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Sample records for retrovirus perv expression

  1. Expression of porcine endogenous retroviruses (PERVs) in melanomas of Munich miniature swine (MMS) Troll.

    PubMed

    Dieckhoff, Britta; Puhlmann, Jenny; Büscher, Kristina; Hafner-Marx, Angela; Herbach, Nadja; Bannert, Norbert; Büttner, Mathias; Wanke, Rüdiger; Kurth, Reinhard; Denner, Joachim

    2007-07-20

    Porcine endogenous retroviruses (PERVs) are integrated in the genome of all pig breeds. Since some of them are able to infect human cells, they might represent a risk for xenotransplantation using pig cells or organs. However, the expression and biological role of PERVs in healthy pigs as well as in porcine tumours is largely unknown. Since we and others have recently shown overexpression of a human endogenous retrovirus, HERV-K, in human melanomas, we studied the expression of PERVs in melanomas of selectively bred Munich miniature swine (MMS) Troll. This breeding herd of MMS Troll is characterised by a high prevalence of melanomas, which histologically resemble various types of cutaneous melanomas in humans. Several genetic factors have been defined when studying inheritance of melanomas and melanocytic nevi in MMS Troll. Here we show that the polytropic PERV-A and PERV-B as well as the ecotropic PERV-C are present in the genome of all melanoma bearing MMS Troll investigated. Most interestingly, in the spleen, but not in other organs, recombinant PERV-A/C proviruses were found. PERV expression was found elevated in melanomas when compared to normal skin and viral proteins were expressed in melanomas and pulmonary metastasis-derived melanoma cell cultures. During passaging of these cells in vitro the expression of PERV mRNA and protein increased and virus particles were released as shown by RT activity in the supernatant and by electron microscopy. Genomic RNA of PERV-A, -B and -C were found in pelleted virus particles. Although PERV expression was elevated in melanomas and pulmonary metastasis-derived cell cultures, the function of the virus in tumour development is still unclear.

  2. How Active Are Porcine Endogenous Retroviruses (PERVs)?

    PubMed Central

    Denner, Joachim

    2016-01-01

    Porcine endogenous retroviruses (PERVs) represent a risk factor if porcine cells, tissues, or organs were to be transplanted into human recipients to alleviate the shortage of human transplants; a procedure called xenotransplantation. In contrast to human endogenous retroviruses (HERVs), which are mostly defective and not replication-competent, PERVs are released from normal pig cells and are infectious. PERV-A and PERV-B are polytropic viruses infecting cells of several species, among them humans; whereas PERV-C is an ecotropic virus infecting only pig cells. Virus infection was shown in co-culture experiments, but also in vivo, in the pig, leading to de novo integration of proviruses in certain organs. This was shown by measurement of the copy number per cell, finding different numbers in different organs. In addition, recombinations between PERV-A and PERV-C were observed and the recombinant PERV-A/C were found to be integrated in cells of different organs, but not in the germ line of the animals. Here, the evidence for such in vivo activities of PERVs, including expression as mRNA, protein and virus particles, de novo infection and recombination, will be summarised. These activities make screening of pigs for provirus number and PERV expression level difficult, especially when only blood or ear biopsies are available for analysis. Highly sