in disease development are also reported. The present work is a detailed overview of the HERV-W contribution to the human genome and provides a robust genetic background useful to clarify HERV-W role in pathologies with poorly understood etiology, representing, to our knowledge, the most complete and exhaustive HERV-W dataset up to date.

HERVs Expression in Autism Spectrum Disorders

PubMed Central

Balestrieri, Emanuela; Arpino, Carla; Matteucci, Claudia; Sorrentino, Roberta; Pica, Francesca; Alessandrelli, Riccardo; Coniglio, Antonella; Curatolo, Paolo; Rezza, Giovanni; Macciardi, Fabio; Garaci, Enrico; Gaudi, Simona; Sinibaldi-Vallebona, Paola

2012-01-01

Background Autistic Spectrum Disorder (ASD) is a heterogeneous neurodevelopmental disorder, resulting from complex interactions among genetic, genomic and environmental factors. Here we have studied the expression of Human Endogenous Retroviruses (HERVs), non-coding DNA elements with potential regulatory functions, and have tested their possible implication in autism. Methods The presence of retroviral mRNAs from four HERV families (E, H, K and W), widely implicated in complex diseases, was evaluated in peripheral blood mononuclear cells (PBMCs) from ASD patients and healthy controls (HCs) by qualitative RT-PCR. We also analyzed the expression of the env sequence from HERV-H, HERV-W and HERV-K families in PBMCs at the time of sampling and after stimulation in culture, in both ASD and HC groups, by quantitative Real-time PCR. Differences between groups were evaluated using statistical methods. Results The percentage of HERV-H and HERV-W positive samples was higher among ASD patients compared to HCs, while HERV-K was similarly represented and HERV-E virtually absent in both groups. The quantitative evaluation shows that HERV-H and HERV-W are differentially expressed in the two groups, with HERV-H being more abundantly expressed and, conversely, HERV-W, having lower abundance, in PBMCs from ASDs compared to healthy controls. PMBCs from ASDs also showed an increased potential to up-regulate HERV-H expression upon stimulation in culture, unlike HCs. Furthermore we report a negative correlation between expression levels of HERV-H and age among ASD patients and a statistically significant higher expression in ASD patients with Severe score in Communication and Motor Psychoeducational Profile-3. Conclusions Specific HERV families have a distinctive expression profile in ASD patients compared to HCs. We propose that HERV-H expression be explored in larger samples of individuals with autism spectrum in order to determine its utility as a novel biological trait of this

Functional Dissection of the PE Domain Responsible for Translocation of PE_PGRS33 across the Mycobacterial Cell Wall

PubMed Central

Cascioferro, Alessandro; Donà, Valentina; Delogu, Giovanni; Palù, Giorgio; Bitter, Wilbert; Manganelli, Riccardo

2011-01-01

PE are peculiar exported mycobacterial proteins over-represented in pathogenic mycobacterial species. They are characterized by an N-terminal domain of about 110 amino acids (PE domain) which has been demonstrated to be responsible for their export and localization. In this paper, we characterize the PE domain of PE_PGRS33 (PERv1818c), one of the best characterized PE proteins. We constructed several mutated proteins in which portions of the PE domain were deleted or subjected to defined mutations. These proteins were expressed in different mycobacterial species and their localization was characterized. We confirmed that the PE domain is essential for PE_PGRS33 surface localization, and demonstrated that a PE domain lacking its first 30 amino acids loses its function. However, single amino acid substitutions in two regions extremely well conserved within the N-terminal domain of all PE proteins had some effect on the stability of PE_PGRS33, but not on its localization. Using Mycobacterium marinum we could show that the type VII secretion system ESX-5 is essential for PE_PGRS33 export. Moreover, in M. marinum, but not in Mycobacterium bovis BCG and in Mycobacterium tuberculosis, the PE domain of PE_PGRS33 is processed and secreted into the culture medium when expressed in the absence of the PGRS domain. Finally, using chimeric proteins in which different portions of the PERv1818c domain were fused to the N-terminus of the green fluorescent protein, we could hypothesize that the first 30 amino acids of the PE domain contain a sequence that allows protein translocation. PMID:22110736

Identification of Transposable Elements Contributing to Tissue-Specific Expression of Long Non-Coding RNAs

PubMed Central

Chishima, Takafumi; Iwakiri, Junichi

2018-01-01

It has been recently suggested that transposable elements (TEs) are re-used as functional elements of long non-coding RNAs (lncRNAs). This is supported by some examples such as the human endogenous retrovirus subfamily H (HERVH) elements contained within lncRNAs and expressed specifically in human embryonic stem cells (hESCs), as required to maintain hESC identity. There are at least two unanswered questions about all lncRNAs. How many TEs are re-used within lncRNAs? Are there any other TEs that affect tissue specificity of lncRNA expression? To answer these questions, we comprehensively identify TEs that are significantly related to tissue-specific expression levels of lncRNAs. We downloaded lncRNA expression data corresponding to normal human tissue from the Expression Atlas and transformed the data into tissue specificity estimates. Then, Fisher’s exact tests were performed to verify whether the presence or absence of TE-derived sequences influences the tissue specificity of lncRNA expression. Many TE–tissue pairs associated with tissue-specific expression of lncRNAs were detected, indicating that multiple TE families can be re-used as functional domains or regulatory sequences of lncRNAs. In particular, we found that the antisense promoter region of L1PA2, a LINE-1 subfamily, appears to act as a promoter for lncRNAs with placenta-specific expression. PMID:29315213

Computer simulation of a cellular automata model for the immune response in a retrovirus system

NASA Astrophysics Data System (ADS)

Pandey, R. B.

1989-02-01

Immune response in a retrovirus system is modeled by a network of three binary cell elements to take into account some of the main functional features of T4 cells, T8 cells, and viruses. Two different intercell interactions are introduced, one of which leads to three fixed points while the other yields bistable fixed points oscillating between a healthy state and a sick state in a mean field treatment. Evolution of these cells is studied for quenched and annealed random interactions on a simple cubic lattice with a nearest neighbor interaction using inhomogenous cellular automata. Populations of T4 cells and viral cells oscillate together with damping (with constant amplitude) for annealed (quenched) interaction on increasing the value of mixing probability B from zero to a characteristic value B ca ( B cq). For higher B, the average number of T4 cells increases while that of the viral infected cells decreases monotonically on increasing B, suggesting a phase transition at B ca ( B cq).

The MHC-II transactivator CIITA, a restriction factor against oncogenic HTLV-1 and HTLV-2 retroviruses: similarities and differences in the inhibition of Tax-1 and Tax-2 viral transactivators

PubMed Central

Forlani, Greta; Abdallah, Rawan; Accolla, Roberto S.; Tosi, Giovanna

2013-01-01

The activation of CD4+ T helper cells is strictly dependent on the presentation of antigenic peptides by MHC class II (MHC-II) molecules. MHC-II expression is primarily regulated at the transcriptional level by the AIR-1 gene product CIITA (class II transactivator). Thus, CIITA plays a pivotal role in the triggering of the adaptive immune response against pathogens. Besides this well known function, we recently found that CIITA acts as an endogenous restriction factor against HTLV-1 (human T cell lymphotropic virus type 1) and HTLV-2 oncogenic retroviruses by targeting their viral transactivators Tax-1 and Tax-2, respectively. Here we review our findings on CIITA-mediated inhibition of viral replication and discuss similarities and differences in the molecular mechanisms by which CIITA specifically counteracts the function of Tax-1 and Tax-2 molecules. The dual function of CIITA as a key regulator of adaptive and intrinsic immunity represents a rather unique example of adaptation of host-derived factors against pathogen infections during evolution. PMID:23986750

Activation of MSRV-Type Endogenous Retroviruses during Infectious Mononucleosis and Epstein-Barr Virus Latency: The Missing Link with Multiple Sclerosis?

PubMed Central

Mameli, Giuseppe; Madeddu, Giordano; Mei, Alessandra; Uleri, Elena; Poddighe, Luciana; Delogu, Lucia G.; Maida, Ivana; Babudieri, Sergio; Serra, Caterina; Manetti, Roberto; Mura, Maria S.; Dolei, Antonina

2013-01-01

The etiology of multiple sclerosis (MS) is still unclear. The immuno-pathogenic phenomena leading to neurodegeneration are thought to be triggered by environmental (viral?) factors operating on predisposing genetic backgrounds. Among the proposed co-factors are the Epstein Barr virus (EBV), and the potentially neuropathogenic HERV-W/MSRV/Syncytin-1 endogenous retroviruses. The ascertained links between EBV and MS are history of late primary infection, possibly leading to infectious mononucleosis (IM), and high titers of pre-onset IgG against EBV nuclear antigens (anti-EBNA IgG). During MS, there is no evidence of MS-specific EBV expression, while a continuous expression of HERV-Ws occurs, paralleling disease behaviour. We found repeatedly extracellular HERV-W/MSRV and MSRV-specific mRNA sequences in MS patients (in blood, spinal fluid, and brain samples), and MRSV presence/load strikingly paralleled MS stages and active/remission phases. Aim of the study was to verify whether HERV-W might be activated in vivo, in hospitalized young adults with IM symptoms, that were analyzed with respect to expression of HERV-W/MSRV transcripts and proteins. Healthy controls were either EBV-negative or latently EBV-infected with/without high titers of anti-EBNA-1 IgG. The results show that activation of HERV-W/MSRV occurs in blood mononuclear cells of IM patients (2Log10 increase of MSRV-type env mRNA accumulation with respect to EBV-negative controls). When healthy controls are stratified for previous EBV infection (high and low, or no anti-EBNA-1 IgG titers), a direct correlation occurs with MSRV mRNA accumulation. Flow cytometry data show increased percentages of cells exposing surface HERV-Wenv protein, that occur differently in specific cell subsets, and in acute disease and past infection. Thus, the data indicate that the two main links between EBV and MS (IM and high anti-EBNA-1-IgG titers) are paralleled by activation of the potentially neuropathogenic HERV-W/MSRV. These

Activation of MSRV-type endogenous retroviruses during infectious mononucleosis and Epstein-Barr virus latency: the missing link with multiple sclerosis?

PubMed

Mameli, Giuseppe; Madeddu, Giordano; Mei, Alessandra; Uleri, Elena; Poddighe, Luciana; Delogu, Lucia G; Maida, Ivana; Babudieri, Sergio; Serra, Caterina; Manetti, Roberto; Mura, Maria S; Dolei, Antonina

2013-01-01

The etiology of multiple sclerosis (MS) is still unclear. The immuno-pathogenic phenomena leading to neurodegeneration are thought to be triggered by environmental (viral?) factors operating on predisposing genetic backgrounds. Among the proposed co-factors are the Epstein Barr virus (EBV), and the potentially neuropathogenic HERV-W/MSRV/Syncytin-1 endogenous retroviruses. The ascertained links between EBV and MS are history of late primary infection, possibly leading to infectious mononucleosis (IM), and high titers of pre-onset IgG against EBV nuclear antigens (anti-EBNA IgG). During MS, there is no evidence of MS-specific EBV expression, while a continuous expression of HERV-Ws occurs, paralleling disease behaviour. We found repeatedly extracellular HERV-W/MSRV and MSRV-specific mRNA sequences in MS patients (in blood, spinal fluid, and brain samples), and MRSV presence/load strikingly paralleled MS stages and active/remission phases. Aim of the study was to verify whether HERV-W might be activated in vivo, in hospitalized young adults with IM symptoms, that were analyzed with respect to expression of HERV-W/MSRV transcripts and proteins. Healthy controls were either EBV-negative or latently EBV-infected with/without high titers of anti-EBNA-1 IgG. The results show that activation of HERV-W/MSRV occurs in blood mononuclear cells of IM patients (2Log10 increase of MSRV-type env mRNA accumulation with respect to EBV-negative controls). When healthy controls are stratified for previous EBV infection (high and low, or no anti-EBNA-1 IgG titers), a direct correlation occurs with MSRV mRNA accumulation. Flow cytometry data show increased percentages of cells exposing surface HERV-Wenv protein, that occur differently in specific cell subsets, and in acute disease and past infection. Thus, the data indicate that the two main links between EBV and MS (IM and high anti-EBNA-1-IgG titers) are paralleled by activation of the potentially neuropathogenic HERV-W/MSRV. These

HEC-1 Flood Hydrograph Package. Users Manual

DTIC Science & Technology

1987-03-01

98 Perv Catchment 200 .02 .35 80 85 SUB7 Ip Catchment 100 .06 .24 15 .96 98 Perv Catchnmnt 190 .03 .35 85 85 TABLE 12.2b CWNEL DATA (Test 2)(RK RECORD...02 .55 TRAP 0 1 AIN CHANNEL 14000 .005 .09 - TRAP 2 2 no SUB7 COLLECTOR CHANNEL 2100 .024 .02 .74 TRAP 0 1 MAIN CHANNEL 7000 .011 .05 TRAP 50 3 yes...35 80 125 RK 2200 .011 .02 .55 TRAP 0 126 RK 14000 .005 .09 TRAP 2 2 127 9K SUB7 128 KM COMBINE RUNOFF FROM SUB4, SUDS, AND SUB6 129 HC 3 130 1IK

The NHLBI Retrovirus Epidemiology Donor Studies (REDS and REDS-II): Twenty years of research to advance blood product safety and availability

PubMed Central

Kleinman, Steven; King, Melissa R; Busch, Michael P; Murphy, Edward L; Glynn, Simone A.

2012-01-01

The Retrovirus Epidemiology Donor Study (REDS), conducted from 1989–2001, and the Retrovirus Epidemiology Donor Study-II (REDS-II), conducted from 2004–2012, were National Heart Lung and Blood Institute (NHLBI) funded multicenter programs focused on improving blood safety and availability in the United States. REDS-II also included international study sites in Brazil and China. The three major research domains of REDS/REDS-II have been infectious disease risk evaluation, blood donation availability, and blood donor characterization. Both programs have made significant contributions to transfusion medicine research methodology by the use of mathematical modeling, large-scale donor surveys, innovative methods of repository sample storage, and establishing an infrastructure that responded to potential emerging blood safety threats such as XMRV. Blood safety studies have included protocols evaluating epidemiologic and/or laboratory aspects of HIV, HTLV I/II, HCV, HBV, WNV, CMV, HHV-8, B19V, malaria, CJD, influenza, and T. cruzi infections. Other analyses have characterized: blood donor demographics, motivations to donate, factors influencing donor return, behavioral risk factors, donors’ perception of the blood donation screening process, and aspects of donor deferral. In REDS-II, two large-scale blood donor protocols examined iron deficiency in donors and the prevalence of leukocyte antibodies. This review describes the major study results from over 150 peer-reviewed articles published by these two REDS programs. In 2011, a new seven year program, the Recipient Epidemiology and Donor Evaluation Study-III (REDS-III), was launched. REDS-III expands beyond donor-based research to include studies of blood transfusion recipients in the hospital setting, and adds a third country, South Africa, to the international program. PMID:22633182

Human endogenous retrovirus-FRD envelope protein (syncytin 2) expression in normal and trisomy 21-affected placenta.

PubMed

Malassiné, André; Frendo, Jean-Louis; Blaise, Sandra; Handschuh, Karen; Gerbaud, Pascale; Tsatsaris, Vassilis; Heidmann, Thierry; Evain-Brion, Danièle

2008-01-23

Human trophoblast expresses two fusogenic retroviral envelope proteins, the widely studied syncytin 1, encoded by HERV-W and the recently characterized syncytin 2 encoded by HERV-FRD. Here we studied syncytin 2 in normal and Trisomy 21-affected placenta associated with abnormal trophoblast differentiation. Syncytin 2 immunolocalization was restricted throughout normal pregnancy to some villous cytotrophoblastic cells (CT). During the second trimester of pregnancy, syncytin 2 was immunolocalized in some cuboidal CT in T21 placentas, whereas in normal placentas it was observed in flat CT, extending into their cytoplasmic processes. In vitro, CT isolated from normal placenta fuse and differentiate into syncytiotrophoblast. At the same time, syncytin 2 transcript levels decreased significantly with syncytiotrophoblast formation. In contrast, CT isolated from T21-affected placentas fused and differentiated poorly and no variation in syncytin 2 transcript levels was observed. Syncytin 2 expression illustrates the abnormal trophoblast differentiation observed in placenta of fetal T21-affected pregnancies.

Human endogenous retrovirus-FRD envelope protein (syncytin 2) expression in normal and trisomy 21-affected placenta

PubMed Central

Malassiné, André; Frendo, Jean-Louis; Blaise, Sandra; Handschuh, Karen; Gerbaud, Pascale; Tsatsaris, Vassilis; Heidmann, Thierry; Evain-Brion, Danièle

2008-01-01

Human trophoblast expresses two fusogenic retroviral envelope proteins, the widely studied syncytin 1, encoded by HERV-W and the recently characterized syncytin 2 encoded by HERV-FRD. Here we studied syncytin 2 in normal and Trisomy 21-affected placenta associated with abnormal trophoblast differentiation. Syncytin 2 immunolocalization was restricted throughout normal pregnancy to some villous cytotrophoblastic cells (CT). During the second trimester of pregnancy, syncytin 2 was immunolocalized in some cuboidal CT in T21 placentas, whereas in normal placentas it was observed in flat CT, extending into their cytoplasmic processes. In vitro, CT isolated from normal placenta fuse and differentiate into syncytiotrophoblast. At the same time, syncytin 2 transcript levels decreased significantly with syncytiotrophoblast formation. In contrast, CT isolated from T21-affected placentas fused and differentiated poorly and no variation in syncytin 2 transcript levels was observed. Syncytin 2 expression illustrates the abnormal trophoblast differentiation observed in placenta of fetal T21-affected pregnancies. PMID:18215254

Differential Expression of HERV-K (HML-2) Proviruses in Cells and Virions of the Teratocarcinoma Cell Line Tera-1

PubMed Central

Bhardwaj, Neeru; Montesion, Meagan; Roy, Farrah; Coffin, John M.

2015-01-01

Human endogenous retrovirus (HERV-K (HML-2)) proviruses are among the few endogenous retroviral elements in the human genome that retain coding sequence. HML-2 expression has been widely associated with human disease states, including different types of cancers as well as with HIV-1 infection. Understanding of the potential impact of this expression requires that it be annotated at the proviral level. Here, we utilized the high throughput capabilities of next-generation sequencing to profile HML-2 expression at the level of individual proviruses and secreted virions in the teratocarcinoma cell line Tera-1. We identified well-defined expression patterns, with transcripts emanating primarily from two proviruses located on chromosome 22, only one of which was efficiently packaged. Interestingly, there was a preference for transcripts of recently integrated proviruses, over those from other highly expressed but older elements, to be packaged into virions. We also assessed the promoter competence of the 5’ long terminal repeats (LTRs) of expressed proviruses via a luciferase assay following transfection of Tera-1 cells. Consistent with the RNASeq results, we found that the activity of most LTRs corresponded to their transcript levels. PMID:25746218

Stable expression of hepatitis delta virus antigen in a eukaryotic cell line.

PubMed

Macnaughton, T B; Gowans, E J; Reinboth, B; Jilbert, A R; Burrell, C J

1990-06-01

The gene encoding the hepatitis delta virus structural antigen (HDAg) was linked to a neomycin resistance gene in a retrovirus expression vector, and human HepG2 cells were transfected with the recombinant plasmid. A stable cell line was cloned that expressed HDAg in the nuclei of 100% of cells, in a pattern indicating a close relationship with cell nucleoli. Analysis of partially purified recombinant HDAg by HPLC showed an Mr in the range of 7 x 10(5) to 2 x 10(6), which appeared to contain conformation-dependent epitopes, whereas the density of the antigen was 1.19 g/ml by equilibrium centrifugation in caesium chloride, and in rate zonal centrifugation it sedimented with a value of 50S, close to that of particulate hepatitis B virus surface antigen. Immunoblotting demonstrated a single polypeptide with an Mr of 24K which corresponded to the smaller of the two HDAg-specific polypeptides present in infected sera. The recombinant HDAg polypeptide was shown to be a RNA-binding protein with specificity for both genomic and antigenomic species of hepatitis delta virus RNA.

Ars insulator identified in sea urchin possesses an activity to ensure the transgene expression in mouse cells.

PubMed

Tajima, Shoji; Shinohara, Keiko; Fukumoto, Maiko; Zaitsu, Reiko; Miyagawa, Junichi; Hino, Shinjiro; Fan, Jun; Akasaka, Koji; Matsuoka, Masao

2006-04-01

Sea urchin arylsulfatase (Ars) gene locus has features of an insulator, i.e., blocking of enhancer and promoter interaction, and protection of a transgene against positional effects [Akasaka et al. (1999) Cell. Mol. Biol. 45, 555-565]. To examine the effect of Ars insulator on long-term expression of a transgene, the insulator was inserted into LTR of retrovirus vector harboring hrGFP gene as a reporter, and then introduced into mouse myoblast cells. The isolated clones transduced with the reporter gene with or without Ars insulator were cultured for more than 20 wk in the absence of a selection reagent, and the expression of hrGFP was periodically determined. Expression of hrGFP in four clones transduced with the reporter gene without Ars insulator was completely silenced after 20 wk of culture. On the other hand, hrGFP was expressed in all clones with Ars insulator inserted in one of the two different orientations. Histone H3 deacetylation and DNA methylation of the 5'LTR promoter region, signs for heterochromatin and silencing, were suppressed in the clones that were expressing hrGFP. Ars insulator is effective in maintaining a transgene in mouse cells in an orientation-dependent manner, and will be a useful tool to ensure stable expression of a transgene.

Computer simulation of a cellular automata model for the immune response in a retrovirus system

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pandey, R.B.

1989-02-01

Immune response in a retrovirus system is modeled by a network of three binary cell elements to take into account some of the main functional features of T4 cells, T8 cells, and viruses. Two different intercell interactions are introduced, one of which leads to three fixed points while the other yields bistable fixed points oscillating between a healthy state and a sick state in a mean field treatment. Evolution of these cells is studied for quenched and annealed random interactions on a simple cubic lattice with a nearest neighbor interaction using inhomogenous cellular automata. Populations of T4 cells and viralmore » cells oscillate together with damping (with constant amplitude) for annealed (quenched) interaction on increasing the value of mixing probability B from zero to a characteristic value B/sub ca/ (B/sub cq/). For higher B, the average number of T4 cells increases while that of the viral infected cells decreases monotonically on increasing B, suggesting a phase transition at B/sub ca/ (B/sub cq/).« less

Structural and Biochemical Studies of ALIX/AlP1 and Its Role in Retrovirus Budding

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fisher,R.; Chung, H.; Zhai, Q.

2007-01-01

ALIX/AIP1 functions in enveloped virus budding, endosomal protein sorting, and many other cellular processes. Retroviruses, including HIV-1, SIV, and EIAV, bind and recruit ALIX through YPXnL late-domain motifs (X = any residue; n = 1-3). Crystal structures reveal that human ALIX is composed of an N-terminal Bro1 domain and a central domain that is composed of two extended three-helix bundles that form elongated arms that fold back into a 'V.'. The structures also reveal conformational flexibility in the arms that suggests that the V domain may act as a flexible hinge in response to ligand binding. YPXnL late domains bindmore » in a conserved hydrophobic pocket on the second arm near the apex of the V, whereas CHMP4/ESCRT-III proteins bind a conserved hydrophobic patch on the Bro1 domain, and both interactions are required for virus budding. ALIX therefore serves as a flexible, extended scaffold that connects retroviral Gag proteins to ESCRT-III and other cellular-budding machinery.« less

Koala retrovirus genotyping analyses reveal a low prevalence of KoRV-A in Victorian koalas and an association with clinical disease.

PubMed

Legione, Alistair R; Patterson, Jade L S; Whiteley, Pam; Firestone, Simon M; Curnick, Megan; Bodley, Kate; Lynch, Michael; Gilkerson, James R; Sansom, Fiona M; Devlin, Joanne M

2017-02-01

Koala retrovirus (KoRV) is undergoing endogenization into the genome of koalas in Australia, providing an opportunity to assess the effect of retrovirus infection on the health of a population. The prevalence of KoRV in north-eastern Australia (Queensland and New South Wales) is 100 %, whereas previous preliminary investigations in south-eastern Australia (Victoria) suggested KoRV is present at a lower prevalence, although the values have varied widely. Here, we describe a large study of free-ranging koalas in Victoria to estimate the prevalence of KoRV and assess the clinical significance of KoRV infection in wild koalas. Blood or spleen samples from 648 koalas where tested for KoRV provirus, and subsequently genotyped, using PCRs to detect the pol and env genes respectively. Clinical data was also recorded where possible and analysed in comparison to infection status. The prevalence of KoRV was 24.7 % (160/648). KoRV-A was detected in 141/160 cases, but KoRV-B, a genotype associated with neoplasia in captive koalas, was not detected. The genotype in 19 cases could not be determined. Genomic differences between KoRV in Victoria and type strains may have impacted genotyping. Factors associated with KoRV infection, based on multivariable analysis, were low body condition score, region sampled, and 'wet bottom' (a staining of the fur around the rump associated with chronic urinary incontinence). Koalas with wet bottom were nearly twice as likely to have KoRV provirus detected than those without wet bottom (odds ratio=1.90, 95 % confidence interval 1.21, 2.98). Our findings have important implications for the conservation of this iconic species, particularly regarding translocation potential of Victorian koalas.

G protein-coupled receptor 30 down-regulates cofactor expression and interferes with the transcriptional activity of glucocorticoid.

PubMed

Ylikomi, Timo; Vienonen, Annika; Ahola, Tytti M

2004-11-01

G protein-coupled receptor 30 (GPR30) has previously been described to be important in steroid-mediated growth and to inhibit cell proliferation. Here we investigated whether the effect of GPR30 on cell growth is dependent on steroid hormone receptors. We stably introduced GPR30 in immortalized normal mammary epithelial (HME) cells using retroviruses for gene delivery. GPR30 inhibited the growth and proliferation of the cells. They expressed glucocorticoid receptor, but not estrogen or progesterone receptor. GPR30 down-regulated the expression of cofactor transcription intermediary factor 2 (TIF2) analyzed using quantitative RT-PCR analysis, and also diminished the expression of TIF2 at protein level analyzed by Western blotting using nuclear extracts from mammary epithelial cells. When HME cells were transiently transfected with the glucocorticoid response element MMTV-luc reporter plasmid, stable expression of GPR30 resulted in the abolition of ligand-induced transactivation of the promoter. In COS cells, transient transfection of GPR30 with glucocorticoid receptor alpha resulted in an abrogation of the MMTV-luc and GRE-luc reporter activities induced by dexamethasone. The results suggest a novel mechanism by which membrane-initiated signaling interferes with steroid signaling.

Expression of young HERV-H loci in the course of colorectal carcinoma and correlation with molecular subtypes

PubMed Central

Naville, Magali; Bressan, Cédric; Hühns, Maja; Gock, Michael; Kühn, Florian; Volff, Jean-Nicolas; Trillet-Lenoir, Véronique

2015-01-01

Background Expression of the human endogenous retrovirus (HERV)-H family has been associated with colorectal carcinomas (CRC), yet no individual HERV-H locus expression has been thoroughly correlated with clinical data. Here, we characterized HERV-H reactivations in clinical CRC samples by integrating expression profiles, molecular patterns and clinical data. Expression of relevant HERV-H sequences was analyzed by qRT-PCR on two well-defined clinical cohorts (n = 139 pairs of tumor and adjacent normal colon tissue) including samples from adenomas (n = 21) and liver metastases (n = 16). Correlations with clinical and molecular data were assessed. Results CRC specific HERV-H sequences were validated and found expressed throughout CRC disease progression. Correlations between HERV-H expression and lymph node invasion of tumor cells (p = 0.0006) as well as microsatellite instable tumors (p < 0.0001) were established. No association with regard to age, tumor localization, grading or common mutations became apparent. Interestingly, CRC expressed elements belonged to specific young HERV-H subfamilies and their 5′ LTR often presented active histone marks. Conclusion These results suggest a functional role of HERV-H sequences in colorectal carcinogenesis. The pronounced connection with microsatellite instability warrants a more detailed investigation. Thus, HERV-H sequences in addition to tumor specific mutations may represent clinically relevant, truly CRC specific markers for diagnostic, prognostic and therapeutic purposes. PMID:26517682

Expression of young HERV-H loci in the course of colorectal carcinoma and correlation with molecular subtypes.

PubMed

Pérot, Philippe; Mullins, Christina Susanne; Naville, Magali; Bressan, Cédric; Hühns, Maja; Gock, Michael; Kühn, Florian; Volff, Jean-Nicolas; Trillet-Lenoir, Véronique; Linnebacher, Michael; Mallet, François

2015-11-24

Expression of the human endogenous retrovirus (HERV)-H family has been associated with colorectal carcinomas (CRC), yet no individual HERV-H locus expression has been thoroughly correlated with clinical data.Here, we characterized HERV-H reactivations in clinical CRC samples by integrating expression profiles, molecular patterns and clinical data. Expression of relevant HERV-H sequences was analyzed by qRT-PCR on two well-defined clinical cohorts (n = 139 pairs of tumor and adjacent normal colon tissue) including samples from adenomas (n = 21) and liver metastases (n = 16). Correlations with clinical and molecular data were assessed. CRC specific HERV-H sequences were validated and found expressed throughout CRC disease progression. Correlations between HERV-H expression and lymph node invasion of tumor cells (p = 0.0006) as well as microsatellite instable tumors (p < 0.0001) were established. No association with regard to age, tumor localization, grading or common mutations became apparent. Interestingly, CRC expressed elements belonged to specific young HERV-H subfamilies and their 5' LTR often presented active histone marks. These results suggest a functional role of HERV-H sequences in colorectal carcinogenesis. The pronounced connection with microsatellite instability warrants a more detailed investigation. Thus, HERV-H sequences in addition to tumor specific mutations may represent clinically relevant, truly CRC specific markers for diagnostic, prognostic and therapeutic purposes.

Pathogenic and host range determinants of the feline aplastic anemia retrovirus.

PubMed Central

Riedel, N; Hoover, E A; Dornsife, R E; Mullins, J I

1988-01-01

Feline leukemia virus (FeLV) C-Sarma (or FSC) is a prototype of subgroup C FeLVs, which induce fatal aplastic anemia in outbred specific-pathogen-free (SPF) cats. FeLV C isolates also possess an extended host range in vitro, including an ability, unique among FeLVs, to replicate in guinea pig cells. To identify the viral determinants responsible for the pathogenicity and host range of FSC we constructed a series of proviral DNAs by exchanging gene fragments between FSC and FeLV-61E (or F6A), the latter of which is minimally pathogenic and whose host range in vitro is restricted to feline cells. Transfer of an 886-base-pair (bp) fragment of FSC, encompassing the codons for 73 amino acids at the 3' end of pol (the integrase/endonuclease gene) and the codons for 241 amino acids of the N-terminal portion of env [the extracellular glycoprotein (gp70) gene], into the F6A genome was sufficient to confer onto chimeric viruses the ability to induce fatal aplastic anemia in SPF cats. In contrast, no chimera lacking this sequence induced disease. When assayed in vitro, all chimeric viruses containing the 886-bp fragment of FSC acquired the ability to replicate in heterologous cells, including dog and guinea pig cells. Thus, the pathogenic and the host range determinants of the feline aplastic anemia retrovirus colocalize to a 3' pol-5' env region of the FSC genome and likely reside within a region encoding 241 amino acid residues of the N terminus of the extracellular glycoprotein. Images PMID:2833751

The LINE-1 DNA sequences in four mammalian orders predict proteins that conserve homologies to retrovirus proteins.

PubMed Central

Fanning, T; Singer, M

1987-01-01

Recent work suggests that one or more members of the highly repeated LINE-1 (L1) DNA family found in all mammals may encode one or more proteins. Here we report the sequence of a portion of an L1 cloned from the domestic cat (Felis catus). These data permit comparison of the L1 sequences in four mammalian orders (Carnivore, Lagomorph, Rodent and Primate) and the comparison supports the suggested coding potential. In two separate, noncontiguous regions in the carboxy terminal half of the proteins predicted from the DNA sequences, there are several strongly conserved segments. In one region, these share homology with known or suspected reverse transcriptases, as described by others in rodents and primates. In the second region, closer to the carboxy terminus, the strongly conserved segments are over 90% homologous among the four orders. One of the latter segments is cysteine rich and resembles the putative metal binding domains of nucleic acid binding proteins, including those of TFIIIA and retroviruses. PMID:3562227

Elevated HERV-K Expression in Soft Tissue Sarcoma Is Associated with Worsened Relapse-Free Survival.

PubMed

Giebler, Maria; Staege, Martin S; Blauschmidt, Sindy; Ohm, Lea I; Kraus, Matthias; Würl, Peter; Taubert, Helge; Greither, Thomas

2018-01-01

A wide variety of endogenous retroviral sequences has been demonstrated in the human genome so far, divided into several different families according to the sequence homology to viral strains. While increased expression of human endogenous retrovirus (HERV) elements has already been linked to unfavorable prognosis in hepatocellular carcinoma, breast cancer, and ovarian carcinoma yet less is known about the impact of the expression of different HERV elements on sarcomagenesis in general as well as the outcome of soft tissue sarcoma (STS) patients. Therefore, in this study the association between expression of HERV-K and HERV-F and the clinicopathological characteristics in a cohort of STSs as well as the patients' prognosis was evaluated. HERV-K and HERV-F expression was assessed by quantitative real-time PCR in 120 patient specimens. HERV-K and HERV-F expression was significantly correlated ( r S = 0.5; p = 6.4 × 10 -9 ; Spearman's rank bivariate correlation). Also, tumor diameter exhibited a significant negative association to HERV-K and HERV-F expression. Levels of several hypoxia-related RNAs like HIF-1α and miR-210 showed a significant positive correlation with both HERV-K and HERV-F expression. Although in survival analyses no impact of HERV expression on disease-specific survival could be detected, patients with elevated HERV-K expression had a significantly shorter relapse-free survival ( p = 0.014, log-rank analysis). In conclusion, we provide evidence for the first time that the increased expression of HERV-K in tumors is associated with STS patients' prognosis.

Fragments of Target Cells are Internalized into Retroviral Envelope Protein-Expressing Cells during Cell-Cell Fusion by Endocytosis

PubMed Central

Izumida, Mai; Kamiyama, Haruka; Suematsu, Takashi; Honda, Eri; Koizumi, Yosuke; Yasui, Kiyoshi; Hayashi, Hideki; Ariyoshi, Koya; Kubo, Yoshinao

2016-01-01

Retroviruses enter into host cells by fusion between viral and host cell membranes. Retroviral envelope glycoprotein (Env) induces the membrane fusion, and also mediates cell-cell fusion. There are two types of cell-cell fusions induced by the Env protein. Fusion-from-within is induced by fusion between viral fusogenic Env protein-expressing cells and susceptible cells, and virions induce fusion-from-without by fusion between adjacent cells. Although entry of ecotropic murine leukemia virus (E-MLV) requires host cell endocytosis, the involvement of endocytosis in cell fusion is unclear. By fluorescent microscopic analysis of the fusion-from-within, we found that fragments of target cells are internalized into Env-expressing cells. Treatment of the Env-expressing cells with an endocytosis inhibitor more significantly inhibited the cell fusion than that of the target cells, indicating that endocytosis in Env-expressing cells is required for the cell fusion. The endocytosis inhibitor also attenuated the fusion-from-without. Electron microscopic analysis suggested that the membrane fusion resulting in fusion-from-within initiates in endocytic membrane dents. This study shows that two types of the viral cell fusion both require endocytosis, and provides the cascade of fusion-from-within. PMID:26834711

Centrosomal Latency of Incoming Foamy Viruses in Resting Cells

PubMed Central

Giron, Marie Lou; Roingeard, Philippe; Clave, Emmanuel; Tobaly-Tapiero, Joelle; Bittoun, Patricia; Toubert, Antoine; de Thé, Hugues; Saïb, Ali

2007-01-01

Completion of early stages of retrovirus infection depends on the cell cycle. While gammaretroviruses require mitosis for proviral integration, lentiviruses are able to replicate in post-mitotic non-dividing cells. Resting cells such as naive resting T lymphocytes from peripheral blood cannot be productively infected by retroviruses, including lentiviruses, but the molecular basis of this restriction remains poorly understood. We demonstrate that in G0 resting cells (primary fibroblasts or peripheral T cells), incoming foamy retroviruses accumulate in close proximity to the centrosome, where they lie as structured and assembled capsids for several weeks. Under these settings, virus uncoating is impaired, but upon cell stimulation, Gag proteolysis and capsid disassembly occur, which allows viral infection to proceed. The data imply that foamy virus uncoating is the rate-limiting step for productive infection of primary G0 cells. Incoming foamy retroviruses can stably persist at the centrosome, awaiting cell stimulation to initiate capsid cleavage, nuclear import, and viral gene expression. PMID:17530924

Generation of mammalian cells stably expressing multiple genes at predetermined levels.

PubMed

Liu, X; Constantinescu, S N; Sun, Y; Bogan, J S; Hirsch, D; Weinberg, R A; Lodish, H F

2000-04-10

Expression of cloned genes at desired levels in cultured mammalian cells is essential for studying protein function. Controlled levels of expression have been difficult to achieve, especially for cell lines with low transfection efficiency or when expression of multiple genes is required. An internal ribosomal entry site (IRES) has been incorporated into many types of expression vectors to allow simultaneous expression of two genes. However, there has been no systematic quantitative analysis of expression levels in individual cells of genes linked by an IRES, and thus the broad use of these vectors in functional analysis has been limited. We constructed a set of retroviral expression vectors containing an IRES followed by a quantitative selectable marker such as green fluorescent protein (GFP) or truncated cell surface proteins CD2 or CD4. The gene of interest is placed in a multiple cloning site 5' of the IRES sequence under the control of the retroviral long terminal repeat (LTR) promoter. These vectors exploit the approximately 100-fold differences in levels of expression of a retrovirus vector depending on its site of insertion in the host chromosome. We show that the level of expression of the gene downstream of the IRES and the expression level and functional activity of the gene cloned upstream of the IRES are highly correlated in stably infected target cells. This feature makes our vectors extremely useful for the rapid generation of stably transfected cell populations or clonal cell lines expressing specific amounts of a desired protein simply by fluorescent activated cell sorting (FACS) based on the level of expression of the gene downstream of the IRES. We show how these vectors can be used to generate cells expressing high levels of the erythropoietin receptor (EpoR) or a dominant negative Smad3 protein and to generate cells expressing two different cloned proteins, Ski and Smad4. Correlation of a biologic effect with the level of expression of the

Prevalence of koala retrovirus in geographically diverse populations in Australia.

PubMed

Simmons, G S; Young, P R; Hanger, J J; Jones, K; Clarke, D; McKee, J J; Meers, J

2012-10-01

To determine the prevalence of koala retrovirus (KoRV) in selected koala populations and to estimate proviral copy number in a subset of koalas. Blood or tissue samples from 708 koalas in Queensland, New South Wales, Victoria and South Australia were tested for KoRV pol provirus gene using standard polymerase chain reaction (PCR), nested PCR and real-time PCR (qPCR). Prevalence of KoRV provirus-positive koalas was 100% in four regions of Queensland and New South Wales, 72.2% in mainland Victoria, 26.6% on four Victorian islands and 14.8% on Kangaroo Island, South Australia. Estimated proviral copy number per cell in four groups of koalas from Queensland and Victoria showed marked variation, ranging from a mean of 165 copies per cell in the Queensland group to 1.29 × 10(-4) copies per cell in one group of Victorian koalas. The higher prevalence of KoRV-positive koalas in the north of Australia and high proviral loads in Queensland koalas may indicate KoRV entered and became endogenous in the north and is spreading southwards. It is also possible there are genetic differences between koalas in northern and southern Australia that affect susceptibility to KoRV infection or endogenisation, or that environmental factors affecting transmission in northern states are absent or uncommon in southern regions. Although further studies are required, the finding of proviral copy numbers orders of magnitude lower than what would be expected for the presence of a single copy in every cell for many Victorian animals suggests that KoRV is not endogenous in these animals and likely reflects ongoing exogenous infection. © 2012 The Authors. Australian Veterinary Journal © 2012 Australian Veterinary Association.

Evidence for retrovirus and paramyxovirus infection of multiple bat species in china.

PubMed

Yuan, Lihong; Li, Min; Li, Linmiao; Monagin, Corina; Chmura, Aleksei A; Schneider, Bradley S; Epstein, Jonathan H; Mei, Xiaolin; Shi, Zhengli; Daszak, Peter; Chen, Jinping

2014-05-16

Bats are recognized reservoirs for many emerging zoonotic viruses of public health importance. Identifying and cataloguing the viruses of bats is a logical approach to evaluate the range of potential zoonoses of bat origin. We characterized the fecal pathogen microbiome of both insectivorous and frugivorous bats, incorporating 281 individual bats comprising 20 common species, which were sampled in three locations of Yunnan province, by combining reverse transcription polymerase chain reaction (RT-PCR) assays and next-generation sequencing. Seven individual bats were paramyxovirus-positive by RT-PCR using degenerate primers, and these paramyxoviruses were mainly classified into three genera (Rubulavirus, Henipavirus and Jeilongvirus). Various additional novel pathogens were detected in the paramyxovirus-positive bats using Illumina sequencing. A total of 7066 assembled contigs (≥200 bp) were constructed, and 105 contigs matched eukaryotic viruses (of them 103 belong to 2 vertebrate virus families, 1 insect virus, and 1 mycovirus), 17 were parasites, and 4913 were homologous to prokaryotic microorganisms. Among the 103 vertebrate viral contigs, 79 displayed low identity (<70%) to known viruses including human viruses at the amino acid level, suggesting that these belong to novel and genetically divergent viruses. Overall, the most frequently identified viruses, particularly in bats from the family Hipposideridae, were retroviruses. The present study expands our understanding of the bat virome in species commonly found in Yunnan, China, and provides insight into the overall diversity of viruses that may be capable of directly or indirectly crossing over into humans.

Evidence for Retrovirus and Paramyxovirus Infection of Multiple Bat Species in China

PubMed Central

Yuan, Lihong; Li, Min; Li, Linmiao; Monagin, Corina; Chmura, Aleksei A.; Schneider, Bradley S.; Epstein, Jonathan H.; Mei, Xiaolin; Shi, Zhengli; Daszak, Peter; Chen, Jinping

2014-01-01

Bats are recognized reservoirs for many emerging zoonotic viruses of public health importance. Identifying and cataloguing the viruses of bats is a logical approach to evaluate the range of potential zoonoses of bat origin. We characterized the fecal pathogen microbiome of both insectivorous and frugivorous bats, incorporating 281 individual bats comprising 20 common species, which were sampled in three locations of Yunnan province, by combining reverse transcription polymerase chain reaction (RT-PCR) assays and next-generation sequencing. Seven individual bats were paramyxovirus-positive by RT-PCR using degenerate primers, and these paramyxoviruses were mainly classified into three genera (Rubulavirus, Henipavirus and Jeilongvirus). Various additional novel pathogens were detected in the paramyxovirus-positive bats using Illumina sequencing. A total of 7066 assembled contigs (≥200 bp) were constructed, and 105 contigs matched eukaryotic viruses (of them 103 belong to 2 vertebrate virus families, 1 insect virus, and 1 mycovirus), 17 were parasites, and 4913 were homologous to prokaryotic microorganisms. Among the 103 vertebrate viral contigs, 79 displayed low identity (<70%) to known viruses including human viruses at the amino acid level, suggesting that these belong to novel and genetically divergent viruses. Overall, the most frequently identified viruses, particularly in bats from the family Hipposideridae, were retroviruses. The present study expands our understanding of the bat virome in species commonly found in Yunnan, China, and provides insight into the overall diversity of viruses that may be capable of directly or indirectly crossing over into humans. PMID:24841387

Human retroviruses and AIDS 1996. A compilation and analysis of nucleic acid and amino acid sequences

DOE Office of Scientific and Technical Information (OSTI.GOV)

Myers, G.; Foley, B.; Korber, B.

1997-04-01

This compendium and the accompanying floppy diskettes are the result of an effort to compile and rapidly publish all relevant molecular data concerning the human immunodeficiency viruses (HIV) and related retroviruses. The scope of the compendium and database is best summarized by the five parts that it comprises: (1) Nuclear Acid Alignments and Sequences; (2) Amino Acid Alignments; (3) Analysis; (4) Related Sequences; and (5) Database Communications. Information within all the parts is updated throughout the year on the Web site, http://hiv-web.lanl.gov. While this publication could take the form of a review or sequence monograph, it is not so conceived.more » Instead, the literature from which the database is derived has simply been summarized and some elementary computational analyses have been performed upon the data. Interpretation and commentary have been avoided insofar as possible so that the reader can form his or her own judgments concerning the complex information. In addition to the general descriptions of the parts of the compendium, the user should read the individual introductions for each part.« less

Monitoring acute phase proteins in retrovirus infected cats undergoing feline interferon-ω therapy.

PubMed

Leal, R O; Gil, S; Sepúlveda, N; McGahie, D; Duarte, A; Niza, M M R E; Tavares, L

2014-01-01

Recombinant feline interferon-ω therapy is an immunomodulator currently used in the treatment of different retroviral diseases including feline immune deficiency virus and feline leukaemia virus. Although its mechanism of action remains unclear, this drug appears to potentiate the innate response. Acute phase proteins are one of the key components of innate immunity and studies describing their use as a monitoring tool for the immune system in animals undergoing interferon-ω therapy are lacking. This study aimed to determine whether interferon-ω therapy influences acute phase protein concentrations namely serum amyloid-A, α-1-glycoprotein and C-reactive protein. A single-arm study was performed using 16 cats, living in an animal shelter, naturally infected with retroviruses and subjected to the interferon-ω therapy licensed protocol. Samples were collected before (D0), during (D10 and D30) and after therapy (D65). Serum amyloid-A and C-reactive protein were measured by specific enzyme-linked immunosorbent assay kits and α-1-glycoprotein by single radial immunodiffusion. All the acute phase proteins significantly increased in cats undergoing interferon-ω therapy (D0/D65: P<0·05) CLINICAL SIGNIFICANCE: Acute phase proteins appear to be reasonable predictors of innate-immune stimulation and may be useful in the individual monitoring of naturally retroviral infected cats undergoing interferon-ω therapy. © 2013 British Small Animal Veterinary Association.

Investigation of Human Cancers for Retrovirus by Low-Stringency Target Enrichment and High-Throughput Sequencing.

PubMed

Vinner, Lasse; Mourier, Tobias; Friis-Nielsen, Jens; Gniadecki, Robert; Dybkaer, Karen; Rosenberg, Jacob; Langhoff, Jill Levin; Cruz, David Flores Santa; Fonager, Jannik; Izarzugaza, Jose M G; Gupta, Ramneek; Sicheritz-Ponten, Thomas; Brunak, Søren; Willerslev, Eske; Nielsen, Lars Peter; Hansen, Anders Johannes

2015-08-19

Although nearly one fifth of all human cancers have an infectious aetiology, the causes for the majority of cancers remain unexplained. Despite the enormous data output from high-throughput shotgun sequencing, viral DNA in a clinical sample typically constitutes a proportion of host DNA that is too small to be detected. Sequence variation among virus genomes complicates application of sequence-specific, and highly sensitive, PCR methods. Therefore, we aimed to develop and characterize a method that permits sensitive detection of sequences despite considerable variation. We demonstrate that our low-stringency in-solution hybridization method enables detection of <100 viral copies. Furthermore, distantly related proviral sequences may be enriched by orders of magnitude, enabling discovery of hitherto unknown viral sequences by high-throughput sequencing. The sensitivity was sufficient to detect retroviral sequences in clinical samples. We used this method to conduct an investigation for novel retrovirus in samples from three cancer types. In accordance with recent studies our investigation revealed no retroviral infections in human B-cell lymphoma cells, cutaneous T-cell lymphoma or colorectal cancer biopsies. Nonetheless, our generally applicable method makes sensitive detection possible and permits sequencing of distantly related sequences from complex material.

[Antiretroviral drug supply in Argentina: National Program to Combat Human Retroviruses, AIDS, and STDs].

PubMed

Colautti, Marisel; Luppi, Irene; Salamano, Mercedes; Traverso, María Luz; Botta, Carina; Palchik, Valeria

2009-01-01

To evaluate the supply cycle of antiretroviral (ARV) drugs, overseen by the National Program to Combat Human Retroviruses, AIDS, and STDs, through its order fulfillment indicators, and to obtain input from supply chain stakeholders. A study was carried out from April-September 2005 in the pharmacies of two hospitals in Rosario, Argentina, involving both a quantitative analysis of indicators and secondary sources and a qualitative evaluation using semistructured interviews. The indicators reveal the impact that interruptions in ARV supply stream from the Program (central level) have and the overstocking that takes place at the pharmacies (local level) to manage the shortages. Changes in ARV treatment account for over 50% of the prescriptions. Fulfillments fall short of the reference value. The interviewees shared possible strategies for overcoming the communication gaps between levels, for building-up stock, for guaranteeing availability, and for shortening waiting times; reached informal agreements to deal with the lack of policies and the shortage of staff; acknowledged the challenges facing the jurisdictions (central, intermediate, and local/community); and recognized local efforts to improve management. These challenges could be the starting point for building teams to work on effectively decentralizing the entire supply chain and allowing the Program to fulfill its much-needed oversight role.

HTLV-1, the Other Pathogenic Yet Neglected Human Retrovirus: From Transmission to Therapeutic Treatment

PubMed Central

Futsch, Nicolas; Mahieux, Renaud; Dutartre, Hélène

2017-01-01

Going back to their discovery in the early 1980s, both the Human T-cell Leukemia virus type-1 (HTLV-1) and the Human Immunodeficiency Virus type-1 (HIV-1) greatly fascinated the virology scene, not only because they were the first human retroviruses discovered, but also because they were associated with fatal diseases in the human population. In almost four decades of scientific research, both viruses have had different fates, HTLV-1 being often upstaged by HIV-1. However, although being very close in terms of genome organization, cellular tropism, and viral replication, HIV-1 and HTLV-1 are not completely commutable in terms of treatment, especially because of the opposite fate of the cells they infect: death versus immortalization, respectively. Nowadays, the antiretroviral therapies developed to treat HIV-1 infected individuals and to limit HIV-1 spread among the human population have a poor or no effect on HTLV-1 infected individuals, and thus, do not prevent the development of HTLV-1-associated diseases, which still lack highly efficient treatments. The present review mainly focuses on the course of HTLV-1 infection, from the initial infection of the host to diseases development and associated treatments, but also investigates HIV-1/HTLV-1 co-infection events and their impact on diseases development. PMID:29267225

Single residues in the surface subunits of oncogenic sheep retrovirus envelopes distinguish receptor-mediated triggering for fusion at low pH and infection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cote, Marceline; Zheng, Yi-Min; Albritton, Lorraine M.

2011-12-20

Jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus (ENTV) are two closely related oncogenic retroviruses that share the same cellular receptor yet exhibit distinct fusogenicity and infectivity. Here, we find that the low fusogenicity of ENTV envelope protein (Env) is not because of receptor binding, but lies in its intrinsic insensitivity to receptor-mediated triggering for fusion at low pH. Distinct from JSRV, shedding of ENTV surface (SU) subunit into culture medium was not enhanced by a soluble form of receptor, Hyal2 (sHyal2), and sHyal2 was unable to effectively inactivate the ENTV pseudovirions. Remarkably, replacing either of the two aminomore » acid residues, N191 or S195, located in the ENTV SU with the corresponding JSRV residues, H191 or G195, markedly increased the Env-mediated membrane fusion activity and infection. Reciprocal amino acid substitutions also partly switched the sensitivities of ENTV and JSRV pseudovirions to sHyal2-mediated SU shedding and inactivation. While N191 is responsible for an extra N-linked glycosylation of ENTV SU relative to that of JSRV, S195 possibly forms a hydrogen bond with a surrounding amino acid residue. Molecular modeling of the pre-fusion structure of JSRV Env predicts that the segment of SU that contains H191 to G195 contacts the fusion peptide and suggests that the H191N and G195S changes seen in ENTV may stabilize its pre-fusion structure against receptor priming and therefore modulate fusion activation by Hyal2. In summary, our study reveals critical determinants in the SU subunits of JSRV and ENTV Env proteins that likely regulate their local structures and thereby differential receptor-mediated fusion activation at low pH, and these findings explain, at least in part, their distinct viral infectivity.« less

Comprehensive Analysis of Human Endogenous Retrovirus Group HERV-W Locus Transcription in Multiple Sclerosis Brain Lesions by High-Throughput Amplicon Sequencing

PubMed Central

Schmitt, Katja; Richter, Christin; Backes, Christina; Meese, Eckart; Ruprecht, Klemens

2013-01-01

Human endogenous retroviruses (HERVs) of the HERV-W group comprise hundreds of loci in the human genome. Deregulated HERV-W expression and HERV-W locus ERVWE1-encoded Syncytin-1 protein have been implicated in the pathogenesis of multiple sclerosis (MS). However, the actual transcription of HERV-W loci in the MS context has not been comprehensively analyzed. We investigated transcription of HERV-W in MS brain lesions and white matter brain tissue from healthy controls by employing next-generation amplicon sequencing of HERV-W env-specific reverse transcriptase (RT) PCR products, thus revealing transcribed HERV-W loci and the relative transcript levels of those loci. We identified more than 100 HERV-W loci that were transcribed in the human brain, with a limited number of loci being predominantly transcribed. Importantly, relative transcript levels of HERV-W loci were very similar between MS and healthy brain tissue samples, refuting deregulated transcription of HERV-W env in MS brain lesions, including the high-level-transcribed ERVWE1 locus encoding Syncytin-1. Quantitative RT-PCR likewise did not reveal differences in MS regarding HERV-W env general transcript or ERVWE1- and ERVWE2-specific transcript levels. However, we obtained evidence for interindividual differences in HERV-W transcript levels. Reporter gene assays indicated promoter activity of many HERV-W long terminal repeats (LTRs), including structurally incomplete LTRs. Our comprehensive analysis of HERV-W transcription in the human brain thus provides important information on the biology of HERV-W in MS lesions and normal human brain, implications for study design, and mechanisms by which HERV-W may (or may not) be involved in MS. PMID:24109235

In vivo treatment with interleukin 12 protects mice from immune abnormalities observed during murine acquired immunodeficiency syndrome (MAIDS)

PubMed Central

1994-01-01

Lymphoproliferation, chronic B cell activation resulting in hypergammaglobulinemia, and profound immunodeficiency are prominent features of a retrovirus-induced syndrome designated murine acquired immunodeficiency syndrome (MAIDS). In vivo treatment of infected mice with recombinant interleukin 12 (IL-12) beginning at the time of infection or up to 9 wk after virus inoculation markedly inhibited the development of splenomegaly and lymphadenopathy, as well as B cell activation and Ig secretion. Treatment with IL-12 also had major effects in preventing induction of several immune defects including impaired production of interferon gamma (IFN-gamma) and IL-2 and depressed proliferative responses to various stimuli. The therapeutic effects of IL-12 on the immune system of mice with MAIDS were also associated with reduced expression of the retrovirus that causes this disease (BM5def), with lesser effects on expression of ecotropic MuLV. IL-12 treatment was not effective in IFN-gamma knockout mice or in infected mice treated simultaneously with IL-12 and anti-IFN-gamma. These results demonstrate that induction and progression of MAIDS are antagonized by IL-12 through high-level expression of IFN-gamma and may provide an experimental basis for developing treatments of retrovirus- induced immune disorders with similar immunopathogenic mechanisms. PMID:7964495

Evidence for retrovirus infections in green turtles Chelonia mydas from the Hawaiian islands

USGS Publications Warehouse

Casey, R.N.; Quackenbush, S.L.; Work, Thierry M.; Balazs, G.H.; Bowser, P.R.; Casey, J.W.

1997-01-01

Apparently normal Hawaiian green turtles Chelonia mydas and those displaying fibropapillomas were analyzed for infection by retroviruses. Strikingly, all samples were positive for polymerase enhanced reverse transcriptase (PERT) with levels high enough to quantitate by the conventional reverse transcriptase (RT) assay. However, samples of skin, even from asymptomatic turtles, were RT positive, although the levels of enzyme activity in healthy turtles hatched and raised in captivity were much lower than those observed in asymptomatic free-ranging turtles. Turtles with fibropapillomas displayed a broad range of reverse transcriptase activity. Skin and eye fibropapillomas and a heart tumor were further analyzed and shown to have reverse transcriptase activity that banded in a sucrose gradient at 1.17 g ml-1. The reverse transcriptase activity purified from the heart tumor displayed a temperature optimum of 37??C and showed a preference for Mn2+ over Mg2+. Sucrose gradient fractions of this sample displaying elevated reverse transcriptase activity contained primarily retrovitalsized particles with prominent envelope spikes, when negatively stained and examined by electron microscopy. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of gradient-purified virions revealed a conserved profile among 4 independent tumors and showed 7 prominent proteins having molecular weights of 116, 83, 51, 43, 40, 20 and 14 kDa. The data suggest that retroviral infections are widespread in Hawaiian green turtles and a comprehensive investigation is warranted to address the possibility that these agents cause green turtle fibropapillomatosis (GTFP).

An exogenous retrovirus isolated from koalas with malignant neoplasias in a US zoo.

PubMed

Xu, Wenqin; Stadler, Cynthia K; Gorman, Kristen; Jensen, Nathaniel; Kim, David; Zheng, HaoQiang; Tang, Shaohua; Switzer, William M; Pye, Geoffrey W; Eiden, Maribeth V

2013-07-09

Leukemia and lymphoma account for more than 60% of deaths in captive koalas (Phascolarctos cinereus) in northeastern Australia. Although the endogenizing gammaretrovirus koala endogenous retrovirus (KoRV) was isolated from these koalas, KoRV has not been definitively associated with leukemogenesis. We performed KoRV screening in koalas from the San Diego Zoo, maintained for more than 45 y with very limited outbreeding, and the Los Angeles Zoo, maintained by continuously assimilating captive-born Australian koalas. San Diego Zoo koalas are currently free of malignant neoplasias and were infected with only endogenous KoRV, which we now term subtype "KoRV-A," whereas Los Angeles Zoo koalas with lymphomas/leukemias are infected in addition to KoRV-A by a unique KoRV we term subtype "KoRV-B." KoRV-B is most divergent in the envelope protein and uses a host receptor distinct from KoRV-A. KoRV-B also has duplicated enhancer regions in the LTR associated with increased pathology in gammaretroviruses. Whereas KoRV-A uses the sodium-dependent phosphate transporter 1 (PiT1) as a receptor, KoRV-B employs a different receptor, the thiamine transporter 1 (THTR1), to infect cells. KoRV-B is transmitted from dam to offspring through de novo infection, rather than via genetic inheritance like KoRV-A. Detection of KoRV-B in native Australian koalas should provide a history, and a mode for remediation, of leukemia/lymphoma currently endemic in this population.

HTLV-1 basic leucine zipper factor downregulates cyclin D1 expression via interactions with NF-κB.

PubMed

Ma, Yunyun; Zhang, Bo; Wang, Dong; Qian, Lili; Song, Xianmei; Wang, Xueyin; Yang, Chaokuan; Zhao, Guoqiang

2017-03-01

Human T cell leukemia virus type 1 (HTLV-1) is an oncogenic retrovirus. It can cause adult T cell leukemia (ATL) and other diseases. The HTLV-1 basic leucine zipper (bZIP) factor (HBZ), which is encoded by the minus-strand of the provirus, is expressed in all cases of ATL and involved in T cell proliferation. However, the exact mechanism underlying its growth-promoting activity is poorly understood. Herein, we demonstrated that HBZ suppressed cyclin D1 expression by inhibiting the nuclear factor (NF)-κB signaling pathway. Among the potential mechanisms of cyclin D1 inhibition mediated by HBZ, we found that HBZ suppressed cyclin D1 promoter activity. Luciferase assay analysis revealed that HBZ repressed cyclin D1 promoter activity by suppressing NF-κB‑driven transcription mediated by the p65 subunit. Using an immunoprecipitation assay, we found that HBZ could bind to p65, but not p50. Finally, we showed that HBZ selectively interacted with p65 via its AD+bZIP domains. By suppressing cyclin D1 expression, HBZ can alter cell cycle progression of HTLV-1-infected cells, which may be critical for oncogenesis.

Capsid-binding retrovirus restriction factors: discovery, restriction specificity and implications for the development of novel therapeutics.

PubMed

Sanz-Ramos, Marta; Stoye, Jonathan P

2013-12-01

The development of drugs against human immunodeficiency virus type 1 infection has been highly successful, and numerous combinational treatments are currently available. However, the risk of the emergence of resistance and the toxic effects associated with prolonged use of antiretroviral therapies have emphasized the need to consider alternative approaches. One possible area of investigation is provided by the properties of restriction factors, cellular proteins that protect organisms against retroviral infection. Many show potent viral inhibition. Here, we describe the discovery, properties and possible therapeutic uses of the group of restriction factors known to interact with the capsid core of incoming retroviruses. This group comprises Fv1, TRIM5α and TRIMCypA: proteins that all act shortly after virus entry into the target cell and block virus replication at different stages prior to integration of viral DNA into the host chromosome. They have different origins and specificities, but share general structural features required for restriction, with an N-terminal multimerization domain and a C-terminal capsid-binding domain. Their overall efficacy makes it reasonable to ask whether they might provide a framework for developing novel antiretroviral strategies.

Murine Cytomegalovirus Downregulates Interleukin-17 in Mice with Retrovirus-induced Immunosuppression that are Susceptible to Experimental Cytomegalovirus Retinitis

PubMed Central

Blalock, Emily L.; Chien, Hsin; Dix, Richard D.

2013-01-01

Interleukin-17 (IL-17), a proinflammatory cytokine produced by CD4+ Th17 cells, has been associated with the pathogenesis of several autoimmune diseases including uveitis. The fate of IL-17 during HIV/AIDS, however, remains unclear, and a possible role for IL-17 in the pathogenesis of AIDS-related diseases has not been investigated. Toward these ends, we performed studies using a well-established animal model of experimental murine cytomegalovirus (MCMV) retinitis that develops in C57/BL6 mice with retrovirus-induced immunosuppression (MAIDS). After establishing baseline levels for IL-17 production in whole splenic cells of healthy mice, we observed a significant increase in IL-17 mRNA levels in whole splenic cells of mice with MAIDS of 4-weeks (MAIDS-4), 8-weeks (MAIDS-8), and 10-weeks (MAIDS-10) duration. In contrast, enriched populations of splenic CD4+ T cells, splenic macrophages, and splenic neutrophils exhibited a reproducible decrease in levels of IL-17 mRNA during MAIDS progression. To explore a possible role for IL-17 during the pathogenesis of MAIDS-related MCMV retinitis, we first demonstrated constitutive IL-17 expression in retinal photoreceptor cells of uninfected eyes of healthy mice. Subsequent studies, however, revealed a significant decrease in intraocular levels of IL-17 mRNA and protein in MCMV-infected eyes of MAIDS-10 mice during retinitis development. That MCMV infection might cause a remarkable downregulation of IL-17 production was supported further by the finding that systemic MCMV infection of healthy, MAIDS-4, or MAIDS-10 mice also significantly decreased IL-17 mRNA production by whole splenic CD4+ T cells. Based on additional studies using IL-10 −/− mice infected systemically with MCMV and IL-10 −/− mice with MAIDS infected intraocularly with MCMV, we propose that MCMV infection downregulates IL-17 production via stimulation of suppressor of cytokine signaling (SOCS)-3 and interleukin-10. PMID:23415673

Design of a Retrovirus-Derived Vector for Expression and Transduction of Exogenous Genes in Mammalian Cells

PubMed Central

Perkins, Archibald S.; Kirschmeier, Paul T.; Gattoni-Celli, Sebastiano; Weinstein, I. Bernard

1983-01-01

We have developed a transfection vector for animal cells that contains long terminal repeat (LTR) sequences to promote expression. Plasmid p101/101, a derivative of plasmid pBR322 containing the complete Moloney murine sarcoma virus genome, was cut with restriction enzymes and religated so that both the 5′ and 3′ LTRs were retained and all but about 700 base pairs of the intervening viral sequences were removed. To test this vector, the Escherichia coli gene gpt was cloned into a unique PstI site, between the two LTRs, with guanine and cytosine tailing, a method that can be generalized for insertion of any DNA segment into this vector. When DNA from recombinant plasmids in which the gpt gene was inserted in the same transcriptional polarity as the LTR sequences was transfected onto murine or rat fibroblast cultures, we obtained a high yield of Gpt+ colonies. However, plasmid constructs with the gpt gene in the opposite polarity were virtually devoid of activity. With gpt in the proper orientation, restriction enzyme cuts within the LTRs or between the 5′ LTR and the gpt gene reduced transfection by more than 98%, whereas a cut between the gpt gene and the 3′ LTR gave an 80% reduction in activity. Thus, both 5′ and 3′ LTR sequences are essential for optimal gpt expression, although the 5′ LTR appears to play a more important role. When the LTR-gpt plasmid was transfected onto murine leukemia virus-infected mouse fibroblasts, we obtained evidence that RNA copies became pseudotyped into viral particles which could transfer the Gpt+ phenotype into rodent cells with extremely high efficiency. This vector should prove useful for high-efficiency transduction of a variety of genes in mammalian cells. Images PMID:6308426

SEPARATIONS RESEARCH AT THE UNITED STATES ENVIRONMENTAL PROTECTION AGENCY - TOWARDS RECOVERY OF VOCS AND METALS USING MEMBRANES AND ADSORPTION PROCESSES

EPA Science Inventory

The USEPA's National Risk Management Research Laboratory is investigating new separations materials and processes for removal and recovery of volatile organic compounds (VOCs) and toxic metals from wastestreams and industrial process streams. Research applying membrane-based perv...

Development of data measurement techniques for traffic operations analysis at intersections

DOT National Transportation Integrated Search

1985-06-01

This study was undertaken for the purpose of developing intersection data collection methods and approaches which reflect the current needs of engineers and the state-of-the-art capabilities of available equipment. A literature review summarizes perv...

Nemo-like kinase (NLK) expression in osteoblastic cells and suppression of osteoblastic differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nifuji, Akira, E-mail: nifuji-a@tsurumi-u.ac.jp; Department of Pharmacology, Tsurumi University School of Dental Medicine, Yokohama; Ideno, Hisashi

2010-04-15

Mitogen-activated protein kinases (MAPKs) regulate proliferation and differentiation in osteoblasts. The vertebral homologue of nemo, nemo-like kinase (NLK), is an atypical MAPK that targets several signaling components, including the T-cell factor/lymphoid enhancer factor (TCF/Lef1) transcription factor. Recent studies have shown that NLK forms a complex with the histone H3-K9 methyltransferase SETDB1 and suppresses peroxisome proliferator-activated receptor (PPAR)-gamma:: action in the mesenchymal cell line ST2. Here we investigated whether NLK regulates osteoblastic differentiation. We showed that NLK mRNA is expressed in vivo in osteoblasts at embryonic day 18.5 (E18.5) mouse calvariae. By using retrovirus vectors, we performed forced expression of NLKmore » in primary calvarial osteoblasts (pOB cells) and the mesenchymal cell line ST2. Wild-type NLK (NLK-WT) suppressed alkaline phosphatase activity and expression of bone marker genes such as alkaline phosphatase, type I procollagen, runx2, osterix, steopontin and osteocalcin in these cells. NLK-WT also decreased type I collagen protein expression in pOB and ST2 cells. Furthermore, mineralized nodule formation was reduced in pOB cells overexpressing NLK-WT. In contrast, kinase-negative form of NLK (NLK-KN) did not suppress or partially suppress ALP activity and bone marker gene expression in pOB and ST2 cells. NLK-KN did not suppress nodule formation in pOB cells. In addition to forced expression, suppression of endogenous NLK expression by siRNA increased bone marker gene expression in pOB and ST2 cells. Finally, transcriptional activity analysis of gene promoters revealed that NLK-WT suppressed Wnt1 activation of TOP flash promoter and Runx2 activation of the osteocalcin promoter. Taken together, these results suggest that NLK negatively regulates osteoblastic differentiation.« less

Suppressive effect on polyclonal B-cell activation of a synthetic peptide homologous to a transmembrane component of oncogenic retroviruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mitani, M.; Cianciolo, G.J.; Snyderman, R.

1987-01-01

Purified feline leukemia virus, UV light-inactivated feline leukemia virus, and a synthetic peptide (CKS-17) homologous to a well-conserved region of the transmembrane components of several human and animal retroviruses were each studied for their effect on IgG production by feline peripheral blood lymphocytes. Using a reverse hemolytic plaque assay, both the viable virus and the UV-inactivated feline leukemia virus, but not the CKS-17, activated B lymphocytes to secrete IgG. When staphylococcal protein A, a polyclonal B-cell activator, was used to stimulate IgG synthesis by feline lymphocytes, the viable virus, the UV-inactivated virus, and the CKS-17 peptide each strongly suppressed IgGmore » secretion without compromising viability of the lymphocytes. These finding suggest that the immunosuppressive influences of feline leukemia virus on immunoglobulin synthesis may reside in a conserved portion of the envelope glycoprotein that includes the region homologous to CKS-17.« less

Not so bad after all: retroviruses and long terminal repeat retrotransposons as a source of new genes in vertebrates.

PubMed

Naville, M; Warren, I A; Haftek-Terreau, Z; Chalopin, D; Brunet, F; Levin, P; Galiana, D; Volff, J-N

2016-04-01

Viruses and transposable elements, once considered as purely junk and selfish sequences, have repeatedly been used as a source of novel protein-coding genes during the evolution of most eukaryotic lineages, a phenomenon called 'molecular domestication'. This is exemplified perfectly in mammals and other vertebrates, where many genes derived from long terminal repeat (LTR) retroelements (retroviruses and LTR retrotransposons) have been identified through comparative genomics and functional analyses. In particular, genes derived from gag structural protein and envelope (env) genes, as well as from the integrase-coding and protease-coding sequences, have been identified in humans and other vertebrates. Retroelement-derived genes are involved in many important biological processes including placenta formation, cognitive functions in the brain and immunity against retroelements, as well as in cell proliferation, apoptosis and cancer. These observations support an important role of retroelement-derived genes in the evolution and diversification of the vertebrate lineage. Copyright © 2016 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Retention of "safe" blood donors. The Retrovirus Epidemiology Donor Study.

PubMed

Thomson, R A; Bethel, J; Lo, A Y; Ownby, H E; Nass, C C; Williams, A E

1998-04-01

There are obvious advantages to increasing donor retention. However, for reasons of blood safety, certain donors may, in fact, be more desirable to retain than others. "Safe" donors are defined as those who provided a blood donation that was negative on all laboratory screening tests and who subsequently reported no behavioral risks in response to an anonymous survey. This study identifies the most important factors affecting the intention of "safe" donors to provide another donation. An anonymous survey asking about donation history, sexual history, injecting drug use, and recent donation experience was mailed to 50,162 randomly selected allogeneic donors (including directed donors) who gave blood from April through July or from October through December 1993 at one of the five United States blood centers participating in the Retrovirus Epidemiology Donor Study. Before mailing, questionnaires were coded to designate donors with nonreactive laboratory screening tests at their most recent donation. A total of 34,726 donors (69%) responded, with substantially higher response among repeat donors. According to reported intentions only, the vast majority of "safe" donors indicated a high likelihood of donating again within the next 12 months. Only 3.4 percent reported a low likelihood of donating again. A comparison of those likely to return and those unlikely to return reveals significant differences in demographics and in ratings of the donation experience. A higher proportion of those unlikely to return were first-time donors, minority-group donors, and donors with less education. The highest projected loss among "safe" donors was seen for those who gave a fair to poor assessment of their treatment by blood center staff or of their physical well-being during or after donating. These data suggest that efforts to improve donors' perceptions of their donation experience, as well as attention to the physical effects of blood donation, may aid in the retention of both

Immortalized bovine mammary epithelial cells express stem cell markers and differentiate in vitro.

PubMed

Hu, Han; Zheng, Nan; Gao, Haina; Dai, Wenting; Zhang, Yangdong; Li, Songli; Wang, Jiaqi

2016-08-01

The bovine mammary epithelial cell is a secretory cell, and its cell number and secretory activity determine milk production. In this study, we immortalized a bovine mammary epithelial cell line by SV40 large T antigen gene using a retrovirus based on Chinese Holstein primary mammary epithelial cells (CMEC) cultured in vitro. An immortalized bovine mammary epithelial cell line surpassed the 50-passage mark and was designated the CMEC-H. The immortalized mammary epithelial cells grew in close contact with each other and exhibited the typical cobblestone morphology characteristic with obvious boundaries. The telomerase expression of CMEC-H has consistently demonstrated the presence of telomerase activity as an immortalized cell line, but the cell line never induced tumor formation in nude mice. CMEC-H expressed epithelial (cytokeratins CK7, CK8, CK18, and CK19), mesenchymal (vimentin), and stem/progenitor (CD44 and p63) cell markers. The induced expression of milk proteins, αS1 -casein, β-casein, κ-casein, and butyrophilin, indicated that CMEC-H maintained the synthesis function of the mammary epithelial cells. The established immortalized bovine mammary epithelial cell line CMEC-H is capable of self-renewal and differentiation and can serve as a valuable reagent for studying the physiological mechanism of the mammary gland. © 2016 International Federation for Cell Biology.

Assessing state efforts to integrate transportation, land use and climate change.

DOT National Transportation Integrated Search

2016-12-01

Climate change is increasingly recognized as a threat to life on earth. Continued emission of greenhouse gases will cause further : warming and long-lasting changes in all components of the climate system, increasing the likelihood of severe, perv...

Human leukemia antigen-A*0201-restricted epitopes of human endogenous retrovirus W family envelope (HERV-W env) induce strong cytotoxic T lymphocyte responses.

PubMed

Tu, Xiaoning; Li, Shan; Zhao, Lijuan; Xiao, Ran; Wang, Xiuling; Zhu, Fan

2017-08-01

Human endogenous retrovirus W family (HERV-W) envelope (env) has been reported to be related to several human diseases, including autoimmune disorders, and it could activate innate immunity. However, there are no reports investigating whether human leukemia antigen (HLA)-A*0201 + restriction is involved in the immune response caused by HERV-W env in neuropsychiatric diseases. In the present study, HERV-W env-derived epitopes presented by HLA-A*0201 are described with the potential for use in adoptive immunotherapy. Five peptides displaying HLA-A*0201-binding motifs were predicted using SYFEPITHI and BIMAS, and synthesized. A CCK-8 assay showed peptides W, Q and T promoted lymphocyte proliferation. Stimulation of peripheral blood mononuclear cells from HLA-A*0201 + donors with each of these peptides induced peptide-specific CD8 + T cells. High numbers of IFN-γ-secreting T cells were also detectable after several weekly stimulations with W, Q and T. Besides lysis of HERV-W env-loaded target cells, specific apoptosis was also observed. These data demonstrate that human T cells can be sensitized toward HERV-W env peptides (W, Q and T) and, moreover, pose a high killing potential toward HERV-W env-expressing U251 cells. In conclusion, peptides W Q and T, which are HERV-W env antigenic epitopes, have both antigenicity and immunogenicity, and can cause strong T cell immune responses. Our data strengthen the view that HERV-W env should be considered as an autoantigen that can induce autoimmunity in neuropsychiatric diseases, such as multiple sclerosis and schizophrenia. These data might provide an experimental foundation for a HERV-W env peptide vaccine and new insight into the treatment of neuropsychiatric diseases.

HPV-16 E7 expression up-regulates phospholipase D activity and promotes rapamycin resistance in a pRB-dependent manner.

PubMed

Rabachini, Tatiana; Boccardo, Enrique; Andrade, Rubiana; Perez, Katia Regina; Nonogaki, Suely; Cuccovia, Iolanda Midea; Villa, Luisa Lina

2018-04-27

Human Papillomavirus (HPV) infection is the main risk factor for the development and progression of cervical cancer. HPV-16 E6 and E7 expression is essential for induction and maintenance of the transformed phenotype. These oncoproteins interfere with the function of several intracellular proteins, including those controlling the PI3K/AKT/mTOR pathway in which Phospolipase D (PLD) and Phosphatidic acid (PA) play a critical role. PLD activity was measured in primary human keratinocytes transduced with retroviruses expressing HPV-16 E6, E7 or E7 mutants. The cytostatic effect of rapamycin, a well-known mTOR inhibitor with potential clinical applications, was evaluated in monolayer and organotypic cultures. HPV-16 E7 expression in primary human keratinocytes leads to an increase in PLD expression and activity. Moreover, this activation is dependent on the ability of HPV-16 E7 to induce retinoblastoma protein (pRb) degradation. We also show that cells expressing HPV-16 E7 or silenced for pRb acquire resistance to the antiproliferative effect of rapamycin. This is the first indication that HPV oncoproteins can affect PLD activity. Since PA can interfere with the ability of rapamycin to bind mTOR, the use of combined strategies to target mTOR and PLD activity might be considered to treat HPV-related malignancies.

Evolutionary dynamics of endogenous Jaagsiekte sheep retroviruses proliferation in the domestic sheep, mouflon and Pyrenean chamois.

PubMed

Sistiaga-Poveda, M; Jugo, B M

2014-06-01

The oncogenic exogenous Jaagsiekte sheep retrovirus (JSRV), responsible for ovine pulmonary adenocarcinoma, has several endogenous counterparts termed enJSRVs. Although many of these elements have been inactivated over time by the accumulation of deleterious mutations or internal recombination leading to solo long terminal repeat (LTR) formation, several members of enJSRVs have been identified as nearly intact and probably represent recent integration events. To determine the level of enJSRV polymorphism in the sheep population and related species, we have undertaken a study by characterizing enJSRVs copies and independent integration sites in six domestic sheep and two wild species of the sheep lineage. enJSRVs copies were detected by amplifying the env-LTR region by PCR, and for the detection of the insertion sites, we used two approaches: (1) an in silico approach based on the recently published Sheep Reference Genome Assembly (OARv3.0) and (2) an experimental approach based on PCR suppression and inverse PCR techniques. In total, 103 enJSRV sequences were generated across 10 individuals and enJSRV integrations were found on 11 of the 28 sheep chromosomes. These findings suggest that there are still uncharacterized enJSRVs, and that some of the integration sites are variable among the different species, breeds of the same species, subspecies and geographic locations.

Gene Expression Analysis of Breast Cancer Progression

DTIC Science & Technology

2005-07-01

Oncogene 22, 761-768. 18. Wroblewski, L. E., Pritchard, D. M., Carter, S. & Varro, A. (20012) Biochent. 36. Smith, L. M., Wise, S. C., Hendricks , D. T...in DMEM medium supplemented with 10% fetal bovine serum (FBS), penicillin , streptomycin and fungizone. Phoenix cells, a helper cell line for retrovirus...fetal bovine serum, 2% l-glutamine, NEAA, 1mM sodium pyruvate, 1.5g/L sodium bicarbonate, 100 I.U./ml penicillin and 100[g/ml streptomycin. Cells

Novel gene expression mechanism in a fission yeast retroelement: Tf1 proteins are derived from a single primary translation product.

PubMed

Levin, H L; Weaver, D C; Boeke, J D

1993-12-01

In sharp contrast to the single ORF of the Schizosaccharomyces pombe retrotransposon Tf1, retroviruses and most retrotransposons employ two different ORFs to separately encode the Gag and Pol proteins. The different ORFs are thought to allow for overexpression of the Gag protein relative to Pol protein presumed necessary for the assembly of functional retrovirus particles and virus-like particles (VLPs). The results of in vivo experiments designed to detect the transposition of Tf1 show that Tf1 is indeed active and can insert itself into the host genome via a true retrotransposition process. Thus, a paradox emerged between the lack of any obvious means of overexpressing Tf1 Gag protein and the demonstrated functionality of the element. Epitope tagging experiments described here confirm that the Tf1 large ORF is intact and that there is no translational or transcriptional mechanism used to overexpress the Tf1 Gag protein. In addition, we used sucrose gradients and antisera specific for Tf1 capsid (CA) and integrase (IN) to show that the Tf1 proteins do assemble into uniform populations of macromolecular particles that also cosediment with Tf1 reverse transcription products. This evidence suggests that Tf1 proteins form VLPs without using the previously described mechanisms that retroviruses and retrotransposons require to overexpress Gag proteins.

Papio cynocephalus endogenous retrovirus among old world monkeys: evidence for coevolution and ancient cross-species transmissions.

PubMed

Mang, R; Maas, J; van Der Kuyl, A C; Goudsmit, J

2000-02-01

To study the evolutionary history of Papio cynocephalus endogenous retrovirus (PcEV), we analyzed the distribution and genetic characteristics of PcEV among 17 different species of primates. The viral pol-env and long terminal repeat and untranslated region (LTR-UTR) sequences could be recovered from all Old World species of the papionin tribe, which includes baboons, macaques, geladas, and mangabeys, but not from the New World monkeys and hominoids we tested. The Old World genera Cercopithecus and Miopithecus hosted either a PcEV variant with an incomplete genome or a virus with substantial mismatches in the LTR-UTR. A complete PcEV was found in the genome of Colobus guereza-but not in Colobus badius-with a copy number of 44 to 61 per diploid genome, comparable to that seen in papionins, and with a sequence most closely related to a virus of the papionin tribe. Analysis of evolutionary distances among PcEV sequences for synonymous and nonsynonymous sites indicated that purifying selection was operational during PcEV evolution. Phylogenetic analysis suggested that possibly two subtypes of PcEV entered the germ line of a common ancestor of the papionins and subsequently coevolved with their hosts. One strain of PcEV was apparently transmitted from a papionin ancestor to an ancestor of the central African lowland C. guereza.

Papio cynocephalus Endogenous Retrovirus among Old World Monkeys: Evidence for Coevolution and Ancient Cross-Species Transmissions

PubMed Central

Mang, Rui; Maas, Jolanda; van der Kuyl, Antoinette C.; Goudsmit, Jaap

2000-01-01

To study the evolutionary history of Papio cynocephalus endogenous retrovirus (PcEV), we analyzed the distribution and genetic characteristics of PcEV among 17 different species of primates. The viral pol-env and long terminal repeat and untranslated region (LTR-UTR) sequences could be recovered from all Old World species of the papionin tribe, which includes baboons, macaques, geladas, and mangabeys, but not from the New World monkeys and hominoids we tested. The Old World genera Cercopithecus and Miopithecus hosted either a PcEV variant with an incomplete genome or a virus with substantial mismatches in the LTR-UTR. A complete PcEV was found in the genome of Colobus guereza—but not in Colobus badius—with a copy number of 44 to 61 per diploid genome, comparable to that seen in papionins, and with a sequence most closely related to a virus of the papionin tribe. Analysis of evolutionary distances among PcEV sequences for synonymous and nonsynonymous sites indicated that purifying selection was operational during PcEV evolution. Phylogenetic analysis suggested that possibly two subtypes of PcEV entered the germ line of a common ancestor of the papionins and subsequently coevolved with their hosts. One strain of PcEV was apparently transmitted from a papionin ancestor to an ancestor of the central African lowland C. guereza. PMID:10627573

Evolutionary dynamics of endogenous Jaagsiekte sheep retroviruses proliferation in the domestic sheep, mouflon and Pyrenean chamois

PubMed Central

Sistiaga-Poveda, M; Jugo, B M

2014-01-01

The oncogenic exogenous Jaagsiekte sheep retrovirus (JSRV), responsible for ovine pulmonary adenocarcinoma, has several endogenous counterparts termed enJSRVs. Although many of these elements have been inactivated over time by the accumulation of deleterious mutations or internal recombination leading to solo long terminal repeat (LTR) formation, several members of enJSRVs have been identified as nearly intact and probably represent recent integration events. To determine the level of enJSRV polymorphism in the sheep population and related species, we have undertaken a study by characterizing enJSRVs copies and independent integration sites in six domestic sheep and two wild species of the sheep lineage. enJSRVs copies were detected by amplifying the env-LTR region by PCR, and for the detection of the insertion sites, we used two approaches: (1) an in silico approach based on the recently published Sheep Reference Genome Assembly (OARv3.0) and (2) an experimental approach based on PCR suppression and inverse PCR techniques. In total, 103 enJSRV sequences were generated across 10 individuals and enJSRV integrations were found on 11 of the 28 sheep chromosomes. These findings suggest that there are still uncharacterized enJSRVs, and that some of the integration sites are variable among the different species, breeds of the same species, subspecies and geographic locations. PMID:24690757

Induction of ebolavirus cross-species immunity using retrovirus-like particles bearing the Ebola virus glycoprotein lacking the mucin-like domain.

PubMed

Ou, Wu; Delisle, Josie; Jacques, Jerome; Shih, Joanna; Price, Graeme; Kuhn, Jens H; Wang, Vivian; Verthelyi, Daniela; Kaplan, Gerardo; Wilson, Carolyn A

2012-01-25

The genus Ebolavirus includes five distinct viruses. Four of these viruses cause hemorrhagic fever in humans. Currently there are no licensed vaccines for any of them; however, several vaccines are under development. Ebola virus envelope glycoprotein (GP1,2) is highly immunogenic, but antibodies frequently arise against its least conserved mucin-like domain (MLD). We hypothesized that immunization with MLD-deleted GP1,2 (GPΔMLD) would induce cross-species immunity by making more conserved regions accessible to the immune system. To test this hypothesis, mice were immunized with retrovirus-like particles (retroVLPs) bearing Ebola virus GPΔMLD, DNA plasmids (plasmo-retroVLP) that can produce such retroVLPs in vivo, or plasmo-retroVLP followed by retroVLPs. Cross-species neutralizing antibody and GP1,2-specific cellular immune responses were successfully induced. Our findings suggest that GPΔMLD presented through retroVLPs may provide a strategy for development of a vaccine against multiple ebolaviruses. Similar vaccination strategies may be adopted for other viruses whose envelope proteins contain highly variable regions that may mask more conserved domains from the immune system.

Role for a Zinc Finger Protein (Zfp111) in Transformation of 208F Rat Fibroblasts by Jaagsiekte Sheep Retrovirus Envelope Protein

PubMed Central

Hsu, Tom; Phung, An; Choe, Kevin; Kim, Jung Woo

2015-01-01

ABSTRACT The native envelope gene (env) of Jaagsiekte sheep retrovirus (JSRV) also acts as an oncogene. To investigate the mechanism of transformation, we performed yeast 2-hybrid screening for cellular proteins that interact with Env. Among several candidates, we identified mouse or rat zinc finger protein 111 (zfp111). The interaction between Env and Zfp111 was confirmed through in vivo coimmunoprecipitation assays. Knockdown of endogenous Zfp111 caused a decrease in cell transformation by JSRV Env, while overexpression of Zfp111 increased overall Env transformation, supporting a role for Zfp111 in Env transformation. Knockdown of Zfp111 had no effect on the growth rate of parental rat 208F cells, while it decreased the proliferation rate of JSRV-transformed 208F cells, suggesting that JSRV-transformed cells became dependent on Zfp111. In addition, Zfp111 preferentially bound to a higher-mobility form of JSRV Env that has not been described previously. The higher-mobility form of Env (P70env) was found exclusively in the nuclear fraction, and size of its polypeptide backbone was the same as that of the cytoplasmic Env polyprotein (Pr80env). The differences in glycosylation between the two versions of Env were characterized. These results identify a novel cellular protein, Zfp111, that binds to the JSRV Env protein, and this binding plays a role in Env transformation. These results indicate that JSRV transformation also involves proteins and interactions in the nucleus. IMPORTANCE The envelope protein (Env) of Jaagsiekte sheep retrovirus (JSRV) is an oncogene, but its mechanism of cell transformation is still unclear. Here we identified seven candidate cellular proteins that can interact with JSRV Env by yeast two-hybrid screening. This study focused on one of the seven candidates, zinc finger protein 111 (Zfp111). Zfp111 was shown to interact with JSRV Env in cells and to be involved in JSRV transformation. Moreover, coexpression of JSRV Env and Zfp111 led to the

Expression of the fusogenic HERV-FRD Env glycoprotein (syncytin 2) in human placenta is restricted to villous cytotrophoblastic cells.

PubMed

Malassiné, A; Blaise, S; Handschuh, K; Lalucque, H; Dupressoir, A; Evain-Brion, D; Heidmann, T

2007-01-01

Recently, the expression of a human endogenous retrovirus HERV-FRD, able to encode a fusogenic envelope protein (syncytin 2), has been observed in human placenta. The aim of the present study was to localize the expression of syncytin 2 in first trimester placenta. In addition, we investigated the presence of HERV-FRD transcripts during the in vitro differentiation of isolated villous and extravillous trophoblastic cells from first trimester chorionic villi. Using a monoclonal antibody specifically raised against the HERV-FRD Env protein, syncytin 2 was immunolocalized only in the villous trophoblast of the chorionic villi, at the level of cytotrophoblastic cells. Interestingly, immunostaining was not observed in all cells but only in some of them, and was detected, more frequently, at the membrane level at the interface between the cytotrophoblastic cells and syncytiotrophoblast. Labeling was observed neither in the syncytiotrophoblast nor in the mesenchymal core of the villi nor in the extravillous trophoblast. In vitro detection of HERV-FRD transcripts was restricted to villous trophoblastic cells and decreased significantly with time in culture. These results suggest that syncytin 2 might play a role in human trophoblastic cell fusion.

Exosomes carring gag/env of ALV-J possess negative effect on immunocytes.

PubMed

Wang, Guihua; Wang, Zhenzhen; Zhuang, Pingping; Zhao, Xiaomin; Cheng, Ziqiang

2017-11-01

J subgroup avian leukosis virus (ALV-J) is an exogenous retrovirus of avian. A key feature of ALV-J infection is leading to severe immunosuppressive characteristic of diseases. Viral components of retrovirus were reported closely associated with immunosuppression, and several similarities between exosomes and retrovirus preparations have lead to the hypotheses of retrovirus hijacker exosomes pathway. In this study, we purified exosomes from DF-1 cells infected and uninfected by ALV-J. Electron microscopy and mass spectrometry (MS) analysis showed that ALV-J not only increased the production of exosomes from ALV-J infected DF-1 cells (Exo-J) but also stimulated some proteins expression, especially ALV-J components secreted in exosomes. Immunosuppressive domain peptide (ISD) of envelope subunit transmembrane (TM) and gag of ALV-J were secreted in Exo-J. It has been reported that HIV gag was budded from endosome-like domains of the T cell plasma membrane. But env protein was first detected in exosomes from retrovirus infected cells. We found that Exo-J caused negative effects on splenocytes in a dose-dependant manner by flow cytometric analysis. And low dose of Exo-J activated immune activity of splenocytes, while high dose possessed immunosuppressive properties. Interestingly, Exo-J has no significant effects on the immunosuppression induced by ALV-J, and the immunosuppressive effects induced by Exo-J lower than that by ALV-J. Taken together, our data indicated that Exo-J supplied a microenvironment for the replication and transformation of ALV-J. Copyright © 2017. Published by Elsevier Ltd.

Aberrant expression of liver microRNA in chickens infected with subgroup J avian leukosis virus

USDA-ARS?s Scientific Manuscript database

Subgroup J avian leukosis virus (ALV-J) is an oncogenic retrovirus primarily causing myeloid leukosis (ML) in broilers. Although ALV is well under control in a few countries including the U.S.A., poultry industry in many parts of the world continues suffering from serious economic loss due to sporad...

Hepa1-6-FLuc cell line with the stable expression of firefly luciferase retains its primary properties with promising bioluminescence imaging ability.

PubMed

Li, Yasha; Liu, Mengnan; Cui, Jiejie; Yang, Ke; Zhao, Li; Gong, Mengjia; Wang, Yi; He, Yun; He, Tongchuan; Bi, Yang

2018-05-01

Reliable animal models are required for the in vivo study of the molecular mechanisms and effects of chemotherapeutic drugs in hepatocarcinoma. In vivo tracing techniques based on firefly luciferase (FLuc) may optimize the non-invasive monitoring of experimental animals. The present study established a murine Hepa1-6-FLuc cell line that stably expressed a retrovirus-delivered FLuc protein gene. The cell morphology, proliferation, migration and invasion ability of Hepa1-6-FLuc cells were the same as that of the Hepa1-6 cells, and thus is suitable to replace Hepa1-6 cells in the construction of hepatocarcinoma animal models. No differences in subcutaneous tumor mass and its pathomorphology from implanted Hepa1-6-FLuc cells were observed compared with Hepa1-6 control tumors. Bioluminescence imaging indicated that the Luc signal of the Hepa1-6-FLuc cells was consistently strengthened with increases in tumor mass; however, the Luc signal of Hepa1-6-AdFLuc became weaker and eventually disappeared during tumor development. Therefore, compared with the transient expression by adenovirus, stable expression of the FLuc gene in Hepa1-6 cells may better reflect cell proliferation and survival in vivo , and provide a reliable source for the establishment of hepatocarcinoma models.

Comparative Methylation of ERVWE1/Syncytin-1 and Other Human Endogenous Retrovirus LTRs in Placenta Tissues

PubMed Central

Gimenez, Juliette; Montgiraud, Cécile; Oriol, Guy; Pichon, Jean-Philippe; Ruel, Karine; Tsatsaris, Vassilis; Gerbaud, Pascale; Frendo, Jean-Louis; Evain-Brion, Danièle; Mallet, François

2009-01-01

Human endogenous retroviruses (HERVs) are globally silent in somatic cells. However, some HERVs display high transcription in physiological conditions. In particular, ERVWE1, ERVFRDE1 and ERV3, three proviruses of distinct families, are highly transcribed in placenta and produce envelope proteins associated with placenta development. As silencing of repeated elements is thought to occur mainly by DNA methylation, we compared the methylation of ERVWE1 and related HERVs to appreciate whether HERV methylation relies upon the family, the integration site, the tissue, the long terminal repeat (LTR) function or the associated gene function. CpG methylation of HERV-W LTRs in placenta-associated tissues was heterogeneous but a joint epigenetic control was found for ERVWE1 5′LTR and its juxtaposed enhancer, a mammalian apparent LTR retrotransposon. Additionally, ERVWE1, ERVFRDE1 and ERV3 5′LTRs were all essentially hypomethylated in cytotrophoblasts during pregnancy, but showed distinct and stage-dependent methylation profiles. In non-cytotrophoblastic cells, they also exhibited different methylation profiles, compatible with their respective transcriptional activities. Comparative analyses of transcriptional activity and LTR methylation in cell lines further sustained a role for methylation in the control of functional LTRs. These results suggest that HERV methylation might not be family related but copy-specific, and related to the LTR function and the tissue. In particular, ERVWE1 and ERV3 could be developmentally epigenetically regulated HERVs. PMID:19561344

Unique Resistance of I/LnJ Mice to a Retrovirus Is Due to Sustained Interferon γ–dependent Production of Virus-neutralizing Antibodies

PubMed Central

Purdy, Alexandra; Case, Laure; Duvall, Melody; Overstrom-Coleman, Max; Monnier, Nilah; Chervonsky, Alexander; Golovkina, Tatyana

2003-01-01

Selection of immune escape variants impairs the ability of the immune system to sustain an efficient antiviral response and to control retroviral infections. Like other retroviruses, mouse mammary tumor virus (MMTV) is not efficiently eliminated by the immune system of susceptible mice. In contrast, MMTV-infected I/LnJ mice are capable of producing IgG2a virus-neutralizing antibodies, sustain this response throughout their life, and secrete antibody-coated virions into the milk, thereby preventing infection of their progeny. Antibodies were produced in response to several MMTV variants and were cross-reactive to them. Resistance to MMTV infection was recessive and was dependent on interferon (IFN)-γ production, because I/LnJ mice with targeted deletion of the INF-γ gene failed to produce any virus-neutralizing antibodies. These findings reveal a novel mechanism of resistance to retroviral infection that is based on a robust and sustained IFN-γ–dependent humoral immune response. PMID:12538662

Evaluation of HTLV-1 activity in HAM/TSP patients using proviral load and Tax mRNA expression after In Vitro lymphocyte activation.

PubMed

Yari, Atefeh; Rezaee, Seyyed Abdolrahim; Valizadeh, Narges; Rajaee, Taraneh; Jazayeri, Seyyed Mohammad; Soltani, Mojdeh; Norouzi, Mehdi

2014-07-01

HTLV-1 is the first human retrovirus that has been recognized and is associated with HAM/TSP and ATLL. Studies have shown that less than five percent of HTLV-1 infected carriers develop HAM/TSP or ATLL and about ninety-five percent remain asymptomatic. Therefore, the proviral load with Tax may affect cellular genes such as cytokines and oncogenes, as well as involve in pathogenicity. Thirty HAM/TSP patients, thirty HTLV-1 healthy carriers, and MT-2 cell line were evaluated for HTLV-1 activity. PBMCs were isolated and activated using PMA and ionomycine. Real-time PCR and TaqMan methods were performed using specific primers and fluorescence probes for Tax expression and proviral load assessment. B2microglobulin (β2m) and albumin were used as controls in Tax expression and in proviral load, respectively. An insignificant increase in Tax expression was observed in rest PBMCs of HAM/TSP patients compared to healthy carriers. However, after lymphocyte activation there was a significant increase in Tax expression in HAM/TSP patients (P=0.042). The Proviral load in patients was significantly higher than in carriers. Moreover, there was a significant correlation between Tax mRNA expression in activated PBMCs and proviral load (R=0.37, P=0.012). Although proviral load had been addressed as a valuable index for monitoring HTLV-1 infected subjects, the results of this study demonstrated that Tax expression in activated PBMCs along with proviral load assessment in HAM/TSP patients are a more reliable factor for determining the prognosis and monitoring healthy carriers and HAM/TSP patients.

Functional characterization of CXCR4 in mediating the expression of protein C system in experimental ulcerative colitis

PubMed Central

Lin, Xuhong; Wang, Huichao; Li, Yuxia; Yang, Jingnan; Yang, Ruilin; Wei, Dandan; Zhang, Junjie; Yang, Desheng; Wang, Bin; Ren, Xuequn; Cheng, Guanchang

2017-01-01

The present study aimed to explore the role of CXCR4 and protein C system (PCS) in the experimental ulcerative colitis (UC). The expression of CXCR3, CCR10, and CXCR4 in dextran sulfate sodium (DSS)-induced colitis mouse model was measured by immunohistochemistry and western blot analysis. In vitro studies with microvascular endothelial cells (MVECs) were performed. The expression of endothelial protein C receptor (EPCR) and thrombomodulin (TM) were detected by RT-PCR and western blot analysis. Activities of protein C (PC), protein S (PS), activated PC (APC) were evaluated in cells pre-treated with JNK inhibitor SP600125 and c-Jun silencing. DSS mice showed up-regulated expression of CXCR4, higher macroscopic score and histological score (P<0.05), as well as elevated levels of SDF-1α (P<0.05) compared with wild type, CXCR4-/-, or CXCR4-/- +DSS mice. In DSS mice, EPCR expression was down-regulated (P<0.05), accompanied by decreased activity of PC and PS (P<0.05 or P<0.01) with an up-regulated expression of pJNK MAPK and pc-Jun (P<0.05). Moreover, the macroscopic score and histological score index, SDF-1α levels, EPCR expression, PC activity, pJNK, and pc-Jun were reversed in CXCR4-/- +DSS mice (P<0.05). In vitro, SDF-1α-induced inhibition of the PCS was blunted by SP600125 (P<0.05). Meanwhile, down-regulation of c-Jun rescued the inhibition of PCS (P<0.05). MVECs with retrovirus-mediated transfection of c-Jun demonstrated a strong trans-inactivation effect on the EPCR promoter (P<0.05). These findings suggest that CXCR4 is involved in UC pathogenesis and could be a promising therapeutic target for UC treatment. PMID:29218082

Induction of ebolavirus cross-species immunity using retrovirus-like particles bearing the Ebola virus glycoprotein lacking the mucin-like domain

PubMed Central

2012-01-01

Background The genus Ebolavirus includes five distinct viruses. Four of these viruses cause hemorrhagic fever in humans. Currently there are no licensed vaccines for any of them; however, several vaccines are under development. Ebola virus envelope glycoprotein (GP1,2) is highly immunogenic, but antibodies frequently arise against its least conserved mucin-like domain (MLD). We hypothesized that immunization with MLD-deleted GP1,2 (GPΔMLD) would induce cross-species immunity by making more conserved regions accessible to the immune system. Methods To test this hypothesis, mice were immunized with retrovirus-like particles (retroVLPs) bearing Ebola virus GPΔMLD, DNA plasmids (plasmo-retroVLP) that can produce such retroVLPs in vivo, or plasmo-retroVLP followed by retroVLPs. Results Cross-species neutralizing antibody and GP1,2-specific cellular immune responses were successfully induced. Conclusion Our findings suggest that GPΔMLD presented through retroVLPs may provide a strategy for development of a vaccine against multiple ebolaviruses. Similar vaccination strategies may be adopted for other viruses whose envelope proteins contain highly variable regions that may mask more conserved domains from the immune system. PMID:22273269

Human retroviruses in leukaemia and AIDS: reflections on their discovery, biology and epidemiology.

PubMed

Karpas, Abraham

2004-11-01

The study of retroviruses has had a profound impact by unveiling an unusual form of viral replication: the multiplication of RNA viruses via a proviral DNA, for which Jan Svoboda provided the experimental model over forty years ago. In 1970 Temin, Mizutani and Baltimore discovered that this group of viruses contains a unique enzyme catalysing the synthesis of a DNA copy of the viral RNA: reverse transcriptase (RT). The discovery of RT has itself had an enormous impact on molecular biology in general, but also stimulated many premature claims of its detection in human disease. Claims by Gallo's laboratory that the cytoplasm of human leukaemia cells contained RT proved to be unfounded, as did his report in collaboration with Weiss that myeloid leukaemia contained HL23 virus, this organism proving not to be human but a laboratory contaminant of three monkey viruses. Conclusive demonstration of a retroviral involvement in human leukaemia was first provided in 1981 by Hinuma and his associates, showing that adult T-cell leukaemia (ATL), a rare form of leukaemia endemic to south-west Japan, is caused by a new retrovirus (ATLV). Other publications in December 1980 and through 1981 claimed the discovery of a new human T-cell leukaemia virus involved in mycosis fungoides (MF) and Sézary's syndrome (SS). This virus was termed HTLV by Gallo. The nucleotide sequence of ATLV is strongly conserved, that of my 1983 isolate from a black British ATL patient being practically identical with the Japanese virus isolates. After AIDS was recognised in 1981 by Gottlieb and coworkers as a new human disease, several papers were published by Gallo and his associates during 1983-4, invoking the oncovirus responsible for adult T-cell leukaemia as the cause of AIDS. In 1983 the French scientist Barré-Sinoussi and her colleagues succeeded in isolating a new agent in the disease, a lentivirus, which they named LAV. The French immunologist Klatzmann and his colleagues discovered that LAV killed

Enhanced retroviral gene delivery in ultrasonic standing wave fields.

PubMed

Lee, Y-H; Peng, C-A

2005-04-01

Enhancement of retroviral transduction efficiency has been achieved by several physical and chemical approaches. However, the application of those methods is hampered by not easily scalable configurations. In this study, instead of looking into the effect of sonoporation, the potential of ultrasonic standing wave fields (USWF) to facilitate retroviral transduction rate was explored. We reasoned that, driven by the primary acoustic radiation force, suspended cells moved to the pressure nodal planes first and formed cell bands. Nanometer-sized retroviruses, circulated between nodal planes by acoustic microstreaming, then used the preformed cell bands as the nucleating sites to attach on. As a result, the encounter opportunity between retroviruses and cells was increased and further facilitated the gene delivery efficiency. Our results showed that mega-Hertz USWF brought K562 erythroleukemia cells (10(6) cells/ml) and vesicular stomatitis virus G-protein (VSV-G) pseudotyped retroviruses (titer of 5 x 10(6) CFU/ml) into close contact at the pressure nodal planes, yielding a four-fold increment of enhanced green fluorescent protein transgene expression after 5-min USWF exposure in the presence of Polybrene. Furthermore, with a fixed titer of retrovirus, the transduction rate was augmented with the increase of cell concentration. In summary, USWF offer a feasible means to enhance retroviral transduction efficiency in large-scale settings.

Transgenic chickens expressing human urokinase-type plasminogen activator.

PubMed

Lee, Sung Ho; Gupta, Mukesh Kumar; Ho, Young Tae; Kim, Teoan; Lee, Hoon Taek

2013-09-01

Urokinase-type plasminogen activator is a serine protease that is clinically used in humans for the treatment of thrombolytic disorders and vascular diseases such as acute ischemic stroke and acute peripheral arterial occlusion. This study explored the feasibility of using chickens as a bioreactor for producing human urokinase-type plasminogen activator (huPA). Recombinant huPA gene, under the control of a ubiquitous Rous sarcoma virus promoter, was injected into the subgerminal cavity of freshly laid chicken eggs at stage X using the replication-defective Moloney murine leukemia virus (MoMLV)-based retrovirus vectors encapsidated with VSV-G (vesicular stomatitis virus G) glycoprotein. A total of 38 chicks, out of 573 virus-injected eggs, hatched and contained the huPA gene in their various body parts. The mRNA transcript of the huPA gene was present in various organs, including blood and egg, and was germ-line transmitted to the next generation. The level of active huPA protein was 16-fold higher in the blood of the transgenic chicken than in the nontransgenic chicken (P < 0.05). The expression of huPA protein in eggs increased from 7.82 IU/egg in the G0 generation to 17.02 IU/egg in the G1 generation. However, huPA-expressing embryos had reduced survival and hatchability at d 18 and 21 of incubation, respectively, and the blood clotting time was significantly higher in transgenic chickens than their nontransgenic counterparts (P < 0.05). Furthermore, adult transgenic rooster showed reduced (P < 0.05) fertility, as revealed by reduced volume of semen ejaculate, sperm concentration, and sperm viability. Taken together, our data suggest that huPA transgenic chickens could be successfully produced by the retroviral vector system. Transgenic chickens, expressing the huPA under the control of a ubiquitous promoter, may not only be used as a bioreactor for pharming of the huPA drug but also be useful for studying huPA-induced bleeding and other disorders.

The Influences of Glycosylation on the Antigenicity, Immunogenicity, and Protective Efficacy of Ebola Virus GP DNA Vaccines

DTIC Science & Technology

2006-11-22

multiple muta- tions were not studied, (iii) a vaccinia virus (VACV)- T7 system was used for transient expression, (iv) pseudotyped retrovi- ruses were used...those studies produced little to no detectable GP1 or GP2 in the transient VACV- T7 expression assays, whereas in our studies with the DNA con- structs...type GP2 was detected in pseudotyped retroviruses, a result seemingly in conflict with these authors’ findings with the VACV- T7 expression. Although

Human endogenous retrovirus type W envelope expression in blood and brain cells provides new insights into multiple sclerosis disease

PubMed Central

Germi, Raphaëlle; Bernard, Corinne; Garcia-Montojo, Marta; Deluen, Cécile; Farinelli, Laurent; Faucard, Raphaël; Veas, Francisco; Stefas, Ilias; Fabriek, Babs O; Van-Horssen, Jack; Van-der-Valk, Paul; Gerdil, Claire; Mancuso, Roberta; Saresella, Marina; Clerici, Mario; Marcel, Sébastien; Creange, Alain; Cavaretta, Rosella; Caputo, Domenico; Arru, Giannina; Morand, Patrice; Lang, Alois B; Sotgiu, Stefano; Ruprecht, Klemens; Rieckmann, Peter; Villoslada, Pablo; Chofflon, Michel; Boucraut, Jose; Pelletier, Jean; Hartung, Hans-Peter

2012-01-01

Background: The envelope protein from multiple sclerosis (MS) associated retroviral element (MSRV), a member of the Human Endogenous Retroviral family ‘W’ (HERV-W), induces dysimmunity and inflammation. Objective: The objective of this study was to confirm and specify the association between HERV-W/MSRV envelope (Env) expression and MS. Methods: 103 MS, 199 healthy controls (HC) and controls with other neurological diseases (28), chronic infections (30) or autoimmunity (30) were analysed with an immunoassay detecting Env in serum. Env RNA or DNA copy numbers in peripheral blood mononuclear cells (PBMC) were determined by a quantitative polymerase chain reaction (PCR). Env was detected by immunohistology in the brains of patients with MS with three specific monoclonals. Results: Env antigen was detected in a serum of 73% of patients with MS with similar prevalence in all clinical forms, and not in chronic infection, systemic lupus, most other neurological diseases and healthy donors (p<0.01). Cases with chronic inflammatory demyelinating polyneuropathy (5/8) and rare HC (4/103) were positive. RNA expression in PBMC and DNA copy numbers were significantly elevated in patients with MS versus HC (p<0.001). In patients with MS, DNA copy numbers were significantly increased in chronic progressive MS (secondary progressive MS vs relapsing–remitting MS (RRMS) p<0.001; primary progressive MS vs RRMS –<0.02). Env protein was evidenced in macrophages within MS brain lesions with particular concentrations around vascular elements. Conclusion: The association between MS disease and the MSRV-type HERV-W element now appears quite strong, as evidenced ex-vivo from serum and PBMC with post-mortem confirmation in brain lesions. Chronic progressive MS, RRMS and clinically isolated syndrome show different ELISA (Enzyme-Linked Immunosorbent Assay) and/or PCR profiles suggestive of an increase with disease evolution, and amplicon sequencing confirms the association with

Characterization of the gag/fusion protein encoded by the defective Duplan retrovirus inducing murine acquired immunodeficiency syndrome.

PubMed Central

Huang, M; Jolicoeur, P

1990-01-01

Murine acquired immunodeficiency syndrome is induced by a defective retrovirus. Sequencing of this defective viral genome revealed a long open reading frame which encodes a putative gag/fusion protein, N-MA-p12-CA-NC-COOH, (D. C. Aziz, Z. Hanna, and P. Jolicoeur, Nature (London) 338:505-508, 1989). We raised a specific antibody to the unique p12 domain of this gag fusion precursor, Pr60gag. We found that Pr60gag was indeed encoded by the defective viral genome both in cell-free translation reticulocyte extracts and in infected mouse fibroblasts. Pr60gag was found to be myristylated, phosphorylated, and attached to the cell membrane, like other helper murine leukemia virus (MuLV) gag precursors. Pr60gag was not substantially cleaved within the nonproducer cells and was not released from these cells. However, in the presence of helper MuLV proteins, it formed phenotypically mixed particles. In these particles, Pr60gag was only partially cleaved. In helper MuLV-producing cells harboring the defective virus, a gag-related p40 intermediate was generated both intracellularly and extracellularly. In these cells, Pr60gag appeared to behave as a dominant negative mutant, interfering with proper cleavage of helper Pr65gag. Our data indicate that Pr60gag is a major (and possibly the only) gene product of the defective murine acquired immunodeficiency syndrome virus and is likely to harbor some determinants of pathogenicity of this virus. Images PMID:2243376

A systematic evaluation of expression of HERV-W elements; influence of genomic context, viral structure and orientation

PubMed Central

2011-01-01

Background One member of the W family of human endogenous retroviruses (HERV) appears to have been functionally adopted by the human host. Nevertheless, a highly diversified and regulated transcription from a range of HERV-W elements has been observed in human tissues and cells. Aberrant expression of members of this family has also been associated with human disease such as multiple sclerosis (MS) and schizophrenia. It is not known whether this broad expression of HERV-W elements represents transcriptional leakage or specific transcription initiated from the retroviral promoter in the long terminal repeat (LTR) region. Therefore, potential influences of genomic context, structure and orientation on the expression levels of individual HERV-W elements in normal human tissues were systematically investigated. Results Whereas intronic HERV-W elements with a pseudogene structure exhibited a strong anti-sense orientation bias, intronic elements with a proviral structure and solo LTRs did not. Although a highly variable expression across tissues and elements was observed, systematic effects of context, structure and orientation were also observed. Elements located in intronic regions appeared to be expressed at higher levels than elements located in intergenic regions. Intronic elements with proviral structures were expressed at higher levels than those elements bearing hallmarks of processed pseudogenes or solo LTRs. Relative to their corresponding genes, intronic elements integrated on the sense strand appeared to be transcribed at higher levels than those integrated on the anti-sense strand. Moreover, the expression of proviral elements appeared to be independent from that of their corresponding genes. Conclusions Intronic HERV-W provirus integrations on the sense strand appear to have elicited a weaker negative selection than pseudogene integrations of transcripts from such elements. Our current findings suggest that the previously observed diversified and tissue

Identification of aberrantly expressed circRNAs in subgroup J avian leucosis virus induced tumor livers by RNA sequencing

USDA-ARS?s Scientific Manuscript database

ALV-J (subgroup J avian leucosis virus) is a kind of oncogenic exogenous retrovirus and diseases associated with ALV-J have caused severe reproduction problems in the poultry industry worldwide. However, the pathogenesis of ALV-J-induced tumor formation is still unclear. In recent years, circRNAs ar...

Retrotransposon expression and incorporation of cloned human and mouse retroelements in human spermatozoa.

PubMed

Lazaros, Leandros; Kitsou, Chrysoula; Kostoulas, Charilaos; Bellou, Sofia; Hatzi, Elissavet; Ladias, Paris; Stefos, Theodoros; Markoula, Sofia; Galani, Vasiliki; Vartholomatos, Georgios; Tzavaras, Theodore; Georgiou, Ioannis

2017-03-01

To investigate the expression of long interspersed element (LINE) 1, human endogenous retrovirus (HERV) K10, and short interspersed element-VNTR-Alu element (SVA) retrotransposons in ejaculated human spermatozoa by means of reverse-transcription (RT) polymerase chain reaction (PCR) analysis as well as the potential incorporation of cloned human and mouse active retroelements in human sperm cell genome. Laboratory study. University research laboratories and academic hospital. Normozoospermic and oligozoospermic white men. RT-PCR analysis was performed to confirm the retrotransposon expression in human spermatozoa. Exogenous retroelements were tagged with a plasmid containing a green fluorescence (EGFP) retrotransposition cassette, and the de novo retrotransposition events were tested with the use of PCR, fluorescence-activated cell sorting analysis, and confocal microscopy. Retroelement expression in human spermatozoa, incorporation of cloned human and mouse active retroelements in human sperm genome, and de novo retrotransposition events in human spermatozoa. RT-PCR products of expressed human LINE-1, HERV-K10, and SVA retrotransposons were observed in ejaculated human sperm samples. The incubation of human spermatozoa with either retrotransposition-active human LINE-1 and HERV-K10 or mouse reverse transcriptase-deficient VL30 retrotransposons tagged with an EGFP-based retrotransposition cassette led to EGFP-positive spermatozo; 16.67% of the samples were positive for retrotransposition. The respective retrotransposition frequencies for the LINE-1, HERV-K10, and VL30 retrotransposons in the positive samples were 0.34 ± 0.13%, 0.37 ± 0.17%, and 0.30 ± 0.14% per sample of 10,000 spermatozoa. Our results show that: 1) LINE-1, HERV-K10, and SVA retrotransposons are transcriptionally expressed in human spermatozoa; 2) cloned active retroelements of human and mammalian origin can be incorporated in human sperm genome; 3) active reverse transcriptases exist in human

[Functional analysis of Grp and Iris, the gag and env domesticated errantivirus genes, in the Drosophila melanogaster genome].

PubMed

Makhnovskii, P A; Kuzmin, I V; Nefedova, L N; Kima, A I

2016-01-01

Drosophila melanogaster is the only invertebrate that contains endogenous retroviruses, which are called errantiviruses. Two domesticated genes, Grp and Iris, which originate from errantivirus gag and env, respectively, have been found in the D. melanogaster genome. The functions performed by the genes in Drosophila are still unclear. To identify the functions of domesticated gag and env in the D. melanogaster genome, expression of Iris and Grp was studied in strains differing by the presence or absence of the functional gypsy errantivirus. In addition, the expression levels were measured after injection of gram-positive and gram-negative bacteria, which activate different immune response pathways, and exposure to various abiotic stress factors. The presence of functional D. melanogaster retrovirus gypsy was found to increase the Grp expression level in somatic tissues of the carcass, while exerting no effect on the Iris expression level. Activation of the immune response in D. melanogaster by bacteria Bacillus cereus increased the Grp expression level and did not affect Iris expression. As for the effects of abiotic stress factors (oxidative stress, starvation, and heat and cold stress), the Grp expression level increased in response to starvation in D. melanogaster females, and the Iris expression level was downregulated in heat shock and oxidative stress. Based on the findings, Grp was assumed to play a direct role in the immune response in D. melanogaster; Iris is not involved in immune responses, but and apparently performs a cell function that is inhibited in stress.

Characterization of retrovirus-based reporter viruses pseudotyped with the precursor membrane and envelope glycoproteins of four serotypes of dengue viruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hu, H.-P.; Hsieh, S.-C.; King, C.-C.

In this study, we successfully established retrovirus-based reporter viruses pseudotyped with the precursor membrane and envelope (PrM/E) proteins of each of the four serotypes of dengue viruses, which caused the most important arboviral diseases in this century. Co-sedimentation of the dengue E protein and HIV-1 core proteins by sucrose gradient analysis of the pseudotype reporter virus of dengue virus type 2, D2(HIVluc), and detection of HIV-1 core proteins by immunoprecipitation with anti-E monoclonal antibody suggested that dengue viral proteins were incorporated into the pseudotype viral particles. The infectivity in target cells, as assessed by the luciferase activity, can be inhibitedmore » by the lysosomotropic agents, suggesting a pH-dependent mechanism of entry. Amino acid substitutions of the leucine at position 107, a critical residue at the fusion loop of E protein, with lysine resulted in severe impairment in infectivity, suggesting that entry of the pseudotype reporter virus is mediated through the fusogenic properties of E protein. With more and more dengue viral sequences available from different outbreaks worldwide, this sensitive and convenient tool has the potential to facilitate molecular characterization of the PrM/E proteins of dengue field isolates.« less

p57KIP2 expression and loss of heterozygosity during immortal conversion of cultured human mammary epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nijjar, Tarlochan; Wigington, Don; Garbe, James C.

1999-08-01

The authors have uncovered a novel role for the cyclin-dependent kinase inhibitor, p57KIP2, during the immortalization of cultured human mammary epithelial cells (HMEC). HMEC immortalized following chemical carcinogen exposure initially expressed little or no telomerase activity, and their telomeres continued to shorten with passage. Cell populations whose mean terminal restriction fragment (TRF) length declined and exhibited slow heterogeneous growth, and contained many non-proliferative cells. These conditionally immortal HMEC cultures accumulated large quantities of p57 protein. With continued passage, the conditionally immortal cell populations very graduall2048nverted to a fully immortal phenotype of good uniform growth, expression of high levels of telomerasemore » activity, and stabilization of telomere length. The fully immortal good growing HMEC did not accumulate p57 in G0 or during the cell cycle. DNA and RNA analysis of mass populations and individual subclones of conditionally immortal HMEC line 184A1 showed that continued growth of conditionally immortal cells with critically short telomeres was repeatedly accompanied by loss of the expressed p57 allele, and transient expression of the previously imprinted allele. Conditionally immortal 184A1 with mean TRF > 3 kb infected with retroviruses containing the p57 gene exhibited premature slow heterogeneous growth. Conversely, exogenous expression of hTERT, the catalytic subunit of telomerase, in 184A1 with mean TRF > 3 kb prevented both the slow heterogeneous growth phase and accumulation of p57 in cycling populations. These data indicate that in HMEC which have overcome replicative senescence, p57 may provide an additional barrier against indefinite proliferation. Overcoming p57 mediated growth inhibition in these cells may be crucial for acquisition of the unlimited growth potential thought to be critical for malignant progression.« less

Human Immunodeficiency Virus Integration Protein Expressed in Escherichia Coli Possesses Selective DNA Cleaving Activity

NASA Astrophysics Data System (ADS)

Sherman, Paula A.; Fyfe, James A.

1990-07-01

The human immunodeficiency virus (HIV) integration protein, a potential target for selective antiviral therapy, was expressed in Escherichia coli. The purified protein, free of detectable contaminating endonucleases, selectively cleaved double-stranded DNA oligonucleotides that mimic the U3 and the U5 termini of linear HIV DNA. Two nucleotides were removed from the 3' ends of both the U5 plus strand and the U3 minus strand; in both cases, cleavage was adjacent to a conserved CA dinucleotide. The reaction was metal-ion dependent, with a preference for Mn2+ over Mg2+. Reaction selectivity was further demonstrated by the lack of cleavage of an HIV U5 substrate on the complementary (minus) strand, an analogous substrate that mimics the U3 terminus of an avian retrovirus, and an HIV U5 substrate in which the conserved CA dinucleotide was replaced with a TA dinucleotide. Such an integration protein-mediated cleavage reaction is expected to occur as part of the integration event in the retroviral life cycle, in which a double-stranded DNA copy of the viral RNA genome is inserted into the host cell DNA.

Extensive retroviral diversity in shark.

PubMed

Han, Guan-Zhu

2015-04-28

Retroviruses infect a wide range of vertebrates. However, little is known about the diversity of retroviruses in basal vertebrates. Endogenous retrovirus (ERV) provides a valuable resource to study the ecology and evolution of retrovirus. I performed a genome-scale screening for ERVs in the elephant shark (Callorhinchus milii) and identified three complete or nearly complete ERVs and many short ERV fragments. I designate these retroviral elements "C. milli ERVs" (CmiERVs). Phylogenetic analysis shows that the CmiERVs form three distinct lineages. The genome invasions by these retroviruses are estimated to take place more than 50 million years ago. My results reveal the extensive retroviral diversity in the elephant shark. Diverse retroviruses appear to have been associated with cartilaginous fishes for millions of years. These findings have important implications in understanding the diversity and evolution of retroviruses.

Contribution of the Retrovirus Epidemiology Donor Study (REDS) to research on blood transfusion safety in Brazil.

PubMed

Loureiro, Paula; de Almeida-Neto, Cesar; Proietti, Anna Bárbara Carneiro; Capuani, Ligia; Gonçalez, Thelma Terezinha; de Oliveira, Claudia Di Lorenzo; Leão, Silvana Carneiro; Lopes, Maria Inês; Sampaio, Divaldo; Patavino, Giuseppina Maria; Ferreira, João Eduardo; Blatyta, Paula Fraiman; Lopes, Maria Esther Duarte; Mendrone-Junior, Alfredo; Salles, Nanci Alves; King, Melissa; Murphy, Edward; Busch, Michael; Custer, Brian; Sabino, Ester Cerdeira

2014-03-01

The Retrovirus Epidemiology Donor Study (REDS) program was established in the United States in 1989 with the purpose of increasing blood transfusion safety in the context of the HIV/AIDS and human T-lymphotropic virus epidemics. REDS and its successor, REDS-II were at first conducted in the US, then expanded in 2006 to include international partnerships with Brazil and China. In 2011, a third wave of REDS renamed the Recipient Epidemiology and Donor Evaluation Study-III (REDS-III) was launched. This seven-year research program focuses on both blood banking and transfusion medicine research in the United States of America, Brazil, China, and South Africa. The main goal of the international programs is to reduce and prevent the transmission of HIV/AIDS and other known and emerging infectious agents through transfusion, and to address research questions aimed at understanding global issues related to the availability of safe blood. This article describes the contribution of REDS-II to transfusion safety in Brazil. Articles published from 2010 to 2013 are summarized, including database analyses to characterize blood donors, deferral rates, and prevalence, incidence and residual risk of the main blood-borne infections. Specific studies were developed to understand donor motivation, the impact of the deferral questions, risk factors and molecular surveillance among HIV-positive donors, and the natural history of Chagas disease. The purpose of this review is to disseminate the acquired knowledge and briefly summarize the findings of the REDS-II studies conducted in Brazil as well as to introduce the scope of the REDS-III program that is now in progress and will continue through 2018.

Transient Expression of an LEDGF/p75 Chimera Retargets Lentivector Integration and Functionally Rescues in a Model for X-CGD

PubMed Central

Vets, Sofie; De Rijck, Jan; Brendel, Christian; Grez, Manuel; Bushman, Frederic; Debyser, Zeger; Gijsbers, Rik

2013-01-01

Retrovirus-based vectors are commonly used as delivery vehicles to correct genetic diseases because of their ability to integrate new sequences stably. However, adverse events in which vector integration activates proto-oncogenes, leading to clonal expansion and leukemogenesis hamper their application. The host cell-encoded lens epithelium-derived growth factor (LEDGF/p75) binds lentiviral integrase and targets integration to active transcription units. We demonstrated earlier that replacing the LEDGF/p75 chromatin interaction domain with an alternative DNA-binding protein could retarget integration. Here, we show that transient expression of the chimeric protein using mRNA electroporation efficiently redirects lentiviral vector (LV) integration in wild-type (WT) cells. We then employed this technology in a model for X-linked chronic granulomatous disease (X-CGD) using myelomonocytic PLB-985 gp91−/− cells. Following electroporation with mRNA encoding the LEDGF-chimera, the cells were treated with a therapeutic lentivector encoding gp91phox. Integration site analysis revealed retargeted integration away from genes and towards heterochromatin-binding protein 1β (CBX1)-binding sites, in regions enriched in marks associated with gene silencing. Nevertheless, gp91phox expression was stable for at least 6 months after electroporation and NADPH-oxidase activity was restored to normal levels as determined by superoxide production. Together, these data provide proof-of-principle that transient expression of engineered LEDGF-chimera can retarget lentivector integration and rescues the disease phenotype in a cell model, opening perspectives for safer gene therapy. PMID:23462964

Long-term erythropoietin gene expression from transduced cells in bioisolator devices.

PubMed

Yanay, Ofer; Barry, Simon C; Flint, Lisa Y; Brzezinski, Margaret; Barton, Randall W; Osborne, William R A

2003-11-20

Recombinant erythropoietin (EPO) is widely administered for long-term treatment of anemia associated with renal failure and other chronic diseases. The ability to deliver EPO by gene therapy would have clinical and economic benefit. We compared autologous and allogeneic transduced primary vascular smooth muscle cells for their ability to provide sustained EPO gene expression when encapsulated in TheraCyte devices implanted subcutaneously (SQ) or intraperitoneally (IP) in rats. Cells were transduced with retrovirus vector LrEpSN encoding rat EPO cDNA. Rats that received either autologous or allogeneic transduced cells showed elevated hematocrits (HCTs) ranging from 50 to 79% that were sustained for more than 12 months. The HCT of control rats remained at baseline (45.8%). Rats that received second SQ implants of either autologous or allogeneic cells showed elevations in hematocrit that were sustained for up to 12 months, suggesting the absence of immunological responses to transduced cells or implant material. All experimental groups had statistically significant elevated HCT (p < 0.001) when compared with controls. Both SQ and IP implantation were equally effective in delivering EPO long term. There were no significant differences in white blood cell (WBC) or platelet (PLT) values between treated and control animals. Implantation of TheraCyte devices was well tolerated and histological evaluation of the devices up to 12 months after surgery revealed a high degree of vascularization and no evidence of host immune response. TheraCyte devices offer a simple and safe gene delivery system that provides sustained therapeutic gene expression, permit removal and implantation of new devices, and do not require immunosuppression of the host.

Retroviruses Hijack Chromatin Loops to Drive Oncogene Expression and Highlight the Chromatin Architecture around Proto-Oncogenic Loci

PubMed Central

Pattison, Jillian M.; Wright, Jason B.; Cole, Michael D.

2015-01-01

The majority of the genome consists of intergenic and non-coding DNA sequences shown to play a major role in different gene regulatory networks. However, the specific potency of these distal elements as well as how these regions exert function across large genomic distances remains unclear. To address these unresolved issues, we closely examined the chromatin architecture around proto-oncogenic loci in the mouse and human genomes to demonstrate a functional role for chromatin looping in distal gene regulation. Using cell culture models, we show that tumorigenic retroviral integration sites within the mouse genome occur near existing large chromatin loops and that this chromatin architecture is maintained within the human genome as well. Significantly, as mutagenesis screens are not feasible in humans, we demonstrate a way to leverage existing screens in mice to identify disease relevant human enhancers and expose novel disease mechanisms. For instance, we characterize the epigenetic landscape upstream of the human Cyclin D1 locus to find multiple distal interactions that contribute to the complex cis-regulation of this cell cycle gene. Furthermore, we characterize a novel distal interaction upstream of the Cyclin D1 gene which provides mechanistic evidence for the abundant overexpression of Cyclin D1 occurring in multiple myeloma cells harboring a pathogenic translocation event. Through use of mapped retroviral integrations and translocation breakpoints, our studies highlight the importance of chromatin looping in oncogene expression, elucidate the epigenetic mechanisms crucial for distal cis-regulation, and in one particular instance, explain how a translocation event drives tumorigenesis through upregulation of a proto-oncogene. PMID:25799187

Reciprocal relationship of T regulatory cells and monocytic myeloid-derived suppressor cells in LP-BM5 murine retrovirus-induced immunodeficiency.

PubMed

O'Connor, Megan A; Vella, Jennifer L; Green, William R

2016-02-01

Immunomodulatory cellular subsets, including myeloid-derived suppressor cells (MDSCs) and T regulatory cells (Tregs), contribute to the immunosuppressive tumour microenvironment and are targets of immunotherapy, but their role in retroviral-associated immunosuppression is less well understood. Due to known crosstalk between Tregs and MDSCs in the tumour microenvironment, and also their hypothesized involvement during human immunodeficiency virus/simian immunodeficiency virus infection, studying the interplay between these immune cells during LP-BM5 retrovirus-induced murine AIDS is of interest. IL-10-producing FoxP3+ Tregs expanded after LP-BM5 infection. Following in vivo adoptive transfer of natural Treg (nTreg)-depleted CD4+T-cells, and subsequent LP-BM5 retroviral infection, enriched monocytic MDSCs (M-MDSCs) from these nTreg-depleted mice displayed altered phenotypic subsets. In addition, M-MDSCs from LP-BM5-infected nTreg-depleted mice exhibited increased suppression of T-cell, but not B-cell, responses, compared with M-MDSCs derived from non-depleted LP-BM5-infected controls. Additionally, LP-BM5-induced M-MDSCs modulated the production of IL-10 by FoxP3+ Tregs in vitro. These collective data highlight in vitro and for the first time, to the best of our knowledge, in vivo reciprocal modulation between retroviral-induced M-MDSCs and Tregs, and may provide insight into the immunotherapeutic targeting of such regulatory cells during retroviral infection.

Molecular Determinants of Antiestrogen and Drug Sensitivity in Breast Carcinoma Cells

DTIC Science & Technology

1997-08-01

W.S. human HT1080 fibrosarcoma cells (a gift of Pear, personal communication). G.R. Stark), susceptible to ecotropic retrovirus In several...transfection with equal amounts of plasmid pOPRSVIluc (generated by replacing CAT of pORRSVICAT with luc) which expresses luc from R02 promoter, and control...retroviral transduction into (Fig. 1). During transient co-transfection of NIH 3T3 human HT1080 fibrosarcoma or mouse NIH 3T3 cell cells with a laac expressing

Genomic landscapes of endogenous retroviruses unveil intricate genetics of conventional and genetically-engineered laboratory mouse strains.

PubMed

Lee, Kang-Hoon; Lim, Debora; Chiu, Sophia; Greenhalgh, David; Cho, Kiho

2016-04-01

Laboratory strains of mice, both conventional and genetically engineered, have been introduced as critical components of a broad range of studies investigating normal and disease biology. Currently, the genetic identity of laboratory mice is primarily confirmed by surveying polymorphisms in selected sets of "conventional" genes and/or microsatellites in the absence of a single completely sequenced mouse genome. First, we examined variations in the genomic landscapes of transposable repetitive elements, named the TREome, in conventional and genetically engineered mouse strains using murine leukemia virus-type endogenous retroviruses (MLV-ERVs) as a probe. A survey of the genomes from 56 conventional strains revealed strain-specific TREome landscapes, and certain families (e.g., C57BL) of strains were discernible with defined patterns. Interestingly, the TREome landscapes of C3H/HeJ (toll-like receptor-4 [TLR4] mutant) inbred mice were different from its control C3H/HeOuJ (TLR4 wild-type) strain. In addition, a CD14 knock-out strain had a distinct TREome landscape compared to its control/backcross C57BL/6J strain. Second, an examination of superantigen (SAg, a "TREome gene") coding sequences of mouse mammary tumor virus-type ERVs in the genomes of the 46 conventional strains revealed a high diversity, suggesting a potential role of SAgs in strain-specific immune phenotypes. The findings from this study indicate that unexplored and intricate genomic variations exist in laboratory mouse strains, both conventional and genetically engineered. The TREome-based high-resolution genetics surveillance system for laboratory mice would contribute to efficient study design with quality control and accurate data interpretation. This genetics system can be easily adapted to other species ranging from plants to humans. Copyright © 2016 Elsevier Inc. All rights reserved.

Functional cis-regulatory modules encoded by mouse-specific endogenous retrovirus

PubMed Central

Sundaram, Vasavi; Choudhary, Mayank N. K.; Pehrsson, Erica; Xing, Xiaoyun; Fiore, Christopher; Pandey, Manishi; Maricque, Brett; Udawatta, Methma; Ngo, Duc; Chen, Yujie; Paguntalan, Asia; Ray, Tammy; Hughes, Ava; Cohen, Barak A.; Wang, Ting

2017-01-01

Cis-regulatory modules contain multiple transcription factor (TF)-binding sites and integrate the effects of each TF to control gene expression in specific cellular contexts. Transposable elements (TEs) are uniquely equipped to deposit their regulatory sequences across a genome, which could also contain cis-regulatory modules that coordinate the control of multiple genes with the same regulatory logic. We provide the first evidence of mouse-specific TEs that encode a module of TF-binding sites in mouse embryonic stem cells (ESCs). The majority (77%) of the individual TEs tested exhibited enhancer activity in mouse ESCs. By mutating individual TF-binding sites within the TE, we identified a module of TF-binding motifs that cooperatively enhanced gene expression. Interestingly, we also observed the same motif module in the in silico constructed ancestral TE that also acted cooperatively to enhance gene expression. Our results suggest that ancestral TE insertions might have brought in cis-regulatory modules into the mouse genome. PMID:28348391

Expression of an Exogenous Growth Hormone Gene by Transplantable Human Epidermal Cells

NASA Astrophysics Data System (ADS)

Morgan, Jeffrey R.; Barrandon, Yann; Green, Howard; Mulligan, Richard C.

1987-09-01

Retrovirus-mediated gene transfer was used to introduce a recombinant human growth hormone gene into cultured human keratinocytes. The transduced keratinocytes secreted biologically active growth hormone into the culture medium. When grafted as an epithelial sheet onto athymic mice, these cultured keratinocytes reconstituted an epidermis that was similar in appearance to that resulting from normal cells, but from which human growth hormone could be extracted. Transduced epidermal cells may prove to be a general vehicle for the delivery of gene products by means of grafting.

Human endogenous retrovirus HERV-K(HML-2) encodes a stable signal peptide with biological properties distinct from Rec

PubMed Central

Ruggieri, Alessia; Maldener, Esther; Sauter, Marlies; Mueller-Lantzsch, Nikolaus; Meese, Eckart; Fackler, Oliver T; Mayer, Jens

2009-01-01

Background The human endogenous retrovirus HERV-K(HML-2) family is associated with testicular germ cell tumors (GCT). Various HML-2 proviruses encode viral proteins such as Env and Rec. Results We describe here that HML-2 Env gives rise to a 13 kDa signal peptide (SP) that harbors a different C-terminus compared to Rec. Subsequent to guiding Env to the endoplasmatic reticulum (ER), HML-2 SP is released into the cytosol. Biochemical analysis and confocal microscopy demonstrated that similar to Rec, SP efficiently translocates to the granular component of nucleoli. Unlike Rec, SP does not shuttle between nucleus and cytoplasm. SP is less stable than Rec as it is subjected to proteasomal degradation. Moreover, SP lacks export activity towards HML-2 genomic RNA, the main function of Rec in the original viral context, and SP does not interfere with Rec's RNA export activity. Conclusion SP is a previously unrecognized HML-2 protein that, besides targeting and translocation of Env into the ER lumen, may exert biological functions distinct from Rec. HML-2 SP represents another functional similarity with the closely related Mouse Mammary Tumor Virus that encodes an Env-derived SP named p14. Our findings furthermore support the emerging concept of bioactive SPs as a conserved retroviral strategy to modulate their host cell environment, evidenced here by a "retroviral fossil". While the specific role of HML-2 SP remains to be elucidated in the context of human biology, we speculate that it may be involved in immune evasion of GCT cells or tumorigenesis. PMID:19220907

Advances in the study of transmissible respiratory tumours in small ruminants.

PubMed

Monot, M; Archer, F; Gomes, M; Mornex, J-F; Leroux, C

2015-12-14

Sheep and goats are widely infected by oncogenic retroviruses, namely Jaagsiekte Sheep RetroVirus (JSRV) and Enzootic Nasal Tumour Virus (ENTV). Under field conditions, these viruses induce transformation of differentiated epithelial cells in the lungs for Jaagsiekte Sheep RetroVirus or the nasal cavities for Enzootic Nasal Tumour Virus. As in other vertebrates, a family of endogenous retroviruses named endogenous Jaagsiekte Sheep RetroVirus (enJSRV) and closely related to exogenous Jaagsiekte Sheep RetroVirus is present in domestic and wild small ruminants. Interestingly, Jaagsiekte Sheep RetroVirus and Enzootic Nasal Tumour Virus are able to promote cell transformation, leading to cancer through their envelope glycoproteins. In vitro, it has been demonstrated that the envelope is able to deregulate some of the important signaling pathways that control cell proliferation. The role of the retroviral envelope in cell transformation has attracted considerable attention in the past years, but it appears to be highly dependent of the nature and origin of the cells used. Aside from its health impact in animals, it has been reported for many years that the Jaagsiekte Sheep RetroVirus-induced lung cancer is analogous to a rare, peculiar form of lung adenocarcinoma in humans, namely lepidic pulmonary adenocarcinoma. The implication of a retrovirus related to Jaagsiekte Sheep RetroVirus is still controversial and under investigation, but the identification of an infectious agent associated with the development of lepidic pulmonary adenocarcinomas might help us to understand cancer development. This review explores the mechanisms of induction of respiratory cancers in small ruminants and the possible link between retrovirus and lepidic pulmonary adenocarcinomas in humans. Copyright © 2015. Published by Elsevier B.V.

Evolution and Distribution of Class II-Related Endogenous Retroviruses†

PubMed Central

Gifford, Robert; Kabat, Peter; Martin, Joanne; Lynch, Clare; Tristem, Michael

2005-01-01

Endogenous retroviruses (ERVs) are widespread in vertebrate genomes and have been loosely grouped into “classes” on the basis of their phylogenetic relatedness to the established genera of exogenous retroviruses. Four of these genera—the lentiviruses, alpharetroviruses, betaretroviruses, and deltaretroviruses—form a well-supported clade in retroviral phylogenies, and ERVs that group with these genera have been termed class II ERVs. We used PCR amplification and sequencing of retroviral fragments from more than 130 vertebrate taxa to investigate the evolution of the class II retroviruses in detail. We confirm that class II retroviruses are largely confined to mammalian and avian hosts and provide evidence for a major novel group of avian retroviruses, and we identify additional members of both the alpha- and the betaretrovirus genera. Phylogenetic analyses demonstrated that the avian and mammalian viruses form distinct monophyletic groups, implying that interclass transmission has occurred only rarely during the evolution of the class II retroviruses. In contrast to previous reports, the lentiviruses clustered as sister taxa to several endogenous retroviruses derived from rodents and insectivores. This topology was further supported by the shared loss of both the class II PR-Pol frameshift site and the class II retrovirus G-patch domain. PMID:15858031

Transient Expression of an LEDGF/p75 Chimera Retargets Lentivector Integration and Functionally Rescues in a Model for X-CGD.

PubMed

Vets, Sofie; De Rijck, Jan; Brendel, Christian; Grez, Manuel; Bushman, Frederic; Debyser, Zeger; Gijsbers, Rik

2013-03-05

Retrovirus-based vectors are commonly used as delivery vehicles to correct genetic diseases because of their ability to integrate new sequences stably. However, adverse events in which vector integration activates proto-oncogenes, leading to clonal expansion and leukemogenesis hamper their application. The host cell-encoded lens epithelium-derived growth factor (LEDGF/p75) binds lentiviral integrase and targets integration to active transcription units. We demonstrated earlier that replacing the LEDGF/p75 chromatin interaction domain with an alternative DNA-binding protein could retarget integration. Here, we show that transient expression of the chimeric protein using mRNA electroporation efficiently redirects lentiviral vector (LV) integration in wild-type (WT) cells. We then employed this technology in a model for X-linked chronic granulomatous disease (X-CGD) using myelomonocytic PLB-985 gp91(-/-) cells. Following electroporation with mRNA encoding the LEDGF-chimera, the cells were treated with a therapeutic lentivector encoding gp91(phox). Integration site analysis revealed retargeted integration away from genes and towards heterochromatin-binding protein 1β (CBX1)-binding sites, in regions enriched in marks associated with gene silencing. Nevertheless, gp91(phox) expression was stable for at least 6 months after electroporation and NADPH-oxidase activity was restored to normal levels as determined by superoxide production. Together, these data provide proof-of-principle that transient expression of engineered LEDGF-chimera can retarget lentivector integration and rescues the disease phenotype in a cell model, opening perspectives for safer gene therapy.Molecular Therapy - Nucleic Acids (2013) 2, e77; doi:10.1038/mtna.2013.4; published online 5 March 2013.

Cooperative transforming activities of ras, myc, and src viral oncogenes in nonestablished rat adrenocortical cells.

PubMed Central

MacAuley, A; Pawson, T

1988-01-01

Early-passage rat adrenocortical cells were infected with Kirsten murine sarcoma virus and MMCV mouse myc virus, two retroviruses carrying the v-Ki-ras and v-myc oncogenes, respectively. Efficient morphological transformation required coinfection with the two viruses, was dependent on the presence of high serum concentrations, and was not immediately accompanied by growth in soft agar. The doubly infected cells coordinately acquired the capacity for anchorage- and serum-independent growth during passage in culture. The appearance of such highly transformed cells was correlated with the emergence of a dominant clone, as suggested by an analysis of retrovirus integration sites. These results indicate that the concerted expression of v-Ki-ras and v-myc could induce rapid morphological transformation of nonestablished adrenocortical cells but that an additional genetic or epigenetic event was required to permit full transformation by these two oncogenes. In contrast, v-src, introduced by retrovirus infection in conjunction with v-myc, rapidly induced serum- and anchorage-independent growth. Therefore, the p60v-src protein-tyrosine kinase, unlike p21v-ras, is apparently not restricted in the induction of a highly transformed phenotype in adrenocortical cells. This system provides an in vitro model for the progressive transformation of epithelial cells by dominantly acting oncogenes. Images PMID:2846881

Parasitism and the retrotransposon life cycle in plants: a hitchhiker's guide to the genome.

PubMed

Sabot, F; Schulman, A H

2006-12-01

LTR (long terminal repeat) retrotransposons are the main components of higher plant genomic DNA. They have shaped their host genomes through insertional mutagenesis and by effects on genome size, gene expression and recombination. These Class I transposable elements are closely related to retroviruses such as the HIV by their structure and presumptive life cycle. However, the retrotransposon life cycle has been closely investigated in few systems. For retroviruses and retrotransposons, individual defective copies can parasitize the activity of functional ones. However, some LTR retrotransposon groups as a whole, such as large retrotransposon derivatives and terminal repeats in miniature, are non-autonomous even though their genomic insertion patterns remain polymorphic between organismal accessions. Here, we examine what is known of the retrotransposon life cycle in plants, and in that context discuss the role of parasitism and complementation between and within retrotransposon groups.

Immunotherapies for Targeting Ancient Retrovirus during Breast Cancer

DTIC Science & Technology

2014-03-01

collection of information if it does not display a currently valid OMB control number. PLEASE DO NOT RETURN YOUR FORM TO THE ABOVE ADDRESS. 1. REPORT DATE...in T cells. HERV-K-specific CAR+T cells were selectively propagated ex vivo on artificial antigen presenting cells (aAPC) using an approach already...demonstrate that it is feasible to generate T cells expressing a HERV-K-specific CAR using a clinically-appealing approach as a treatment strategy for HERV-K

A herpes simplex virus 2 glycoprotein D mutant generated by bacterial artificial chromosome mutagenesis is severely impaired for infecting neuronal cells and infects only Vero cells expressing exogenous HVEM.

PubMed

Wang, Kening; Kappel, Justin D; Canders, Caleb; Davila, Wilmer F; Sayre, Dean; Chavez, Mayra; Pesnicak, Lesley; Cohen, Jeffrey I

2012-12-01

We constructed a herpes simplex virus 2 (HSV-2) bacterial artificial chromosome (BAC) clone, bHSV2-BAC38, which contains full-length HSV-2 inserted into a BAC vector. Unlike previously reported HSV-2 BAC clones, the virus genome inserted into this BAC clone has no known gene disruptions. Virus derived from the BAC clone had a wild-type phenotype for growth in vitro and for acute infection, latency, and reactivation in mice. HVEM, expressed on epithelial cells and lymphocytes, and nectin-1, expressed on neurons and epithelial cells, are the two principal receptors used by HSV to enter cells. We used the HSV-2 BAC clone to construct an HSV-2 glycoprotein D mutant (HSV2-gD27) with point mutations in amino acids 215, 222, and 223, which are critical for the interaction of gD with nectin-1. HSV2-gD27 infected cells expressing HVEM, including a human epithelial cell line. However, the virus lost the ability to infect cells expressing only nectin-1, including neuronal cell lines, and did not infect ganglia in mice. Surprisingly, we found that HSV2-gD27 could not infect Vero cells unless we transduced the cells with a retrovirus expressing HVEM. High-level expression of HVEM in Vero cells also resulted in increased syncytia and enhanced cell-to-cell spread in cells infected with wild-type HSV-2. The inability of the HSV2-gD27 mutant to infect neuronal cells in vitro or sensory ganglia in mice after intramuscular inoculation suggests that this HSV-2 mutant might be an attractive candidate for a live attenuated HSV-2 vaccine.

A Herpes Simplex Virus 2 Glycoprotein D Mutant Generated by Bacterial Artificial Chromosome Mutagenesis Is Severely Impaired for Infecting Neuronal Cells and Infects Only Vero Cells Expressing Exogenous HVEM

PubMed Central

Kappel, Justin D.; Canders, Caleb; Davila, Wilmer F.; Sayre, Dean; Chavez, Mayra; Pesnicak, Lesley; Cohen, Jeffrey I.

2012-01-01

We constructed a herpes simplex virus 2 (HSV-2) bacterial artificial chromosome (BAC) clone, bHSV2-BAC38, which contains full-length HSV-2 inserted into a BAC vector. Unlike previously reported HSV-2 BAC clones, the virus genome inserted into this BAC clone has no known gene disruptions. Virus derived from the BAC clone had a wild-type phenotype for growth in vitro and for acute infection, latency, and reactivation in mice. HVEM, expressed on epithelial cells and lymphocytes, and nectin-1, expressed on neurons and epithelial cells, are the two principal receptors used by HSV to enter cells. We used the HSV-2 BAC clone to construct an HSV-2 glycoprotein D mutant (HSV2-gD27) with point mutations in amino acids 215, 222, and 223, which are critical for the interaction of gD with nectin-1. HSV2-gD27 infected cells expressing HVEM, including a human epithelial cell line. However, the virus lost the ability to infect cells expressing only nectin-1, including neuronal cell lines, and did not infect ganglia in mice. Surprisingly, we found that HSV2-gD27 could not infect Vero cells unless we transduced the cells with a retrovirus expressing HVEM. High-level expression of HVEM in Vero cells also resulted in increased syncytia and enhanced cell-to-cell spread in cells infected with wild-type HSV-2. The inability of the HSV2-gD27 mutant to infect neuronal cells in vitro or sensory ganglia in mice after intramuscular inoculation suggests that this HSV-2 mutant might be an attractive candidate for a live attenuated HSV-2 vaccine. PMID:22993162

Possible Application of Biotechnology to the Development of Biological Agents by Potential Enemies

DTIC Science & Technology

1987-06-01

of enzyme catalyzed reactions. Although cloning techniques are directly applicable to the manipulation of proteinaceous toxins, they would be less...useful for nonproteinaceous toxins because the corresponding gene for each enzyme must be cloned and expressed in a coordinated manner. Effective...to produce a synthetic DNA. The enzyme reverse transcriptase (RNA dependent DNA polymerase), which is obtained from retroviruses, is the only enzyme

A1AT polymorphisms may be associated with clinical characteristics of retrovirus infections in a mixed ethnic population from the Brazilian Amazon region.

PubMed

Ferreira, Thalita Camêlo da Silva; Queiroz, Maria Alice Freitas; Argañaraz, Gustavo Adolfo; Ishak, Ricardo; Vallinoto, Antonio Carlos Rosário; Argañaraz, Enrique Roberto

2017-12-01

This study investigated the association of alpha-1-antrypsin deficiency (A1AT; S and Z polymorphisms) with HIV-1 and HTLV-1 infection. Blood samples from 201 HIV-1-infected and 115 HTLV-1-infected individuals were examined and compared with those from 300 healthy controls. Genotyping of A1AT (S and Z) and quantification of plasma viral load were performed using RT-PCR, and the CD4+/CD8+ T-cell count was determined by flow cytometry. The wild-type MM genotype showed the highest frequency in each of the three groups investigated. SS and ZZ homozygous genotypes (variants) were observed only among HTLV-1 patients and controls, respectively. Genotype MS was significantly less frequent in HTLV-1-positive persons than in controls. Statistically significant differences were observed when comparing genotype frequencies between symptomatic and asymptomatic HTLV-1-infected persons. The distribution of plasma HIV-1 viral load among individuals with different genotypes of A1AT polymorphism also differed significantly. The results suggest that A1AT polymorphisms may be associated with human retrovirus infections when dealing with an ethnically mixed population from the Amazon region of Brazil. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Mutations altering the gammaretrovirus endoproteolytic motif affect glycosylation of the envelope glycoprotein and early events of the virus life cycle

DOE Office of Scientific and Technical Information (OSTI.GOV)

Argaw, Takele; Wilson, Carolyn A., E-mail: carolyn.wilson@fda.hhs.gov

Previously, we found that mutation of glutamine to proline in the endoproteolytic cleavage signal of the PERV-C envelope (RQKK to RPKK) resulted in non-infectious vectors. Here, we show that RPKK results in a non-infectious vector when placed in not only a PERV envelope, but also the envelope of a related gammaretrovirus, FeLV-B. The amino acid substitutions do not prevent envelope precursor cleavage, viral core and genome assembly, or receptor binding. Rather, the mutations result in the formation of hyperglycosylated glycoprotein and a reduction in the reverse transcribed minus strand synthesis and undetectable 2-LTR circular DNA in cells exposed to vectorsmore » with these mutated envelopes. Our findings suggest novel functions associated with the cleavage signal sequence that may affect trafficking through the glycosylation machinery of the cell. Further, the glycosylation status of the envelope appears to impact post-binding events of the viral life cycle, either membrane fusion, internalization, or reverse transcription. - Highlights: • Env cleavage signal impacts infectivity of gammaretroviruses. • Non-infectious mutants have hyper-glycosylated envelope that bind target cells. • Non-infectious mutants have defects in the formation of the double-stranded DNA. • Env cleavage motif has functions beyond cleavage of the env precursor.« less

A novel role for APOBEC3: Susceptibility to sexual transmission of murine acquired immunodeficiency virus (mAIDS) is aggravated in APOBEC3 deficient mice

PubMed Central

2012-01-01

Background APOBEC3 proteins are host factors that restrict infection by retroviruses like HIV, MMTV, and MLV and are variably expressed in hematopoietic and non-hematopoietic cells, such as macrophages, lymphocytes, dendritic, and epithelia cells. Previously, we showed that APOBEC3 expressed in mammary epithelia cells function to limit milk-borne transmission of the beta-retrovirus, mouse mammary tumor virus. In this present study, we used APOBEC3 knockout mice and their wild type counterpart to query the role of APOBEC3 in sexual transmission of LP-BM5 MLV – the etiological agent of murine AIDs (mAIDs). Results We show that mouse APOBEC3 is expressed in murine genital tract tissues and gametes and that genital tract tissue of APOBEC3-deficient mice are more susceptible to infection by LP-BM5 virus. APOBEC3 expressed in genital tract tissues most likely plays a role in decreasing virus transmission via the sexual route, since mice deficient in APOBEC3 gene have higher genitalia and seminal plasma virus load and sexually transmit the virus more efficiently to their partners compared to APOBEC3+ mice. Moreover, we show that female mice sexually infected with LP-BM5 virus transmit the virus to their off-spring in APOBEC3-dependent manner. Conclusion Our data indicate that genital tissue intrinsic APOBEC3 restricts genital tract infection and limits sexual transmission of LP-BM5 virus. PMID:22691411

The baculovirus-integrated retrotransposon TED encodes gag and pol proteins that assemble into viruslike particles with reverse transcriptase.

PubMed Central

Lerch, R A; Friesen, P D

1992-01-01

TED is a lepidopteran retrotransposon found inserted within the DNA genome of the Autographa californica nuclear polyhedrosis virus mutant, FP-D. To examine the proteins and functions encoded by this representative of the gypsy family of retrotransposons, the gag- and pol-like open reading frames (ORFs 1 and 2) were expressed in homologous lepidopteran cells by using recombinant baculovirus vectors. Expression of ORF 1 resulted in synthesis of an abundant TED-specific protein (Pr55gag) that assembled into viruslike particles with a diameter of 55 to 60 nm. Expression of ORF 2, requiring a -1 translational frameshift, resulted in synthesis of a protease that mediated cleavage of Pr55gag to generate p37, the major protein component of the resulting particles. Expression of ORF 2 also produced reverse transcriptase that associated with these particles. Both protease and reverse transcriptase activities mapped to domains within ORF 2 that contain sequence similarities with the corresponding functional domains of the pol gene of the vertebrate retroviruses. These results indicated that TED ORFs 1 and 2 functionally resemble the retrovirus gag and pol genes and demonstrated for the first time that an invertebrate member of the gypsy family of elements encodes active forms of the structural and enzymatic functions necessary for transposition via an RNA intermediate. TED integration within the baculovirus genome thus represents one of the first examples of transposon-mediated transfer of host-derived genes to an eukaryotic virus. Images PMID:1371168

Connexins and Cadherin Crosstalk in the Pathogenesis of Prostate Cancer

DTIC Science & Technology

2015-09-01

the plaque as double membrane vesicles, by endocytosis and targeted to the lysosome for degradation. Alternatively, undocked connexons may be...endocytosed by clathrin mediated or non-clathrin mediated endocytosis (Figure 2) [13-16]. Tasks of Aim 1: 1. Prepare recombinant retroviruses that...results were described in 2014 report. 7) Determine if N-cadherin induces endocytosis of gap junctions in connexin-expressing LNCaP (ATCC) and

Purification and protein composition of endogenous rat viruses.

PubMed

Hlubinová, K; Prachar, J; Vrbenská, A; Matoska, J; Simkovic, D

1984-01-01

Endogenous retroviruses are not in the majority of cases the cause of any neoplasia, except for the laboratory conditions. As far as they might serve for the evolution of pathogenic retroviruses more attention should have been paid to them. In this paper we introduce some approaches to the purification of rat endogenous retroviruses to such a degree of purity that enabled satisfactory SDS-PAGE analysis of its structural proteins. Purities of samples obtained by usual purification methods, long-term isopycnic centrifugation at a high gravity force and velocity centrifugation are compared. Protein profile of rat endogenous virus in SDS-PAGE is compared with the ones of other retroviruses. For the first time the evidence was obtained for the striking similarity between electrophoretic protein profile of rat endogenous virus WERC and feline leukemia virus. The major structural proteins of rat endogenous retrovirus and feline leukemia virus cannot be distinguished even when resolution long gradient PAGE had been employed. The accordance of electrophoretic mobilities of major structural proteins in SDS-PAGE can indicate the relatedness of retroviruses.

Humoral immunity response to human endogenous retroviruses K/W differentiates between amyotrophic lateral sclerosis and other neurological diseases.

PubMed

Arru, G; Mameli, G; Deiana, G A; Rassu, A L; Piredda, R; Sechi, E; Caggiu, E; Bo, M; Nako, E; Urso, D; Mariotto, S; Ferrari, S; Zanusso, G; Monaco, S; Sechi, G; Sechi, L A

2018-03-31

Human endogenous retroviruses (HERV) K/W seem to play a role in fostering and exacerbation of some neurological diseases, including amyotrophic lateral sclerosis (ALS). Given these findings, the immunity response against HERV-K and HERV-W envelope surface (env-su) glycoprotein antigens in serum and cerebrospinal fluid (CSF) was investigated for ALS, multiple sclerosis (MS) and Alzheimer's disease patients and in healthy controls. Four antigenic peptides derived respectively from HERV-K and HERV-W env-su proteins were studied in 21 definite or probable ALS patients, 26 possible or definite relapsing-remitting MS patients, 18 patients with Alzheimer's disease and 39 healthy controls. An indirect enzyme-linked immunosorbent assay was set up to detect specific antibodies (Abs) against env-su peptides. Amongst the measured levels of Abs against the four different HERV-K peptide fragments, only HERV-K env-su 19-37 was significantly elevated in ALS compared to other groups, both in serum and CSF. Instead, amongst the Abs levels directed against the four different HERV-W peptide fragments, only HERV-W env-su 93-108 and HERV-W env-su 248-262 were significantly elevated, in the serum and CSF of the MS group compared to other groups. In ALS patients, the HERV-K env-su 19-37 Abs levels were significantly correlated with clinical measures of disease severity, both in serum and CSF. Increased circulating levels of Abs directed against the HERV-W env-su 93-108 and HERV-W env-su 248-262 peptide fragments could serve as possible biomarkers in patients with MS. Similarly, increased circulating levels of Abs directed against the HERV-K env-su 19-37 peptide fragment could serve as a possible early novel biomarker in patients with ALS. © 2018 EAN.

Detection of osteoclastic cell-cell fusion through retroviral vector packaging.

PubMed

Kondo, Takako; Ikeda, Kyoji; Matsuo, Koichi

2004-11-01

Cell-cell fusion generates multinucleated cells such as osteoclasts in bone, myotubes in muscle, and trophoblasts in placenta. Molecular details governing these fusion processes are still largely unknown. As a step toward identification of fusogenic genes, we tested the concept that retroviral vectors can be packaged as a result of cell-cell fusion. First, we introduced replication-deficient retroviral vectors expressing mCAT-1, which mediates fusogenic interaction with the retroviral envelope protein Env, into Chinese hamster ovary (CHO) cells to generate vector cells. Plasmids expressing virion proteins Gag, Pol, and Env were introduced into a separate culture of CHO cells to generate packaging cells. Co-culturing vector and packaging cells resulted in production of infectious retroviruses carrying the mCAT-1 gene as a consequence of cell-cell fusion. Second, we introduced a retroviral vector into primary osteoclast precursors and co-cultured them with established osteoclast precursor RAW264.7 cells, which turned out to harbor packaging activity. Packaged retroviral vector was detected in culture supernatants only where the osteoclast differentiation factor receptor activator for NF-kappaB ligand (RANKL) induced fusion between these two cell types. These data suggest that retrovirus production can occur as a result of cell-cell fusion. This provides a novel approach for isolating and characterizing fusogenic genes using retroviral expression vectors.

Lung adenocarcinoma originates from retrovirus infection of proliferating type 2 pneumocytes during pulmonary post-natal development or tissue repair.

PubMed

Murgia, Claudio; Caporale, Marco; Ceesay, Ousman; Di Francesco, Gabriella; Ferri, Nicola; Varasano, Vincenzo; de las Heras, Marcelo; Palmarini, Massimo

2011-03-01

Jaagsiekte sheep retrovirus (JSRV) is a unique oncogenic virus with distinctive biological properties. JSRV is the only virus causing a naturally occurring lung cancer (ovine pulmonary adenocarcinoma, OPA) and possessing a major structural protein that functions as a dominant oncoprotein. Lung cancer is the major cause of death among cancer patients. OPA can be an extremely useful animal model in order to identify the cells originating lung adenocarcinoma and to study the early events of pulmonary carcinogenesis. In this study, we demonstrated that lung adenocarcinoma in sheep originates from infection and transformation of proliferating type 2 pneumocytes (termed here lung alveolar proliferating cells, LAPCs). We excluded that OPA originates from a bronchioalveolar stem cell, or from mature post-mitotic type 2 pneumocytes or from either proliferating or non-proliferating Clara cells. We show that young animals possess abundant LAPCs and are highly susceptible to JSRV infection and transformation. On the contrary, healthy adult sheep, which are normally resistant to experimental OPA induction, exhibit a relatively low number of LAPCs and are resistant to JSRV infection of the respiratory epithelium. Importantly, induction of lung injury increased dramatically the number of LAPCs in adult sheep and rendered these animals fully susceptible to JSRV infection and transformation. Furthermore, we show that JSRV preferentially infects actively dividing cell in vitro. Overall, our study provides unique insights into pulmonary biology and carcinogenesis and suggests that JSRV and its host have reached an evolutionary equilibrium in which productive infection (and transformation) can occur only in cells that are scarce for most of the lifespan of the sheep. Our data also indicate that, at least in this model, inflammation can predispose to retroviral infection and cancer.

Functions, structure, and read-through alternative splicing of feline APOBEC3 genes

PubMed Central

Münk, Carsten; Beck, Thomas; Zielonka, Jörg; Hotz-Wagenblatt, Agnes; Chareza, Sarah; Battenberg, Marion; Thielebein, Jens; Cichutek, Klaus; Bravo, Ignacio G; O'Brien, Stephen J; Lochelt, Martin; Yuhki, Naoya

2008-01-01

Background Over the past years a variety of host restriction genes have been identified in human and mammals that modulate retrovirus infectivity, replication, assembly, and/or cross-species transmission. Among these host-encoded restriction factors, the APOBEC3 (A3; apolipoprotein B mRNA-editing catalytic polypeptide 3) proteins are potent inhibitors of retroviruses and retrotransposons. While primates encode seven of these genes (A3A to A3H), rodents carry only a single A3 gene. Results Here we identified and characterized several A3 genes in the genome of domestic cat (Felis catus) by analyzing the genomic A3 locus. The cat genome presents one A3H gene and three very similar A3C genes (a-c), probably generated after two consecutive gene duplications. In addition to these four one-domain A3 proteins, a fifth A3, designated A3CH, is expressed by read-through alternative splicing. Specific feline A3 proteins selectively inactivated only defined genera of feline retroviruses: Bet-deficient feline foamy virus was mainly inactivated by feA3Ca, feA3Cb, and feA3Cc, while feA3H and feA3CH were only weakly active. The infectivity of Vif-deficient feline immunodeficiency virus and feline leukemia virus was reduced only by feA3H and feA3CH, but not by any of the feA3Cs. Within Felidae, A3C sequences show significant adaptive selection, but unexpectedly, the A3H sequences present more sites that are under purifying selection. Conclusion Our data support a complex evolutionary history of expansion, divergence, selection and individual extinction of antiviral A3 genes that parallels the early evolution of Placentalia, becoming more intricate in taxa in which the arms race between host and retroviruses is harsher. PMID:18315870

Proviruses with Long-Term Stable Expression Accumulate in Transcriptionally Active Chromatin Close to the Gene Regulatory Elements: Comparison of ASLV-, HIV- and MLV-Derived Vectors

PubMed Central

Miklík, Dalibor; Šenigl, Filip; Hejnar, Jiří

2018-01-01

Individual groups of retroviruses and retroviral vectors differ in their integration site preference and interaction with the host genome. Hence, immediately after infection genome-wide distribution of integrated proviruses is non-random. During long-term in vitro or persistent in vivo infection, the genomic position and chromatin environment of the provirus affects its transcriptional activity. Thus, a selection of long-term stably expressed proviruses and elimination of proviruses, which have been gradually silenced by epigenetic mechanisms, helps in the identification of genomic compartments permissive for proviral transcription. We compare here the extent and time course of provirus silencing in single cell clones of the K562 human myeloid lymphoblastoma cell line that have been infected with retroviral reporter vectors derived from avian sarcoma/leukosis virus (ASLV), human immunodeficiency virus type 1 (HIV) and murine leukaemia virus (MLV). While MLV proviruses remain transcriptionally active, ASLV proviruses are prone to rapid silencing. The HIV provirus displays gradual silencing only after an extended time period in culture. The analysis of integration sites of long-term stably expressed proviruses shows a strong bias for some genomic features—especially integration close to the transcription start sites of active transcription units. Furthermore, complex analysis of histone modifications enriched at the site of integration points to the accumulation of proviruses of all three groups in gene regulatory segments, particularly close to the enhancer loci. We conclude that the proximity to active regulatory chromatin segments correlates with stable provirus expression in various retroviral species. PMID:29517993

Infection of chicken bone marrow mononuclear cells with subgroup J avian leukosis virus inhibits dendritic cell differentiation and alters cytokine expression.

PubMed

Liu, Di; Qiu, Qianqian; Zhang, Xu; Dai, Manman; Qin, Jianru; Hao, Jianjong; Liao, Ming; Cao, Weisheng

2016-10-01

Subgroup J avian leukosis virus (ALV-J) is an oncogenic retrovirus known to induce tumor formation and immunosuppression in infected chickens. One of the organs susceptible to ALV-J is the bone marrow, from which specialized antigen-presenting cells named dendritic cells (BM-DCs) are derived. Notably, these cells possess the unique ability to induce primary immune responses. In the present study, a method of cultivating and purifying DCs from chicken bone marrow in vitro was established to investigate the effects of ALV-J infection on BM-DC differentiation or generation. The results indicated that ALV-J not only infects the chicken bone marrow mononuclear cells but also appears to inhibit the differentiation and maturation of BM-DCs and to trigger apoptosis. Moreover, substantial reductions in the mRNA expression of TLR1, TLR2, TLR3, MHCI, and MHCII and in cytokine production were detected in the surviving BM-DCs following ALV-J infection. These findings indicate that ALV-J infection disrupts the process of bone marrow mononuclear cell differentiation into BM-DCs likely via altered antigen presentation, resulting in a downstream immune response in affected chickens. Copyright © 2016 Elsevier B.V. All rights reserved.

Localization and Sub-Cellular Shuttling of HTLV-1 Tax with the miRNA Machinery

PubMed Central

Van Duyne, Rachel; Guendel, Irene; Klase, Zachary; Narayanan, Aarthi; Coley, William; Jaworski, Elizabeth; Roman, Jessica; Popratiloff, Anastas; Mahieux, Renaud; Kehn-Hall, Kylene; Kashanchi, Fatah

2012-01-01

The innate ability of the human cell to silence endogenous retroviruses through RNA sequences encoding microRNAs, suggests that the cellular RNAi machinery is a major means by which the host mounts a defense response against present day retroviruses. Indeed, cellular miRNAs target and hybridize to specific sequences of both HTLV-1 and HIV-1 viral transcripts. However, much like the variety of host immune responses to retroviral infection, the virus itself contains mechanisms that assist in the evasion of viral inhibition through control of the cellular RNAi pathway. Retroviruses can hijack both the enzymatic and catalytic components of the RNAi pathway, in some cases to produce novel viral miRNAs that can either assist in active viral infection or promote a latent state. Here, we show that HTLV-1 Tax contributes to the dysregulation of the RNAi pathway by altering the expression of key components of this pathway. A survey of uninfected and HTLV-1 infected cells revealed that Drosha protein is present at lower levels in all HTLV-1 infected cell lines and in infected primary cells, while other components such as DGCR8 were not dramatically altered. We show colocalization of Tax and Drosha in the nucleus in vitro as well as coimmunoprecipitation in the presence of proteasome inhibitors, indicating that Tax interacts with Drosha and may target it to specific areas of the cell, namely, the proteasome. In the presence of Tax we observed a prevention of primary miRNA cleavage by Drosha. Finally, the changes in cellular miRNA expression in HTLV-1 infected cells can be mimicked by the add back of Drosha or the addition of antagomiRs against the cellular miRNAs which are downregulated by the virus. PMID:22808228

Induction of complex immune responses and strong protection against retrovirus challenge by adenovirus-based immunization depends on the order of vaccine delivery.

PubMed

Kaulfuß, Meike; Wensing, Ina; Windmann, Sonja; Hrycak, Camilla Patrizia; Bayer, Wibke

2017-02-06

In the Friend retrovirus mouse model we developed potent adenovirus-based vaccines that were designed to induce either strong Friend virus GagL 85-93 -specific CD8 + T cell or antibody responses, respectively. To optimize the immunization outcome we evaluated vaccination strategies using combinations of these vaccines. While the vaccines on their own confer strong protection from a subsequent Friend virus challenge, the simple combination of the vaccines for the establishment of an optimized immunization protocol did not result in a further improvement of vaccine effectivity. We demonstrate that the co-immunization with GagL 85-93 /leader-gag encoding vectors together with envelope-encoding vectors abrogates the induction of GagL 85-93 -specific CD8 + T cells, and in successive immunization protocols the immunization with the GagL 85-93 /leader-gag encoding vector had to precede the immunization with an envelope encoding vector for the efficient induction of GagL 85-93 -specific CD8 + T cells. Importantly, the antibody response to envelope was in fact enhanced when the mice were adenovirus-experienced from a prior immunization, highlighting the expedience of this approach. To circumvent the immunosuppressive effect of envelope on immune responses to simultaneously or subsequently administered immunogens, we developed a two immunizations-based vaccination protocol that induces strong immune responses and confers robust protection of highly Friend virus-susceptible mice from a lethal Friend virus challenge.

Testing New Drugs for Treatment of Melanoma Patients Applying Connectivity Map Database Analysis with Melanoma Gene Signatures

DTIC Science & Technology

2012-10-01

use of R packages implemented in Bioconductor. Each dataset was normalized from raw data using the Frozen RMA (fRMA) algorithm . We applied the same...because development of the specific algorithms and fine tuning of the analytic strategy to accomplish this task was not immediately straightforward. We...express firefly luciferase using a retrovirus that encodes a fusion of luciferase and neomycin phosphotransferase (LucNeo), will be implanted and followed

In Vivo Selection of CD4+ T Cells Transduced with a Gamma-Retroviral Vector Expressing a Single-Chain Intrabody Targeting HIV-1 Tat

PubMed Central

Braun, Stephen E.; Taube, Ran; Zhu, Quan; Wong, Fay Eng; Murakami, Akikazu; Kamau, Erick; Dwyer, Markryan; Qiu, Gang; Daigle, Janet; Carville, Angela; Johnson, R. Paul

2012-01-01

Abstract We evaluated the potential of an anti–human immunodeficiency virus (HIV) Tat intrabody (intracellular antibody) to promote the survival of CD4+ cells after chimeric simian immunodeficiency virus (SIV)/HIV (SHIV) infection in rhesus macaques. Following optimization of stimulation and transduction conditions, purified CD4+ T cells were transduced with GaLV-pseudotyped retroviral vectors expressing either an anti-HIV-1 Tat or a control single-chain intrabody. Ex vivo intrabody-gene marking was highly efficient, averaging four copies per CD4+ cell. Upon reinfusion of engineered autologous CD4+ cells into two macaques, high levels of gene marking (peak of 0.6% and 6.8% of peripheral blood mononuclear cells (PBMCs) and 0.3% or 2.2% of the lymph node cells) were detected in vivo. One week post cell infusion, animals were challenged with SHIV 89.6p and the ability of the anti-HIV Tat intrabody to promote cell survival was evaluated. The frequency of genetically modified CD4+ T cells progressively decreased, concurrent with loss of CD4+ cells and elevated viral loads in both animals. However, CD4+ T cells expressing the therapeutic anti-Tat intrabody exhibited a relative survival advantage over an 8- and 21-week period compared with CD4+ cells expressing a control intrabody. In one animal, this survival benefit of anti-Tat transduced cells was associated with a reduction in viral load. Overall, these results indicate that a retrovirus-mediated anti-Tat intrabody provided significant levels of gene marking in PBMCs and peripheral tissues and increased relative survival of transduced cells in vivo. PMID:22734618

The Effects of Curcuma longa L., Purple Sweet Potato, and Mixtures of the Two on Immunomodulation in C57BL/6J Mice Infected with LP-BM5 Murine Leukemia Retrovirus.

PubMed

Park, Soo-Jeung; Lee, Dasom; Lee, Minhee; Kwon, Han-Ol; Kim, Hyesook; Park, Jeongjin; Jeon, Woojin; Cha, Minseok; Jun, Suhwa; Park, Kwangjin; Lee, Jeongmin

2018-06-04

The immune response is stimulated to protect the body from external antigens and is controlled by several types of immune cells. In the present study, the immunomodulatory effects of Curcuma longa L., purple sweet potato, and mixtures of the two (CPM) were investigated in C57BL/6 mice infected with LP-BM5 murine leukemia virus (MuLV). Mice were divided into seven groups as follows: normal control, infected control (LP-BM5 MuLV infection), positive control (LP-BM5 MuLV infection+dietary supplement of red ginseng 300 mg/kg body weight), the original powder of C. longa L. (C; LP-BM5 MuLV infection+dietary supplement of C 189 mg/kg body weight), the original powder of purple sweet potato (P; LP-BM5 MuLV infection+dietary supplement of P 1811 mg/kg body weight), CPM Low (CPL; LP-BM5 MuLV infection+CPM 2 g/kg body weight), and CPM High (CPH; LP-BM5 MuLV infection+CPM 5 g/kg body weight). Dietary supplementation lasted for 12 weeks. Dietary supplementation of CPM inhibited LP-BM5 MuLV-induced lymphadenopathy and splenomegaly and inhibited reduction of messenger RNA (mRNA) expression of major histocompatibility complex (MHC) I and II. Moreover, CPM reduced the decrease in T- and B cell proliferation, reduced the population of CD4(+)/CD8(+) T cells, and remedied the unbalanced production of T helper-1 (Th1)/T helper-2 (Th2) cytokines in LP-BM5 MuLV-infected mice. In addition, CPM inhibited reduction of phagocytosis in peritoneal macrophages and decreased serum levels of immunoglobulin A (IgA), immunoglobulin E (IgE), and immunoglobulin G (IgG). These results suggest that CPM had a positive effect on immunomodulation in C57BL/6 mice induced by LP-BM5 leukemia retrovirus infection.

Resistance to experimental tumorigenesis in cells of a long-lived mammal, the naked mole-rat (Heterocephalus glaber)

PubMed Central

Liang, Sitai; Mele, James; Wu, Yuehong; Buffenstein, Rochelle; Hornsby, Peter J.

2013-01-01

Summary The naked mole-rat (NMR, Heterocephalus glaber) is a long-lived mammal in which spontaneous cancer has not been observed. In order to investigate possible mechanisms for cancer resistance in this species, we studied the properties of skin fibroblasts from the NMR following transduction with oncogenes that cause cells of other mammalian species to form malignant tumors. NMR fibroblasts were transduced with a retrovirus encoding SV40 large T antigen and oncogenic RasG12V. Following transplantation of transduced cells into immunodeficient mice, cells rapidly entered crisis, as evidenced by the presence of anaphase bridges, giant cells with enlarged nuclei, multinucleated cells, and cells with large number of chromosomes or abnormal chromatin material. In contrast, similarly transduced mouse and rat fibroblasts formed tumors that grew rapidly without crisis. Crisis was also observed after >40 population doublings in SV40 TAg/Ras-expressing NMR cells in culture. Crisis in culture was prevented by additional infection of the cells with a retrovirus encoding hTERT (telomerase reverse transcriptase). SV40 TAg/Ras/hTERT-expressing NMR cells formed tumors that grew rapidly in immunodeficient mice without evidence of crisis. Crisis could also be induced in SV40 TAg/Ras-expressing NMR cells by loss of anchorage, but after hTERT transduction cells were able to proliferate normally following loss of anchorage. Thus, rapid crisis is a response of oncogene-expressing NMR cells to growth in an in vivo environment, which requires anchorage independence, and hTERT permits cells to avoid crisis and to achieve malignant tumor growth. The unique reaction of NMR cells to oncogene expression may form part of the cancer resistance of this species. PMID:20550519

Targeted disruption of Pten in ovarian granulosa cells enhances ovulation and extends the life span of luteal cells.

PubMed

Fan, Heng-Yu; Liu, Zhilin; Cahill, Nicola; Richards, JoAnne S

2008-09-01

FSH activates the phosphatidylinositol-3 kinase (PI3K)/acute transforming retrovirus thymoma protein kinase pathway and thereby enhances granulosa cell differentiation in culture. To identify the physiological role of the PI3K pathway in vivo we disrupted the PI3K suppressor, Pten, in developing ovarian follicles. To selectively disrupt Pten expression in granulosa cells, Ptenfl/fl mice were mated with transgenic mice expressing cAMP response element recombinase driven by Cyp19 promoter (Cyp19-Cre). The resultant Pten mutant mice were fertile, ovulated more oocytes, and produced moderately more pups than control mice. These physiological differences in the Pten mutant mice were associated with hyperactivation of the PI3K/acute transforming retrovirus thymoma protein kinase pathway, decreased susceptibility to apoptosis, and increased proliferation of mutant granulosa cells. Strikingly, corpora lutea of the Pten mutant mice persisted longer than those of control mice. Although the follicular and luteal cell steroidogenesis in Ptenfl/fl;Cyp19-Cre mice was similar to controls, viable nonsteroidogenic luteal cells escaped structural luteolysis. These findings provide the novel evidence that Pten impacts the survival/life span of granulosa/luteal cells and that its loss not only results in the facilitated ovulation but also in the persistence of nonsteroidogenic luteal structures in the adult mouse ovary.

Silver nanoparticles exhibit size-dependent differential toxicity and induce expression of syncytin-1 in FA-AML1 and MOLT-4 leukaemia cell lines.

PubMed

Alqahtani, Sultan; Promtong, Pawika; Oliver, Anthony W; He, Xiaotong T; Walker, Thomas D; Povey, Andrew; Hampson, Lynne; Hampson, Ian N

2016-11-01

Human endogenous retrovirus (HERV) sequences make up ~8% of the human genome and increased expression of some HERV proteins has been observed in various pathologies including leukaemia and multiple sclerosis. However, little is known about the function of these HERV proteins or environmental factors which regulate their expression. Silver nanoparticles (AgNPs) are used very extensively as antimicrobials and antivirals in numerous consumer products although their effect on the expression of HERV gene products is unknown. Cell proliferation and cell toxicity assays were carried out on human acute T lymphoblastic leukaemia (MOLT-4) and Fanconi anaemia associated acute myeloid leukaemia (FA-AML1) cells treated with two different sizes of AgNPs (7nm and 50nm diameter). Reverse-transcriptase polymerase chain reaction and western blotting were then used to the assess expression of HERV-W syncytin-1 mRNA and protein in these cells. FA-AML1 cells were more sensitive overall than MOLT-4 to treatment with the smaller 7nm sized AgNp's being the most toxic in these cells. MOLT-4 cell were more resistant and showed no evidence of differential toxicity to the different sized particles. Syncytin-1 mRNA and protein were induced by both 7 and 50nm AgNPs in both cell types yet with different kinetics. In summary, the observation that AgNPs induce expression of syncytin-1 in FA-AML1 and MOLT-4 cells at doses as little as 5 µg/ml is grounds for concern since this protein is up-regulated in both malignant and neurodegenerative diseases. Considering the widespread use of AgNPs in the environment it is clear that their ability to induce syncytin-1 should be investigated further in other cell types. © The Author 2016. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Investigations of High Resolution Imaging through the Earth’s Atmosphere Using Speckle Interferometry

DTIC Science & Technology

1984-03-07

improved the SO/AFGL speckle camera for use at any conventionai`teescop" by pErv’-ion ofmicro-processor controlled " Risley prisms, shutter mechanisms...condition in the IR because of the all-reflecting system (the wedges used for mode . C are normally not used for IR work). In the 2ptical Experiments the...of pairs of •’ 16 fl**S ’ ,**S * !, S glass wedges (mode C). Beckers and Roddier (1982) used the Roddier rotation sholaring interferometer to obtain

Reduced frequency of murine cytomegalovirus retinitis in C57BL/6 mice correlates with low levels of suppressor of cytokine signaling (SOCS)1 and SOCS3 expression within the eye during corticosteroid-induced immunosuppression.

PubMed

Alston, Christine I; Dix, Richard D

2017-09-01

AIDS-related human cytomegalovirus retinitis remains a leading cause of blindness worldwide. We compared two C57BL/6 mouse models of experimental murine cytomegalovirus (MCMV) retinitis for intraocular expression of suppressors of cytokine signaling (SOCS)1 and SOCS3, host proteins that are inducible negative feedback regulators of cytokine signaling. These mouse models differed in method of immune suppression, one by retrovirus-induced immune suppression (MAIDS) and the other by corticosteroid-induced immune suppression. Following subretinal injection of MCMV to induce retinitis, intraocular SOCS1 and SOCS3 were only mildly stimulated, and often without significance, within MCMV-infected eyes during the progression of MCMV retinitis in corticosteroid-immunosuppressed mice, contrary to MCMV-infected eyes of mice with MAIDS that showed significant high stimulation of SOCS1 and SOCS3 expression in agreement with previous findings. Frequency and severity of retinitis as well as amounts of intraocular infectious MCMV in corticosteroid-immunosuppressed mice were also unexpectedly lower than values previously reported for MAIDS animals during MCMV retinitis. These data reveal a major difference between two mouse models of experimental MCMV retinitis and suggest a possible link between the amplitude of SOCS1 and SOCS3 stimulation and severity of disease in these models. Copyright © 2017 Elsevier Ltd. All rights reserved.

Cross- and Co-Packaging of Retroviral RNAs and Their Consequences

PubMed Central

Ali, Lizna M.; Rizvi, Tahir A.; Mustafa, Farah

2016-01-01

Retroviruses belong to the family Retroviridae and are ribonucleoprotein (RNP) particles that contain a dimeric RNA genome. Retroviral particle assembly is a complex process, and how the virus is able to recognize and specifically capture the genomic RNA (gRNA) among millions of other cellular and spliced retroviral RNAs has been the subject of extensive investigation over the last two decades. The specificity towards RNA packaging requires higher order interactions of the retroviral gRNA with the structural Gag proteins. Moreover, several retroviruses have been shown to have the ability to cross-/co-package gRNA from other retroviruses, despite little sequence homology. This review will compare the determinants of gRNA encapsidation among different retroviruses, followed by an examination of our current understanding of the interaction between diverse viral genomes and heterologous proteins, leading to their cross-/co-packaging. Retroviruses are well-known serious animal and human pathogens, and such a cross-/co-packaging phenomenon could result in the generation of novel viral variants with unknown pathogenic potential. At the same time, however, an enhanced understanding of the molecular mechanisms involved in these specific interactions makes retroviruses an attractive target for anti-viral drugs, vaccines, and vectors for human gene therapy. PMID:27727192

Cross- and Co-Packaging of Retroviral RNAs and Their Consequences.

PubMed

Ali, Lizna M; Rizvi, Tahir A; Mustafa, Farah

2016-10-11

Retroviruses belong to the family Retroviridae and are ribonucleoprotein (RNP) particles that contain a dimeric RNA genome. Retroviral particle assembly is a complex process, and how the virus is able to recognize and specifically capture the genomic RNA (gRNA) among millions of other cellular and spliced retroviral RNAs has been the subject of extensive investigation over the last two decades. The specificity towards RNA packaging requires higher order interactions of the retroviral gRNA with the structural Gag proteins. Moreover, several retroviruses have been shown to have the ability to cross-/co-package gRNA from other retroviruses, despite little sequence homology. This review will compare the determinants of gRNA encapsidation among different retroviruses, followed by an examination of our current understanding of the interaction between diverse viral genomes and heterologous proteins, leading to their cross-/co-packaging. Retroviruses are well-known serious animal and human pathogens, and such a cross-/co-packaging phenomenon could result in the generation of novel viral variants with unknown pathogenic potential. At the same time, however, an enhanced understanding of the molecular mechanisms involved in these specific interactions makes retroviruses an attractive target for anti-viral drugs, vaccines, and vectors for human gene therapy.

Caveolin-1 interacts with the Gag precursor of murine leukaemia virus and modulates virus production

PubMed Central

Yu, Zheng; Beer, Christiane; Koester, Mario; Wirth, Manfred

2006-01-01

Background Retroviral Gag determines virus assembly at the plasma membrane and the formation of virus-like particles in intracellular multivesicular bodies. Thereby, retroviruses exploit by interaction with cellular partners the cellular machineries for vesicular transport in various ways. Results The retroviral Gag precursor protein drives assembly of murine leukaemia viruses (MLV) at the plasma membrane (PM) and the formation of virus like particles in multivesicular bodies (MVBs). In our study we show that caveolin-1 (Cav-1), a multifunctional membrane-associated protein, co-localizes with Gag in a punctate pattern at the PM of infected NIH 3T3 cells. We provide evidence that Cav-1 interacts with the matrix protein (MA) of the Gag precursor. This interaction is mediated by a Cav-1 binding domain (CBD) within the N-terminus of MA. Interestingly, the CBD motif identified within MA is highly conserved among most other γ-retroviruses. Furthermore, Cav-1 is incorporated into MLV released from NIH 3T3 cells. Overexpression of a GFP fusion protein containing the putative CBD of the retroviral MA resulted in a considerable decrease in production of infectious retrovirus. Moreover, expression of a dominant-negative Cav-1 mutant affected retroviral titres significantly. Conclusion This study demonstrates that Cav-1 interacts with MLV Gag, co-localizes with Gag at the PM and affects the production of infectious virus. The results strongly suggest a role for Cav-1 in the process of virus assembly. PMID:16956408

Biallelic expression of the L-arginine:glycine amidinotransferase gene with different methylation status between male and female primordial germ cells in chickens.

PubMed

Jang, H J; Lee, M O; Kim, S; Kim, T H; Kim, S K; Song, G; Womack, J E; Han, J Y

2013-03-01

The basic functions of DNA methylation include in gene silencing by methylation of specific gene promoters, defense of the host genome from retrovirus, and transcriptional suppression of transgenes. In addition, genomic imprinting, by which certain genes are expressed in a parent-of-origin-specific manner, has been observed in a wide range of plants and animals and has been associated with differential methylation. However, imprinting phenomena of DNA methylation effects have not been revealed in chickens. To analyze whether genomic imprinting occurs in chickens, methyl-DNA immunoprecipitation array analysis was applied across the entire genome of germ cells in early chick embryos. A differentially methylated region (DMR) was detected in the eighth intron of the l-arginine:glycine amidinotransferase (GATM) gene. When the DMR in GATM was analyzed by bisulfite sequencing, the methylation in male primordial germ cells (PGC) of 6-d-old embryos was higher than that in female PGC (57.5 vs. 35.0%). At 8 d, the DMR methylation of GATM in male PGC was 3.7-fold higher than that in female PGC (65.0 vs. 17.5%). Subsequently, to investigate mono- or biallelic expression of the GATM gene during embryo development, we found 2 indel sequences (GTTTAATGC and CAAAAA) within the GATM 3'-untranslated region in Korean Oge (KO) and White Leghorn (WL) chickens. When individual WL and KO chickens were genotyped for indel sequences, 3 allele combinations (homozygous insertion, homozygous deletion, and heterozygotes) were detected in both breeds using a gel shift assay and high-resolution melt assay. The deletion allele was predominant in KO, whereas the insertion allele was predominant in WL. Heterozygous animals were evenly distributed in both breeds (P < 0.01). Despite the different methylation status between male and female PGC, the GATM gene conclusively displayed biallelic expression in PGC as well as somatic embryonic, extraembryonic, and adult chicken tissues.

Initial preclinical safety of non-replicating human endogenous retrovirus envelope protein-coated baculovirus vector-based vaccines against human papillomavirus.

PubMed

Han, Su-Eun; Kim, Mi-Gyeong; Lee, Soondong; Cho, Hee-Jeong; Byun, Youngro; Kim, Sujeong; Kim, Young Bong; Choi, Yongseok; Oh, Yu-Kyoung

2013-12-01

Human endogenous retrovirus (HERV) envelope protein-coated, baculovirus vector-based HPV 16 L1 (AcHERV-HPV16L1) is a non-replicating recombinant baculoviral vaccine. Here, we report an initial evaluation of the preclinical safety of AcHERV-HPV16L1 vaccine. In an acute toxicity study, a single administration of AcHERV-HPV16L1 DNA vaccine given intramuscularly (i.m.) to mice at a dose of 1 × 10(8) plaque-forming units (PFU) did not cause significant changes in body weight compared with vehicle-treated controls. It did cause a brief increase in the weights of some organs on day 15 post-treatment, but by day 30, all organ weights were not significantly different from those in the vehicle-treated control group. No hematological changes were observed on day 30 post-treatment. In a range-finding toxicity study with three doses of 1 × 10(7) , 2 × 10(7) and 5 × 10(7) PFU once daily for 5 days, the group treated with 5 × 10(7) PFU showed a transient decrease in the body weights from day 5 to day 15 post-treatment, but recovery to the levels similar to those in the vehicle-treated control group by post-treatment day 20. Organ weights were slightly higher for lymph nodes, spleen, thymus and liver after repeated dosing with 5 × 10(7) PFU on day 15, but had normalized by day 30. Moreover, repeated administration of AcHERV-HPV16L1 did not induce myosin-specific autoantibody in serum, and did not cause immune complex deposition or tissue damage at injection sites. Taken together, these results provide preliminary evidence of the preclinical safety of AcHERV-based HPV16L1 DNA vaccines in mice. Copyright © 2012 John Wiley & Sons, Ltd.

Characterization of replication-competent retroviruses from nonhuman primates with virus-induced T-cell lymphomas and observations regarding the mechanism of oncogenesis.

PubMed Central

Vanin, E F; Kaloss, M; Broscius, C; Nienhuis, A W

1994-01-01

Rapidly progressive T-cell lymphomas were observed in 3 of 10 rhesus monkeys several months after autologous transplantation of enriched bone marrow stem cells that had been transduced with a retroviral vector preparation containing replication-competent virus (R. E. Donahue, S. W. Kessler, D. Bodice, K. McDonagh, C. Dunbar, S. Goodman, B. Agricola, E. Byrne, M. Raffeld, R. Moen, J. Bacher, K. M. Zsebo, and A. W. Nienhuis, J. Exp. Med. 176:1124-1135, 1992). The animals with lymphoma appeared to be tolerant to retroviral antigens in that their sera lacked antibodies reactive with viral proteins and contained 10(4) to 10(5) infectious virus particles per ml. By molecular cloning and DNA sequencing, we have now demonstrated that the serum from one of the monkeys contained a replication-competent retrovirus that arose by recombination between vector and packaging encoding sequences (vector/helper [V/H] recombinant) in the producer clone used for transduction of bone marrow stem cells. Southern blot analysis demonstrated 14 or 25 copies of this genome per cell where present in two animals. The genome of a second replication-competent virus was also recovered by molecular cloning; it arose by recombination involving the genome of the V/H recombinant and endogenous murine retroviral genomes in the producer clone. Twelve copies of this amphotropic virus/mink cell focus-forming virus genome were present in tumor DNA of one animal, but it was not found in tumor DNA of the other two animals with lymphoma. Southern blot analysis of DNA from various tissues demonstrated common insertion site bands in several samples of tumor DNA from one animal, suggesting clonal origin of the lymphoma. Our data are most consistent with a pathogenic mechanism in which chronic productive retroviral infection allowed insertional mutagenesis of critical growth control genes, leading to cell transformation and clonal tumor evolution. Images PMID:8207799

Functional analysis of human foamy virus accessory reading frames.

PubMed Central

Baunach, G; Maurer, B; Hahn, H; Kranz, M; Rethwilm, A

1993-01-01

Foamy viruses belong to the retroviruses which possess a complex genome structure. The human foamy virus (HFV) isolate bears three open reading frames (the so-called bel genes) in the 3' region of the genome which have been reported to give rise to possibly six different proteins via alternative splicing (W. Muranyi and R. M. Flügel, J. Virol. 65:727-735, 1991). In order to analyze the requirements of these proteins for HFV replication in vitro, we constructed a set of single and combinatory bel gene mutants of an infectious molecular clone of HFV. The mutant which lacked the transacting activator, bel-1, was found to be replication incompetent. All other mutants replicated equally well and gave rise to comparable titers of infectious cell-free virus. When HFV proviruses were put under the control of a heterologous promoter (simian virus 40), none of the accessory gene products was found to be required for expression of structural (gag) proteins. There was no evidence for a posttranscriptional regulatory protein that is present in other complex retroviruses. Images PMID:8394455

LINE-1 Retroelements Complexed and Inhibited by Activation Induced Cytidine Deaminase

PubMed Central

Metzner, Mirjam; Jäck, Hans-Martin; Wabl, Matthias

2012-01-01

LINE-1 (abbreviated L1) is a major class of retroelements in humans and mice. If unrestricted, retroelements accumulate in the cytoplasm and insert their DNA into the host genome, with the potential to cause autoimmune disease and cancer. Retroviruses and other retroelements are inhibited by proteins of the APOBEC family, of which activation-induced cytidine deaminase (AID) is a member. Although AID is mainly known for being a DNA mutator shaping the antibody repertoire in B lymphocytes, we found that AID also restricts de novo L1 integrations in B- and non-B-cell lines. It does so by decreasing the protein level of open reading frame 1 (ORF1) of both exogenous and endogenous L1. In activated B lymphocytes, AID deficiency increased L1 mRNA 1.6-fold and murine leukemia virus (MLV) mRNA 2.7-fold. In cell lines and activated B lymphocytes, AID forms cytoplasmic high-molecular-mass complexes with L1 mRNA, which may contribute to L1 restriction. Because AID-deficient activated B lymphocytes do not express ORF1 protein, we suggest that ORF1 protein expression is inhibited by additional restriction factors in these cells. The greater increase in MLV compared to L1 mRNA in AID-deficient activated B lymphocytes may indicate less strict surveillance of retrovirus. PMID:23133680

Analysis of gene transfer rate with immobilized retroviral vectors.

PubMed

Peng, Ching-An

2009-04-01

Efficient delivery of transgenes into the cell nucleus by retroviral vectors in a static culture system is limited by the intrinsic features of incompetent retroviruses (i.e., thermodynamically unstable envelope proteins and low titers). Although several physicochemical approaches (e.g., adding polycationic polymer and applying magnetic force) have been reported to augment the retroviral gene transfer rate, none are suitable for scaling up to a setting for clinical use. The study of using acoustic fields with the form of standing waves has recently been reported to be a feasible way to enhance retroviral gene delivery efficiency in large-scale settings. The concept of using ultrasound standing-wave fields to increase retrovirus-mediated gene transfer is based on quickly established cell bands on acoustic nodal planes as nucleating sites to capture unstable colloidlike retroviruses. In this study, instead of having retroviral nanoparticles circulated between nodal planes, we proposed to immobilize retroviruses onto acoustic transparent films arranged in an acoustic chamber. Then, cells inoculated into the acoustic chamber can be driven by the primary radiation forces to the retrovirus-coated films that are constructed on the nodal planes. To obtain the optimal time of immobilizing retroviruses onto the acoustic transparent film prior to the inception of acoustic fields, we developed a retroviral diffusion-reaction model to describe such a static retroviral system. Analysis of viral transport model has its merit to guide experimental design for attaining high gene transfer efficiency.

Characterization of the fusion core in zebrafish endogenous retroviral envelope protein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shi, Jian; State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei 430071; Zhang, Huaidong

2015-05-08

Zebrafish endogenous retrovirus (ZFERV) is the unique endogenous retrovirus in zebrafish, as yet, containing intact open reading frames of its envelope protein gene in zebrafish genome. Similarly, several envelope proteins of endogenous retroviruses in human and other mammalian animal genomes (such as syncytin-1 and 2 in human, syncytin-A and B in mouse) were identified and shown to be functional in induction of cell–cell fusion involved in placental development. ZFERV envelope protein (Env) gene appears to be also functional in vivo because it is expressible. After sequence alignment, we found ZFERV Env shares similar structural profiles with syncytin and other type Imore » viral envelopes, especially in the regions of N- and C-terminal heptad repeats (NHR and CHR) which were crucial for membrane fusion. We expressed the regions of N + C protein in the ZFERV Env (residues 459–567, including predicted NHR and CHR) to characterize the fusion core structure. We found N + C protein could form a stable coiled-coil trimer that consists of three helical NHR regions forming a central trimeric core, and three helical CHR regions packing into the grooves on the surface of the central core. The structural characterization of the fusion core revealed the possible mechanism of fusion mediated by ZFERV Env. These results gave comprehensive explanation of how the ancient virus infects the zebrafish and integrates into the genome million years ago, and showed a rational clue for discovery of physiological significance (e.g., medicate cell–cell fusion). - Highlights: • ZFERV Env shares similar structural profiles with syncytin and other type I viral envelopes. • The fusion core of ZFERV Env forms stable coiled-coil trimer including three NHRs and three CHRs. • The structural mechanism of viral entry mediated by ZFERV Env is disclosed. • The results are helpful for further discovery of physiological function of ZFERV Env in zebrafish.« less

Detection of receptor-specific murine leukemia virus binding to cells by immunofluorescence analysis.

PubMed Central

Kadan, M J; Sturm, S; Anderson, W F; Eglitis, M A

1992-01-01

Four classes of murine leukemia virus (MuLV) which display distinct cellular tropisms and bind to different retrovirus receptors to initiate virus infection have been described. In the present study, we describe a rapid, sensitive immunofluorescence assay useful for characterizing the initial binding of MuLV to cells. By using the rat monoclonal antibody 83A25 (L. H. Evans, R. P. Morrison, F. G. Malik, J. Portis, and W. J. Britt, J. Virol. 64:6176-6183, 1990), which recognizes an epitope of the envelope gp70 molecule common to the different classes of MuLV, it is possible to analyse the binding of ecotropic, amphotropic, or xenotropic MuLV by using only a single combination of primary and secondary antibodies. The MuLV binding detected by this assay is envelope receptor specific and matches the susceptibility to infection determined for cells from a variety of species. The binding of amphotropic MuLV to NIH 3T3 cells was shown to be rapid, saturable, and temperature dependent. Chinese hamster ovary (CHO-K1) cells normally lack the ability to bind ecotropic virus and are not infectible by ecotropic vectors. Expression of the cloned ecotropic retrovirus receptor gene (Rec) in CHO-K1 cells confers high levels of ecotropic virus-specific binding and confers susceptibility to infection. Characterization of MuLV binding to primary cells may provide insight into the infectibility of cells by retroviruses and aid in the selection of appropriate vectors for gene transfer experiments. PMID:1312632

Molecular characterization of a novel gammaretrovirus in killer whales (Orcinus orca).

PubMed

Lamere, Sarah A; St Leger, Judy A; Schrenzel, Mark D; Anthony, Simon J; Rideout, Bruce A; Salomon, Daniel R

2009-12-01

There are currently no published data documenting the presence of retroviruses in cetaceans, though the occurrences of cancers and immunodeficiency states suggest the potential. We examined tissues from adult killer whales and detected a novel gammaretrovirus by degenerate PCR. Reverse transcription-PCR also demonstrated tissue and serum expression of retroviral mRNA. The full-length sequence of the provirus was obtained by PCR, and a TaqMan-based copy number assay did not demonstrate evidence of productive infection. PCR on blood samples from 11 healthy captive killer whales and tissues from 3 free-ranging animals detected the proviral DNA in all tissues examined from all animals. A survey of multiple cetacean species by PCR for gag, pol, and env sequences showed homologs of this virus in the DNA of eight species of delphinids, pygmy and dwarf sperm whales, and harbor porpoises, but not in beluga or fin whales. Analysis of the bottlenose dolphin genome revealed two full-length proviral sequences with 97.4% and 96.9% nucleotide identity to the killer whale gammaretrovirus. The results of single-cell PCR on killer whale sperm and Southern blotting are also consistent with the conclusion that the provirus is endogenous. We suggest that this gammaretrovirus entered the delphinoid ancestor's genome before the divergence of modern dolphins or that an exogenous variant existed following divergence that was ultimately endogenized. However, the transcriptional activity demonstrated in tissues and the nearly intact viral genome suggest a more recent integration into the killer whale genome, favoring the latter hypothesis. The proposed name for this retrovirus is killer whale endogenous retrovirus.

Calorie restriction suppresses subgenomic mink cytopathic focus-forming murine leukemia virus transcription and frequency of genomic expression while impairing lymphoma formation.

PubMed Central

Shields, B A; Engelman, R W; Fukaura, Y; Good, R A; Day, N K

1991-01-01

Calorie restriction suppresses mammary proviral mRNA expression and protooncogene activation in breast tumor-prone C3H/Ou mice while inhibiting tumor formation. To determine whether the beneficial effects of chronic energy-intake restriction (CEIR) can be extended to an organ site of retrovirus-induced tumorigenesis where the dynamics of growth and sexual maturity are not paramount as they are in breast tissue, calorie restriction of 40% was imposed on thymic lymphoma-prone AKR mice when 4 weeks old. Recombination between various murine leukemia virus (MuLV) mRNAs, resulting in the generation of an 8.4-kilobase genomic-length transcript with mink cytopathic focus-forming (MCF) characteristics, is considered the proximal retroviral event in AKR lymphomagenesis. Thymic expression of subgenomic MCF MuLV mRNA was uniformly suppressed among 6- and 8-week-old CEIR mice (P less than 0.02). This suppression of MuLV transcription preceded a 25% reduction in the appearance of genomic-length MCF transcripts among CEIR mice and a 28% reduction in cumulative lymphoma mortality. The latency to median tumor incidence was extended greater than 3 months by calorie restriction, and median lifespan was extended approximately 50%. Survival curves for the full-fed and CEIR dietary cohorts were found to be significantly different (P less than 0.0001), with full-fed mice experiencing a 3 times greater risk of lymphoma mortality. These findings extend the known range of pathologic states influenced by CEIR in inbred mice and show that retroviral mechanisms involved in generation of lymphoid malignancy can be significantly impaired by calorie restriction. Images PMID:1763029

High prevalences and a wide genetic diversity of simian retroviruses in non-human primate bushmeat in rural areas of the Democratic Republic of Congo

PubMed Central

Steve, Ahuka-Mundeke; Ahidjo, Ayouba; Placide, Mbala-Kingebeni; Caroline, Foncelle; Mukulumanya, Mubonga; Simon-Pierre, Ndimbo-Kumugo; Octavie, Lunguya-Metila; Valentin, Mbenzo-Abokome; Jean-Jacques, Muyembe-Tamfum; Eric, Delaporte; Martine, Peeters

2017-01-01

Like the majority of emerging infectious diseases, HIV and HTLV are of zoonotic origin. Here we assess the risk of cross-species transmissions of their simian counterparts, SIV and STLV, from nonhuman primates (NHP) to humans in the Democratic Republic of Congo (DRC). A total of 331 samples, derived from NHP bushmeat, were collected as dried blood spots (DBS, n=283) or as tissue samples (n=36) at remote forest sites in northern and eastern DRC. SIV antibody prevalences in DBS were estimated with a novel high throughput immune assay with antigens representing the actual known diversity of HIV/SIV lineages. Antibody positive samples were confirmed by PCR and sequence analysis. Screening for STLV infection was done with universal primers in tax and new strains were further characterized in LTR. SIV and STLV infection in tissue samples was done by PCR only. Overall, 5% and 15.4% of NHP bushmeat was infected with SIV and STLV respectively. A new SIV lineage was identified in Allen's swamp monkeys (Allenopithecus nigroviridis). Three new STLV-1 subtypes were identified in Allen's swamp monkeys (Allenopithecus nigroviridis), blue monkeys (Cercopithecus mitis), red tailed guenons (Cercopithecus ascanius schmidti) and agile mangabeys (Cercocebus agilis). SIV and STLV prevalences varied according to species and geographic region. Our study illustrates clearly, even on a small sample size from a limited number of geograhic areas, that our knowledge on the genetic diversity and geographic distribution of simian retroviruses is still limited and that humans continue to be exposed to relative high proportions on infected NHP bushmeat. PMID:28050688

A Long Terminal Repeat-Containing Retrotransposon of Schizosaccharomyces pombe Expresses a Gag-Like Protein That Assembles into Virus-Like Particles Which Mediate Reverse Transcription

PubMed Central

Teysset, Laure; Dang, Van-Dinh; Kim, Min Kyung; Levin, Henry L.

2003-01-01

The Tf1 element of Schizosaccharomyces pombe is a long terminal repeat-containing retrotransposon that encodes functional protease, reverse transcriptase, and integrase proteins. Although these proteins are known to be necessary for protein processing, reverse transcription, and integration, respectively, the function of the protein thought to be Gag has not been determined. We present here the first electron microscopy of Tf1 particles. We tested whether the putative Gag of Tf1 was required for particle formation, packaging of RNA, and reverse transcription. We generated deletions of 10 amino acids in each of the four hydrophilic domains of the protein and found that all four mutations reduced transposition activity. The N-terminal deletion removed a nuclear localization signal and inhibited nuclear import of the transposon. The two mutations in the center of Gag destabilized the protein and resulted in no virus-like particles. The C-terminal deletion caused a defect in RNA packaging and, as a result, low levels of cDNA. The electron microscopy of cells expressing a truncated Tf1 showed that Gag alone was sufficient for the formation of virus-like particles. Taken together, these results indicate that Tf1 encodes a Gag protein that is a functional equivalent of the Gag proteins of retroviruses. PMID:12692246

Endogenous Retrovirus EAV-HP Linked to Blue Egg Phenotype in Mapuche Fowl

PubMed Central

Alcalde, José A.; Wang, Chen; Han, Jian-Lin; Gongora, Jaime; Gourichon, David; Tixier-Boichard, Michèle; Hanotte, Olivier

2013-01-01

Oocyan or blue/green eggshell colour is an autosomal dominant trait found in native chickens (Mapuche fowl) of Chile and in some of their descendants in European and North American modern breeds. We report here the identification of an endogenous avian retroviral (EAV-HP) insertion in oocyan Mapuche fowl and European breeds. Sequencing data reveals 100% retroviral identity between the Mapuche and European insertions. Quantitative real-time PCR analysis of European oocyan chicken indicates over-expression of the SLCO1B3 gene (P<0.05) in the shell gland and oviduct. Predicted transcription factor binding sites in the long terminal repeats (LTR) indicate AhR/Ar, a modulator of oestrogen, as a possible promoter/enhancer leading to reproductive tissue-specific over-expression of the SLCO1B3 gene. Analysis of all jungle fowl species Gallus sp. supports the retroviral insertion to be a post-domestication event, while identical LTR sequences within domestic chickens are in agreement with a recent de novo mutation. PMID:23990950

Endogenous retrovirus EAV-HP linked to blue egg phenotype in Mapuche fowl.

PubMed

Wragg, David; Mwacharo, Joram M; Alcalde, José A; Wang, Chen; Han, Jian-Lin; Gongora, Jaime; Gourichon, David; Tixier-Boichard, Michèle; Hanotte, Olivier

2013-01-01

Oocyan or blue/green eggshell colour is an autosomal dominant trait found in native chickens (Mapuche fowl) of Chile and in some of their descendants in European and North American modern breeds. We report here the identification of an endogenous avian retroviral (EAV-HP) insertion in oocyan Mapuche fowl and European breeds. Sequencing data reveals 100% retroviral identity between the Mapuche and European insertions. Quantitative real-time PCR analysis of European oocyan chicken indicates over-expression of the SLCO1B3 gene (P<0.05) in the shell gland and oviduct. Predicted transcription factor binding sites in the long terminal repeats (LTR) indicate AhR/Ar, a modulator of oestrogen, as a possible promoter/enhancer leading to reproductive tissue-specific over-expression of the SLCO1B3 gene. Analysis of all jungle fowl species Gallus sp. supports the retroviral insertion to be a post-domestication event, while identical LTR sequences within domestic chickens are in agreement with a recent de novo mutation.

Role of Accessory Proteins of HTLV-1 in Viral Replication, T Cell Activation, and Cellular Gene Expression

PubMed Central

Michael, Bindhu; Nair, Amithraj; Lairmore, Michael D.

2010-01-01

Human T-cell lymphotropic virus type 1 (HTLV-1), causes adult T cell leukemia/lymphoma (ATLL), and initiates a variety of immune mediated disorders. The viral genome encodes common structural and enzymatic proteins characteristic of all retroviruses and utilizes alternative splicing and alternate codon usage to make several regulatory and accessory proteins encoded in the pX region (pX ORF I to IV). Recent studies indicate that the accessory proteins p12I, p27I, p13II, and p30II, encoded by pX ORF I and II, contribute to viral replication and the ability of the virus to maintain typical in vivo expression levels. Proviral clones that are mutated in either pX ORF I or II, while fully competent in cell culture, are severely limited in their replicative capacity in a rabbit model. These HTLV-1 accessory proteins are critical for establishment of viral infectivity, enhance T- lymphocyte activation and potentially alter gene transcription and mitochondrial function. HTLV-1 pX ORF I expression is critical to the viral infectivity in resting primary lymphocytes suggesting a role for the calcineurin-binding protein p12I in lymphocyte activation. The endoplasmic reticulum and cis-Golgi localizing p12I activates NFAT, a key T cell transcription factor, through calcium-mediated signaling pathways and may lower the threshold of lymphocyte activation via the JAK/STAT pathway. In contrast p30II localizes to the nucleus and represses viral promoter activity, but may regulate cellular gene expression through p300/CBP or related co-activators of transcription. The mitochondrial localizing p13II induces morphologic changes in the organelle and may influence energy metabolism infected cells. Future studies of the molecular details HTLV-1 “accessory” proteins interactions will provide important new directions for investigations of HTLV-1 and related viruses associated with lymphoproliferative diseases. Thus, the accessory proteins of HTLV-1, once thought to be dispensable for

A novel endogenous betaretrovirus group characterized from polar bears (Ursus maritimus) and giant pandas (Ailuropoda melanoleuca).

PubMed

Mayer, Jens; Tsangaras, Kyriakos; Heeger, Felix; Avila-Arcos, María; Stenglein, Mark D; Chen, Wei; Sun, Wei; Mazzoni, Camila J; Osterrieder, Nikolaus; Greenwood, Alex D

2013-08-15

Transcriptome analysis of polar bears (Ursus maritimus) yielded sequences with highest similarity to the human endogenous retrovirus group HERV-K(HML-2). Further analysis of the polar bear draft genome identified an endogenous betaretrovirus group comprising 26 proviral copies and 231 solo LTRs. Molecular dating indicates the group originated before the divergence of bears from a common ancestor but is not present in all carnivores. Closely related sequences were identified in the giant panda (Ailuropoda melanoleuca) and characterized from its genome. We have designated the polar bear and giant panda sequences U. maritimus endogenous retrovirus (UmaERV) and A. melanoleuca endogenous retrovirus (AmeERV), respectively. Phylogenetic analysis demonstrated that the bear virus group is nested within the HERV-K supergroup among bovine and bat endogenous retroviruses suggesting a complex evolutionary history within the HERV-K group. All individual remnants of proviral sequences contain numerous frameshifts and stop codons and thus, the virus is likely non-infectious. Copyright © 2013 Elsevier Inc. All rights reserved.

A novel endogenous betaretrovirus group characterized from polar bears (Ursus maritimus) and giant pandas (Ailuropoda melanoleuca)

PubMed Central

Mayer, Jens; Tsangaras, Kyriakos; Heeger, Felix; Ávila-Arcos, Maria; Stenglein, Mark D.; Chen, Wei; Sun, Wei; Mazzoni, Camila; Osterrieder, Nikolaus; Greenwood, Alex D.

2013-01-01

Transcriptome analysis of polar bears (Ursus maritimus) yielded sequences with highest similarity to the human endogenous retrovirus group HERV-K(HML-2). Further analysis of the polar bear draft genome identified an endogenous betaretrovirus group comprising 26 proviral copies and 231 solo LTRs. Molecular dating indicates the group originated before the divergence of bears from a common ancestor but is not present in all carnivores. Closely related sequences were identified in the giant panda (Ailuropoda melanoleuca) and characterized from its genome. We have designated the polar bear and giant panda sequences Ursus maritimus endogenous retrovirus (UmaERV) and Ailuropoda melanoleuca endogenous retrovirus (AmeERV), respectively. Phylogenetic analysis demonstrated that the bear virus group is nested within the HERV-K supergroup among bovine and bat endogenous retroviruses suggesting a complex evolutionary history within the HERV-K group. All individual remnants of proviral sequences contain numerous frameshifts and stop codons and thus, the virus is likely non-infectious. PMID:23725819

Lineage-specific expansions of retroviral insertions within the genomes of African great apes but not humans and orangutans.

PubMed

Yohn, Chris T; Jiang, Zhaoshi; McGrath, Sean D; Hayden, Karen E; Khaitovich, Philipp; Johnson, Matthew E; Eichler, Marla Y; McPherson, John D; Zhao, Shaying; Pääbo, Svante; Eichler, Evan E

2005-04-01

Retroviral infections of the germline have the potential to episodically alter gene function and genome structure during the course of evolution. Horizontal transmissions between species have been proposed, but little evidence exists for such events in the human/great ape lineage of evolution. Based on analysis of finished BAC chimpanzee genome sequence, we characterize a retroviral element (Pan troglodytes endogenous retrovirus 1 [PTERV1]) that has become integrated in the germline of African great ape and Old World monkey species but is absent from humans and Asian ape genomes. We unambiguously map 287 retroviral integration sites and determine that approximately 95.8% of the insertions occur at non-orthologous regions between closely related species. Phylogenetic analysis of the endogenous retrovirus reveals that the gorilla and chimpanzee elements share a monophyletic origin with a subset of the Old World monkey retroviral elements, but that the average sequence divergence exceeds neutral expectation for a strictly nuclear inherited DNA molecule. Within the chimpanzee, there is a significant integration bias against genes, with only 14 of these insertions mapping within intronic regions. Six out of ten of these genes, for which there are expression data, show significant differences in transcript expression between human and chimpanzee. Our data are consistent with a retroviral infection that bombarded the genomes of chimpanzees and gorillas independently and concurrently, 3-4 million years ago. We speculate on the potential impact of such recent events on the evolution of humans and great apes.

U1snRNP-mediated suppression of polyadenylation in conjunction with the RNA structure controls poly (A) site selection in foamy viruses

PubMed Central

2013-01-01

Background During reverse transcription, retroviruses duplicate the long terminal repeats (LTRs). These identical LTRs carry both promoter regions and functional polyadenylation sites. To express full-length transcripts, retroviruses have to suppress polyadenylation in the 5′LTR and activate polyadenylation in the 3′LTR. Foamy viruses have a unique LTR structure with respect to the location of the major splice donor (MSD), which is located upstream of the polyadenylation signal. Results Here, we describe the mechanisms of foamy viruses regulating polyadenylation. We show that binding of the U1 small nuclear ribonucleoprotein (U1snRNP) to the MSD suppresses polyadenylation at the 5′LTR. In contrast, polyadenylation at the 3′LTR is achieved by adoption of a different RNA structure at the MSD region, which blocks U1snRNP binding and furthers RNA cleavage and subsequent polyadenylation. Conclusion Recently, it was shown that U1snRNP is able to suppress the usage of intronic cryptic polyadenylation sites in the cellular genome. Foamy viruses take advantage of this surveillance mechanism to suppress premature polyadenylation at the 5’end of their RNA. At the 3’end, Foamy viruses use a secondary structure to presumably block access of U1snRNP and thereby activate polyadenylation at the end of the genome. Our data reveal a contribution of U1snRNP to cellular polyadenylation site selection and to the regulation of gene expression. PMID:23718736

Nuclear Export and Expression of Human T-Cell Leukemia Virus Type 1 tax/rex mRNA Are RxRE/Rex Dependent

PubMed Central

Bai, X. T.; Sinha-Datta, U.; Ko, N. L.; Bellon, M.

2012-01-01

Human T-cell leukemia virus type 1 (HTLV-1) is a complex retrovirus associated with the lymphoproliferative disease adult T-cell leukemia/lymphoma (ATL) and the neurodegenerative disorder tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM). Replication of HTLV-1 is under the control of two major trans-acting proteins, Tax and Rex. Previous studies suggested that Tax activates transcription from the viral long terminal repeat (LTR) through recruitment of cellular CREB and transcriptional coactivators. Other studies reported that Rex acts posttranscriptionally and allows the cytoplasmic export of unspliced or incompletely spliced viral mRNAs carrying gag/pol and env only. As opposed to HIV's Rev-responsive element (RRE), the Rex-responsive element (RxRE) is present in all viral mRNAs in HTLV-1. However, based on indirect observations, it is believed that nuclear export and expression of the doubly spliced tax/rex RNA are Rex independent. In this study, we demonstrate that Rex does stimulate Tax expression, through nuclear-cytoplasmic export of the tax/rex RNA, even though a Rex-independent basal export mechanism exists. This effect was dependent upon the RxRE element and the RNA-binding activity of Rex. In addition, Rex-mediated export of tax/rex RNA was CRM1 dependent and inhibited by leptomycin B treatment. RNA immunoprecipitation (RNA-IP) experiments confirmed Rex binding to the tax/rex RNA in both transfected cells with HTLV-1 molecular clones and HTLV-1-infected T cells. Since both Rex and p30 interact with the tax/rex RNA and with one another, this may offer a temporal and dynamic regulation of HTLV-1 replication. Our results shed light on HTLV-1 replication and reveal a more complex regulatory network than previously anticipated. PMID:22318152

Prospects for inhibiting the post-transcriptional regulation of gene expression in hepatitis B virus

PubMed Central

Chen, Augustine; Panjaworayan T-Thienprasert, Nattanan; Brown, Chris M

2014-01-01

There is a continuing need for novel antivirals to treat hepatitis B virus (HBV) infection, as it remains a major health problem worldwide. Ideally new classes of antivirals would target multiple steps in the viral lifecycle. In this review, we consider the steps in which HBV RNAs are processed, exported from the nucleus and translated. These are often overlooked steps in the HBV life-cycle. HBV, like retroviruses, incorporates a number of unusual steps in these processes, which use a combination of viral and host cellular machinery. Some of these unusual steps deserve a closer scrutiny. They may provide alternative targets to existing antiviral therapies, which are associated with increasing drug resistance. The RNA post-transcriptional regulatory element identified 20 years ago promotes nucleocytoplasmic export of all unspliced HBV RNAs. There is evidence that inhibition of this step is part of the antiviral action of interferon. Similarly, the structured RNA epsilon element situated at the 5’ end of the polycistronic HBV pregenomic RNA also performs key roles during HBV replication. The pregenomic RNA, which is the template for translation of both the viral core and polymerase proteins, is also encapsidated and used in replication. This complex process, regulated at the epsilon element, also presents an attractive antiviral target. These RNA elements that mediate and regulate gene expression are highly conserved and could be targeted using novel strategies employing RNAi, miRNAs or aptamers. Such approaches targeting these functionally constrained genomic regions should avoid escape mutations. Therefore understanding these regulatory elements, along with providing potential targets, may also facilitate the development of other new classes of antiviral drugs. PMID:25009369

Monitoring the Transcriptional Activity of Human Endogenous Retroviral HERV-W Family Using PNA Strand Invasion into Double-Stranded DNA.

PubMed

Machnik, Grzegorz; Skudrzyk, Estera; Bułdak, Łukasz; Ruczyński, Jarosław; Kozłowska, Agnieszka; Mucha, Piotr; Rekowski, Piotr; Szkróbka, Witold; Basiak, Marcin; Bołdys, Aleksandra; Sławska, Helena; Okopień, Bogusław

2018-02-01

In the presented assay, we elaborated a method for distinguishing sequences that are genetically closely related to each other. This is particularly important in a situation where a fine balance of the allele abundance is a point of research interest. We developed a peptide nucleic acid (PNA) strand invasion technique for the differentiation between multiple sclerosis-associated retrovirus (MSRV) and ERVWE1 sequences, both molecularly similar, belonging to the human endogenous retrovirus HERV-W family. We have found that this method may support the PCR technique in screening for minor alleles which, in certain conditions, may be undetected by the standard PCR technique. We performed the analysis of different ERVWE1 and MSRV template mixtures ranging from 0 to 100% of ERVWE1 in the studied samples, finding the linear correlation between template composition and signal intensity of final reaction products. Using the PNA strand invasion assay, we were able to estimate the relative ERVWE1 expression level in human specimens such as U-87 MG, normal human astrocytes cell lines and placental tissue. The results remained in concordance with those obtained by semi-quantitative or quantitative PCR.

Preclinical Corrective Gene Transfer in Xeroderma Pigmentosum Human Skin Stem Cells

PubMed Central

Warrick, Emilie; Garcia, Marta; Chagnoleau, Corinne; Chevallier, Odile; Bergoglio, Valérie; Sartori, Daniela; Mavilio, Fulvio; Angulo, Jaime F; Avril, Marie-Françoise; Sarasin, Alain; Larcher, Fernando; Del Rio, Marcela; Bernerd, Françoise; Magnaldo, Thierry

2012-01-01

Xeroderma pigmentosum (XP) is a devastating disease associated with dramatic skin cancer proneness. XP cells are deficient in nucleotide excision repair (NER) of bulky DNA adducts including ultraviolet (UV)-induced mutagenic lesions. Approaches of corrective gene transfer in NER-deficient keratinocyte stem cells hold great hope for the long-term treatment of XP patients. To face this challenge, we developed a retrovirus-based strategy to safely transduce the wild-type XPC gene into clonogenic human primary XP-C keratinocytes. De novo expression of XPC was maintained in both mass population and derived independent candidate stem cells (holoclones) after more than 130 population doublings (PD) in culture upon serial propagation (>1040 cells). Analyses of retrovirus integration sequences in isolated keratinocyte stem cells suggested the absence of adverse effects such as oncogenic activation or clonal expansion. Furthermore, corrected XP-C keratinocytes exhibited full NER capacity as well as normal features of epidermal differentiation in both organotypic skin cultures and in a preclinical murine model of human skin regeneration in vivo. The achievement of a long-term genetic correction of XP-C epidermal stem cells constitutes the first preclinical model of ex vivo gene therapy for XP-C patients. PMID:22068429

Mass Determination of Rous Sarcoma Virus Virions by Scanning Transmission Electron Microscopy

PubMed Central

Vogt, Volker M.; Simon, Martha N.

1999-01-01

The internal structural protein of retroviruses, Gag, comprises most of the mass of the virion, and Gag itself can give rise to virus-like particles when expressed in appropriate cells. Previously the stoichiometry of Gag in virions was inferred from indirect measurements carried out 2 decades ago. We now have directly determined the masses of individual particles of the prototypic avian retrovirus, Rous sarcoma virus (RSV), by using scanning transmission electron microscopy. In this technique, the number of scattered electrons in the dark-field image integrated over an individual freeze-dried virus particle on a grid is directly proportional to its mass. The RSV virions had a mean mass of 2.5 × 108 Da, corresponding to about 1,500 Gag molecules per virion. The population of virions was not homogeneous, with about one-third to two-thirds of the virions deviating from the mean by more than 10% of the mass in two respective preparations. The mean masses for virions carrying genomes of 7.4 or 9.3 kb were indistinguishable, suggesting that mass variability is not due to differences in RNA incorporation. PMID:10400808

Evidence For Multiple Cell Death Pathways during Development of Experimental Cytomegalovirus Retinitis in Mice with Retrovirus-Induced Immunosuppression: Apoptosis, Necroptosis, and Pyroptosis

PubMed Central

Chien, Hsin

2012-01-01

AIDS-related human cytomegalovirus (HCMV) retinitis remains a major ophthalmologic problem worldwide. Although this sight-threatening disease is well characterized clinically, many pathogenic issues remain unresolved, among them a basic understanding of the relative roles of cell death pathways during development of retinal tissue destruction. Using an established model of experimental murine cytomegalovirus (MCMV) retinitis in mice with retrovirus-induced immunosuppression (MAIDS), we initially investigated MCMV-infected eyes for evidence of apoptosis-associated molecules in mice with MAIDS of 4 weeks' (MAIDS-4) and 10 weeks' (MAIDS-10) duration, which were resistant and susceptible to retinal disease, respectively, but which harbored equivalent amounts of infectious MCMV. Whereas MCMV-infected eyes of MAIDS-4 mice showed little evidence of apoptosis-associated molecules, MCMV-infected eyes of MAIDS-10 mice showed significant amounts of tumor necrosis factor alpha (TNF-α), TNF receptors 1 and 2, active caspase 8, active caspase 3, TNF-related apoptosis-inducing ligand (TRAIL), TRAIL-R(DR5), Fas, and Fas ligand mRNAs and/or proteins, all detected at peak amounts prior to development of most severe retinal disease. Immunohistochemical staining showed macrophages, granulocytes (neutrophils), Müller cells, and microglial cells as TNF-α sources. Remarkably, quantification of apoptosis by terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assay suggested that apoptosis contributed minimally to retinal disease in MCMV-infected eyes of MAIDS-10 mice. Subsequent studies demonstrated that MCMV-infected eyes of MAIDS-10 mice, but not MAIDS-4 mice, showed evidence of significant increases in molecules associated with two additional cell death pathways, necroptosis (receptor-interacting protein 1 [RIP1] and RIP3 mRNAs) and pyroptosis (caspase 1, interleukin 1β [IL-1β], and IL-18 mRNAs). We conclude that apoptosis, necroptosis, and

GCN5 regulates the activation of PI3K/Akt survival pathway in B cells exposed to oxidative stress via controlling gene expressions of Syk and Btk.

PubMed

Kikuchi, Hidehiko; Kuribayashi, Futoshi; Takami, Yasunari; Imajoh-Ohmi, Shinobu; Nakayama, Tatsuo

2011-02-25

Histone acetyltransferase(s) (HATs) are involved in the acetylation of core histones, which is an important event for transcription regulation through alterations in the chromatin structure in eukaryotes. General control non-depressible 5 (GCN5) was first identified as a global coactivator and transcription-related HAT. Here we report that GCN5 regulates the activation of phosphatidylinositol 3-kinase (PI3K)/acutely transforming retrovirus AKT8 in rodent T cell lymphoma (Akt) survival pathway in B cells exposed to oxidative stress via controlling gene expressions of spleen tyrosine kinase (Syk) and Bruton's tyrosine kinase (Btk). The GCN5-deficiency remarkably caused apoptotic cell death by treatment with exogenous hydrogen peroxide (H(2)O(2)) in chicken DT40 cells. In GCN5-deficient DT40 cells, gene expressions of Syk and Btk, which are involved in activation of PI3K/Akt survival pathway in DT40 cells exposed to exogenous H(2)O(2), were remarkably decreased compared with those in wild type DT40 cells. In addition, phosphorylation of Akt in H(2)O(2)-treated GCN5-deficient cells was remarkably suppressed as compared to that of DT40. Chromatin immunoprecipitation assay revealed that GCN5 binds to proximal 5'-upstream regions of Syk and Btk genes in vivo. These results suggest that GCN5 takes part in transcriptional regulations of the Syk and Btk genes, and plays a key role in epigenetic regulation of PI3K/Akt survival pathway in B cells exposed to reactive oxygen species such as H(2)O(2). Copyright © 2011 Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yin, Zhiqi; Shi, Ke; Banerjee, Surajit

Integration of the reverse-transcribed viral DNA into the host genome is an essential step in the life cycle of retroviruses. Retrovirus integrase catalyses insertions of both ends of the linear viral DNA into a host chromosome. Integrase from HIV-1 and closely related retroviruses share the three-domain organization, consisting of a catalytic core domain flanked by amino- and carboxy-terminal domains essential for the concerted integration reaction. Although structures of the tetrameric integrase–DNA complexes have been reported for integrase from prototype foamy virus featuring an additional DNA-binding domain and longer interdomain linkers, the architecture of a canonical three-domain integrase bound to DNAmore » remained elusive. In this paper, we report a crystal structure of the three-domain integrase from Rous sarcoma virus in complex with viral and target DNAs. The structure shows an octameric assembly of integrase, in which a pair of integrase dimers engage viral DNA ends for catalysis while another pair of non-catalytic integrase dimers bridge between the two viral DNA molecules and help capture target DNA. The individual domains of the eight integrase molecules play varying roles to hold the complex together, making an extensive network of protein–DNA and protein–protein contacts that show both conserved and distinct features compared with those observed for prototype foamy virus integrase. Finally, our work highlights the diversity of retrovirus intasome assembly and provides insights into the mechanisms of integration by HIV-1 and related retroviruses.« less

Crystal structure of the Rous sarcoma virus intasome

DOE PAGES

Yin, Zhiqi; Shi, Ke; Banerjee, Surajit; ...

2016-02-17

Integration of the reverse-transcribed viral DNA into the host genome is an essential step in the life cycle of retroviruses. Retrovirus integrase catalyses insertions of both ends of the linear viral DNA into a host chromosome. Integrase from HIV-1 and closely related retroviruses share the three-domain organization, consisting of a catalytic core domain flanked by amino- and carboxy-terminal domains essential for the concerted integration reaction. Although structures of the tetrameric integrase–DNA complexes have been reported for integrase from prototype foamy virus featuring an additional DNA-binding domain and longer interdomain linkers, the architecture of a canonical three-domain integrase bound to DNAmore » remained elusive. In this paper, we report a crystal structure of the three-domain integrase from Rous sarcoma virus in complex with viral and target DNAs. The structure shows an octameric assembly of integrase, in which a pair of integrase dimers engage viral DNA ends for catalysis while another pair of non-catalytic integrase dimers bridge between the two viral DNA molecules and help capture target DNA. The individual domains of the eight integrase molecules play varying roles to hold the complex together, making an extensive network of protein–DNA and protein–protein contacts that show both conserved and distinct features compared with those observed for prototype foamy virus integrase. Finally, our work highlights the diversity of retrovirus intasome assembly and provides insights into the mechanisms of integration by HIV-1 and related retroviruses.« less

Persistence of antigen is required to maintain transplantation tolerance induced by genetic modification of bone marrow stem cells.

PubMed

Tian, C; Bagley, J; Iacomini, J

2006-09-01

Genetic modification of hematopoietic stem cells (HSCs) resulting in a state of molecular chimerism can be used to induce donor-specific tolerance to allografts. However, the requirements for maintaining tolerance in molecular chimeras remain unknown. Here, we examined whether long-term expression of a retrovirally encoded alloantigen in hematopoietic cells is required to maintain donor-specific tolerance in molecular chimeras. To this end, mice were reconstituted with syngeneic bone marrow transduced with retroviruses carrying the gene encoding the allogeneic MHC class I molecule Kb. Following induction of molecular chimerism, mice were depleted of cells expressing Kb by administration of the anti-Kb monoclonal antibody Y-3. Mice that were effectively depleted of cells expressing the retrovirally encoded MHC class I antigen rejected Kb disparate skin allografts. In contrast, control molecular chimeras accepted Kb disparate skin allografts indefinitely. These data suggest maintenance of tolerance in molecular chimeras requires long-term expression of retrovirally transduced alloantigen on the progeny of retrovirally transduced HSCs.

Innate Immune Recognition of HIV-1

PubMed Central

Iwasaki, Akiko

2012-01-01

In contrast to the extraordinary body of knowledge gained over the past three decades on the virology, pathogenesis, and immunology of HIV-1 infection, innate sensors that detect HIV-1 had remained elusive until recently. By virtue of integration, retroviridae makes up a substantial portion of our genome. Thus, immune strategies that deal with endogenous retroviruses are, by necessity, those of self-preservation and not of virus elimination. Some of the principles of such strategies may also apply for defense against exogenous retroviruses including HIV-1. Here, I highlight several sensors that have recently been revealed to be capable of recognizing distinct features of HIV-1 infection, while taking into account the host-retrovirus relationship that converges on avoiding pathogenic inflammatory consequences. PMID:22999945

Sites of Retroviral DNA Integration: From Basic Research to Clinical Applications

PubMed Central

Serrao, Erik; Engelman, Alan N.

2016-01-01

One of the most crucial steps in the life cycle of a retrovirus is the integration of the viral DNA (vDNA) copy of the RNA genome into the genome of an infected host cell. Integration provides for efficient viral gene expression as well as for the segregation of the viral genomes to daughter cells upon cell division. Some integrated viruses are not well expressed, and cells latently infected with HIV-1 can resist the action of potent antiretroviral drugs and remain dormant for decades. Intensive research has been dedicated to understanding the catalytic mechanism of integration, as well as the viral and cellular determinants that influence integration site distribution throughout the host genome. In this review we summarize the evolution of techniques that have been used to recover and map retroviral integration sites, from the early days that first indicated that integration could occur in multiple cellular DNA locations, to current technologies that map upwards of millions of unique integration sites from single in vitro integration reactions or cell culture infections. We further review important insights gained from the use of such mapping techniques, including the monitoring of cell clonal expansion in patients treated with retrovirus-based gene therapy vectors, or AIDS patients on suppressive antiretroviral therapy (ART). These insights span from integrase (IN) enzyme sequence preferences within target DNA (tDNA) at the sites of integration, to the roles of host cellular proteins in mediating global integration distribution, to the potential relationship between genomic location of vDNA integration site and retroviral latency. PMID:26508664

Sequence comparison in the crossover region of an oncogenic avian retrovirus recombinant and its nononcogenic parent: Genetic regions that control growth rate and oncogenic potential

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tsichlis, P.N.; Donehower, L.; Hager, G.

1982-11-01

NTRE is an avian retrovirus recombinant of the endogeneous nononcogenic Rous-associated virus-0 (RAV-0) and the oncogenic, exogeneous, transformation-defective (td) Prague strain of Rous sarcoma virus B (td-PrRSV-B). Oligonucleotide mapping had shown that the recombinant virus is indistinguishable from its RAV-0 parent except for the 3'-end sequences, which were derived from td-PrRSV-B. However, the virus exhibits properties which are typical of an exogenous virus: it grows to high titers in tissue culture, and it is oncogenic in vivo. To accurately define the genetic region responsible for these properties, the authors determined the nucleotide sequences of the recombinant and its RAV-0 parentmore » by using molecular clones of their DNA. These were compared with sequences already available for PrRSV-C, a virus closely related to the exogenous parent td-PrRSV-B. The results suggested that the crossover event which generated NTRE 7 took place in a region -501 to -401 nucleotides from the 3' end of the td-PrRSV parental genome and that sequences to the right of the recombination region were responsible for its growth properties and oncogenic potential. Since the exogenous-virus-specific sequences are expected to be missing from transformation-defective mutants of the Schmidt-Ruppin strain of RSV, which, like other exogeneous viruses, grow to high tiers in tissue culture and are oncogenic in vivo, the authors concluded that the growth properties and oncogenic potential of the exogeneous viruses are determined by sequences in the U3 region of the long terminal repeat. However, the authors propose that the exogeneous-virus-specific region may play a role in determining the oncogenic spectrum of a given oncogenic virus.« less

Lentiviral infection of proliferating brain macrophages in HIV and simian immunodeficiency virus encephalitis despite sterile alpha motif and histidine-aspartate domain-containing protein 1 expression

PubMed Central

Lindgren, Allison A.; Filipowicz, Adam R.; Hattler, Julian B.; Kim, Soon Ok; Chung, Hye Kyung; Kuroda, Marcelo J.; Johnson, Edward M.; Kim, Woong-Ki

2018-01-01

Objective: HIV-1 infection of the brain and related cognitive impairment remain prevalent in HIV-1-infected individuals despite combination antiretroviral therapy. Sterile alpha motif and histidine-aspartate domain-containing protein 1 (SAMHD1) is a newly identified host restriction factor that blocks the replication of HIV-1 and other retroviruses in myeloid cells. Cell cycle-regulated phosphorylation at residue Thr592 and viral protein X (Vpx)-mediated degradation of SAMHD1 have been shown to bypass SAMHD1 restriction in vitro. Herein, we investigated expression and phosphorylation of SAMHD1 in vivo in relation to macrophage infection and proliferation during the neuropathogenesis of HIV-1 and simian immunodeficiency virus (SIV) encephalitis. Methods: Using brain and other tissues from uninfected and SIV-infected macaques with or without encephalitis, we performed immunohistochemistry, multilabel fluorescence microscopy and western blot to examine the expression, localization and phosphorylation of SAMHD1. Results: The number of SAMHD1+ nuclei increased in encephalitic brains despite the presence of Vpx. Many of these cells were perivascular macrophages, although subsets of SAMHD1+ microglia and endothelial cells were also observed. The SAMHD1+ macrophages were shown to be both infected and proliferating. Moreover, the presence of cycling SAMHD1+ brain macrophages was confirmed in the tissue of HIV-1-infected patients with encephalitis. Finally, western blot analysis of brain-protein extracts from SIV-infected macaques showed that SAMHD1 protein exists in the brain mainly as an inactive Thr592-phosphorylated form. Conclusion: The ability of SAMHD1 to act as a restriction factor for SIV/HIV in the brain is likely bypassed in proliferating brain macrophages through the phosphorylation-mediated inactivation, not Vpx-mediated degradation of SAMHD1. PMID:29698322

v-src induces clonal sarcomas and rapid metastasis following transduction with a replication-defective retrovirus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoker, A.W.; Sieweke, M.H.

1989-12-01

v-src is an effective carcinogen when expressed from Rous sarcoma virus (RSV) in vivo. Whereas RSV tumors require sustained oncogene expression, their growth is largely a balance between viral recruitment of tissues and host immune destruction of infected cells. The authors have therefore examined the tumorigenic potential of v-src in the absence of viral recruitment and viral antigen expression. v-src was introduced with high efficiency into chicken wing web tissues using replication-defective (rd) retroviral vectors. Clonal sarcomas were induced rapidly, and furthermore, v-src potentiated metastatic progression in {approx} 0.1%-1% of tumor clones with unexpectedly short latency. rd vectors proved effectivemore » not only in transducing v-src into tissues but also as insertional markers of tumor clonality. The rd vector present in most primary and metastatic tumors was a highly truncated form of RSV derived by viral transmission of spliced v-src mRNA; this vector should thus avoid viral recruitment and host anti-viral immune reaction through its complete lack of viral structural genes. Under such conditions v-src maintains strong carcinogenicity in vivo when restricted to clonal tumor growth and can confer rapid metastatic potential on a discrete subset of tumor clones.« less

Transplantation of autoimmune regulator-encoding bone marrow cells delays the onset of experimental autoimmune encephalomyelitis.

PubMed

Ko, Hyun-Ja; Kinkel, Sarah A; Hubert, François-Xavier; Nasa, Zeyad; Chan, James; Siatskas, Christopher; Hirubalan, Premila; Toh, Ban-Hock; Scott, Hamish S; Alderuccio, Frank

2010-12-01

The autoimmune regulator (AIRE) promotes "promiscuous" expression of tissue-restricted antigens (TRA) in thymic medullary epithelial cells to facilitate thymic deletion of autoreactive T-cells. Here, we show that AIRE-deficient mice showed an earlier development of myelin oligonucleotide glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE). To determine the outcome of ectopic Aire expression, we used a retroviral transduction system to over-express Aire in vitro, in cell lines and in bone marrow (BM). In the cell lines that included those of thymic medullary and dendritic cell origin, ectopically expressed Aire variably promoted expression of TRA including Mog and Ins2 (proII) autoantigens associated, respectively, with the autoimmune diseases multiple sclerosis and type 1 diabetes. BM chimeras generated from BM transduced with a retrovirus encoding Aire displayed elevated levels of Mog and Ins2 expression in thymus and spleen. Following induction of EAE with MOG(35-55), transplanted mice displayed significant delay in the onset of EAE compared with control mice. To our knowledge, this is the first example showing that in vivo ectopic expression of AIRE can modulate TRA expression and alter autoimmune disease development. Copyright © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Interferon-α (IFN-α) suppresses HTLV-1 gene expression and cell cycling, while IFN-α combined with zidovudin induces p53 signaling and apoptosis in HTLV-1-infected cells

PubMed Central

2013-01-01

Background Human T-cell leukemia virus type-1 (HTLV-1) is the causative retrovirus of adult T-cell leukemia/lymphoma (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). HTLV-1 gene expression is maintained at low levels in vivo by unknown mechanisms. A combination therapy of interferon-α (IFN-α) and zidovudin (AZT) shows therapeutic effects in ATL patients, although its mechanism is also obscure. We previously found that viral gene expression in IL-2-dependent HTLV-1-infected T-cells (ILTs) derived from ATL patients was markedly suppressed by stromal cells through a type I IFN response. Here, we investigated the effects of IFN-α with or without AZT on viral gene expression and cell growth in ILTs. Results ILTs expressed variable but lower amounts of HTLV-1 Tax protein than HTLV-1-transformed HUT102 cells. Following the addition of IFN-α, the amounts of HTLV-1 p19 in the supernatants of these cells decreased in three days, while HTLV-1 gene expression decreased only in ILTs but not HUT102 cells. IFN-α also suppressed the spontaneous HTLV-1 induction in primary ATL cells cultured for 24 h. A time course study using ILTs revealed that the levels of intracellular Tax proteins decreased in the first 24 h after addition of IFN-α, before the reduction in HTLV-1 mRNA levels. The initial decreases of Tax protein following IFN-α treatment were observed in 6 of 7 ILT lines tested, although the reduction rates varied among ILT lines. An RNA-dependent protein kinase (PKR)-inhibitor reversed IFN-mediated suppression of Tax in ILTs. IFN-α also induced cell cycle arrest at the G0/G1 phase and suppressed NF-κB activities in these cells. AZT alone did not affect HTLV-1 gene expression, cell viability or NF-κB activities. AZT combined with IFN-α markedly induced cell apoptosis associated with phosphorylation of p53 and induction of p53-responsive genes in ILTs. Conclusions IFN-α suppressed HTLV-1 gene expression at least through a PKR

Hili Inhibits HIV Replication in Activated T Cells.

PubMed

Peterlin, B Matija; Liu, Pingyang; Wang, Xiaoyun; Cary, Daniele; Shao, Wei; Leoz, Marie; Hong, Tian; Pan, Tao; Fujinaga, Koh

2017-06-01

P-element-induced wimpy-like (Piwil) proteins restrict the replication of mobile genetic elements in the germ line. They are also expressed in many transformed cell lines. In this study, we discovered that the human Piwil 2 (Hili) protein can also inhibit HIV replication, especially in activated CD4 + T cells that are the preferred target cells for this virus in the infected host. Although resting cells did not express Hili, its expression was rapidly induced following T cell activation. In these cells and transformed cell lines, depletion of Hili increased levels of viral proteins and new viral particles. Further studies revealed that Hili binds to tRNA. Some of the tRNAs represent rare tRNA species, whose codons are overrepresented in the viral genome. Targeting tRNA Arg (UCU) with an antisense oligonucleotide replicated effects of Hili and also inhibited HIV replication. Finally, Hili also inhibited the retrotransposition of the endogenous intracysternal A particle (IAP) by a similar mechanism. Thus, Hili joins a list of host proteins that inhibit the replication of HIV and other mobile genetic elements. IMPORTANCE Piwil proteins inhibit the movement of mobile genetic elements in the germ line. In their absence, sperm does not form and male mice are sterile. This inhibition is thought to occur via small Piwi-interacting RNAs (piRNAs). However, in some species and in human somatic cells, Piwil proteins bind primarily to tRNA. In this report, we demonstrate that human Piwil proteins, especially Hili, not only bind to select tRNA species, including rare tRNAs, but also inhibit HIV replication. Importantly, T cell activation induces the expression of Hili in CD4 + T cells. Since Hili also inhibited the movement of an endogenous retrovirus (IAP), our finding shed new light on this intracellular resistance to exogenous and endogenous retroviruses as well as other mobile genetic elements. Copyright © 2017 American Society for Microbiology.

Comparing the landcapes of common retroviral insertion sites across tumor models

NASA Astrophysics Data System (ADS)

Weishaupt, Holger; Čančer, Matko; Engström, Cristopher; Silvestrov, Sergei; Swartling, Fredrik J.

2017-01-01

Retroviral tagging represents an important technique, which allows researchers to screen for candidate cancer genes. The technique is based on the integration of retroviral sequences into the genome of a host organism, which might then lead to the artificial inhibition or expression of proximal genetic elements. The identification of potential cancer genes in this framework involves the detection of genomic regions (common insertion sites; CIS) which contain a number of such viral integration sites that is greater than expected by chance. During the last two decades, a number of different methods have been discussed for the identification of such loci and the respective techniques have been applied to a variety of different retroviruses and/or tumor models. We have previously established a retrovirus driven brain tumor model and reported the CISs which were found based on a Monte Carlo statistics derived detection paradigm. In this study, we consider a recently proposed alternative graph theory based method for identifying CISs and compare the resulting CIS landscape in our brain tumor dataset to those obtained when using the Monte Carlo approach. Finally, we also employ the graph-based method to compare the CIS landscape in our brain tumor model with those of other published retroviral tumor models.

Autoimmune disease: A role for new anti-viral therapies?

PubMed

Dreyfus, David H

2011-12-01

Many chronic human diseases may have an underlying autoimmune mechanism. In this review, the author presents a case of autoimmune CIU (chronic idiopathic urticaria) in stable remission after therapy with a retroviral integrase inhibitor, raltegravir (Isentress). Previous reports located using the search terms "autoimmunity" and "anti-viral" and related topics in the pubmed data-base are reviewed suggesting that novel anti-viral agents such as retroviral integrase inhibitors, gene silencing therapies and eventually vaccines may provide new options for anti-viral therapy of autoimmune diseases. Cited epidemiologic and experimental evidence suggests that increased replication of epigenomic viral pathogens such as Epstein-Barr Virus (EBV) in chronic human autoimmune diseases such as rheumatoid arthritis (RA), systemic lupus Erythematosus (SLE), and multiple sclerosis (MS) may activate endogenous human retroviruses (HERV) as a pathologic mechanism. Memory B cells are the reservoir of infection of EBV and also express endogenous retroviruses, thus depletion of memory b-lymphocytes by monoclonal antibodies (Rituximab) may have therapeutic anti-viral effects in addition to effects on B-lymphocyte presentation of both EBV and HERV superantigens. Other novel anti-viral therapies of chronic autoimmune diseases, such as retroviral integrase inhibitors, could be effective, although not without risk. Copyright © 2011 Elsevier B.V. All rights reserved.

In vivo modification of retroviral gag gene-encoded polyproteins by myristic acid.

PubMed Central

Schultz, A M; Oroszlan, S

1983-01-01

It has recently been shown by mass spectral analysis (Henderson et al., Proc. Natl. Acad. Sci. U.S.A. 80:339-343, 1983) that the p15gag protein of murine leukemia viruses contains a novel post-translational modification, an amino-terminal myristyl (tetradecanoyl) amide. In this report we show that p15gag is the only structural protein to contain this fatty acid. In addition, the gag precursor polyproteins of type B, C, and D retroviruses have been examined for the presence of myristic acid by metabolic labeling and immunoprecipitation studies. In a panel of mammalian type C retroviruses we found that the precursor polyprotein Pr65gag homologs, but not the glycosylated forms (gPr80gag homologs), were specifically labeled after a 5-min incubation of infected cells with [3H]myristic acid. The gag precursor polyprotein was also labeled in mouse mammary tumor virus and in Mason-Pfizer monkey virus, but Pr76gag of Rous sarcoma virus failed to incorporate [3H]myristate. Under similar conditions, [3H]palmitate was not found to be incorporated into any viral gag proteins. Thus, myristylation appears to be a common feature of mammalian type B, C, and D retroviruses but not of avian retroviruses. Images PMID:6302307

Discovery of an endogenous Deltaretrovirus in the genome of long-fingered bats (Chiroptera: Miniopteridae).

PubMed

Farkašová, Helena; Hron, Tomáš; Pačes, Jan; Hulva, Pavel; Benda, Petr; Gifford, Robert James; Elleder, Daniel

2017-03-21

Retroviruses can create endogenous forms on infiltration into the germline cells of their hosts. These forms are then vertically transmitted and can be considered as genetic fossils of ancient viruses. All retrovirus genera, with the exception of deltaretroviruses, have had their representation identified in the host genome as a virus fossil record. Here we describe an endogenous Deltaretrovirus, identified in the germline of long-fingered bats (Miniopteridae). A single, heavily deleted copy of this retrovirus has been found in the genome of miniopterid species, but not in the genomes of the phylogenetically closest bat families, Vespertilionidae and Cistugonidae. Therefore, the endogenization occurred in a time interval between 20 and 45 million years ago. This discovery closes the last major gap in the retroviral fossil record and provides important insights into the history of deltaretroviruses in mammals.

Tolerance to MHC class II disparate allografts through genetic modification of bone marrow

PubMed Central

Jindra, Peter T.; Tripathi, Sudipta; Tian, Chaorui; Iacomini, John; Bagley, Jessamyn

2012-01-01

Induction of molecular chimerism through genetic modification of bone marrow is a powerful tool for the induction of tolerance. Here we demonstrate for the first time that expression of an allogeneic MHC class II gene in autologous bone marrow cells, resulting in a state of molecular chimerism, induces tolerance to MHC class II mismatched skin grafts, a stringent test of transplant tolerance. Reconstitution of recipients with syngeneic bone marrow transduced with retrovirus encoding H-2I-Ab (I-Ab) resulted the long-term expression of the retroviral gene product on the surface of MHC class II-expressing bone marrow derived cell types. Mechanistically, tolerance was maintained by the presence of regulatory T cells, which prevented proliferation and cytokine production by alloreactive host T cells. Thus, the introduction of MHC class II genes into bone marrow derived cells through genetic engineering results in tolerance. These results have the potential to extend the clinical applicability of molecular chimerism for tolerance induction. PMID:22833118

Retrovirally mediated correction of bone marrow-derived mesenchymal stem cells from patients with mucopolysaccharidosis type I.

PubMed

Baxter, Melissa A; Wynn, Robert F; Deakin, Jonathan A; Bellantuono, Ilaria; Edington, Kirsten G; Cooper, Alan; Besley, Guy T N; Church, Heather J; Wraith, J Ed; Carr, Trevor F; Fairbairn, Leslie J

2002-03-01

We have investigated the utility of bone marrow-derived mesenchymal stem cells (MSCs) as targets for gene therapy of the autosomal recessive disorder mucopolysaccharidosis type IH (MPS-IH, Hurler syndrome). Cultures of MSCs were initially exposed to a green fluorescent protein-expressing retrovirus. Green fluorescent protein-positive cells maintained their proliferative and differentiation capacity. Next we used a vector encoding alpha-L-iduronidase (IDUA), the enzyme that is defective in MPS-IH. Following transduction, MPS-IH MSCs expressed high levels of IDUA and secreted supernormal levels of this enzyme into the extracellular medium. Exogenous IDUA expression led to a normalization of glycosaminoglycan storage in MPS-IH cells, as evidenced by a dramatic decrease in the amount of (35)SO(4) sequestered within the heparan sulfate and dermatan sulfate compartments of these cells. Finally, gene-modified MSCs were able to cross-correct the enzyme defect in untransduced MPS-IH fibroblasts via protein transfer.

Who's Expressing in "Expressive Writing"?

ERIC Educational Resources Information Center

Reed, Janine

In an attempt to understand what expressive writing means to themselves and to their students, teachers should explore and reflect on various questions regarding expressive writing theories and practices. For many, self-expression is the basis of all serious writing and an important stage in any act of learning, so it is essential to uncover the…

Evaluation of the immunomodulatory and antiviral effects of the cytokine combination IFN-α and IL-7 in the lymphocytic choriomeningitis virus and Friend retrovirus mouse infection models.

PubMed

Audigé, Annette; Hofer, Ursula; Dittmer, Ulf; van den Broek, Maries; Speck, Roberto F

2011-10-01

Existing therapies for chronic viral infections are still suboptimal or have considerable side effects, so new therapeutic strategies need to be developed. One option is to boost the host's immune response with cytokines. We have recently shown in an acute ex vivo HIV infection model that co-administration of interferon (IFN)-α and interleukin (IL)-7 allows us to combine the potent anti-HIV activity of IFN-α with the beneficial effects of IL-7 on T-cell survival and function. Here we evaluated the effect of combining IFN-α and IL-7 on viral replication in vivo in the chronic lymphocytic choriomeningitis virus (LCMV) and acute Friend retrovirus (FV) infection models. In the chronic LCMV model, cytokine treatment was started during the early replication phase (i.e., on day 7 post-infection [pi]). Under the experimental conditions used, exogenous IFN-α inhibited FV replication, but had no effect on viral replication in the LCMV model. There was no therapeutic benefit of IL-7 either alone or in combination with IFN-α in either of the two infection models. In the LCMV model, dose-dependent effects of the cytokine combination on T-cell phenotype/function were observed. It is possible that these effects would translate into antiviral activity in re-challenged mice. It is also possible that another type of IFN-α/β or induction of endogenous IFN-α/β alone or in combination with IL-7 would have antiviral activity in the LCMV model. Furthermore, we cannot exclude that some effect on viral titers would have been seen at later time points not investigated here (i.e., beyond day 34 pi). Finally, IFN-α/IL-7 may inhibit the replication of other viruses. Thus it might be worth testing these cytokines in other in vivo models of chronic viral infections.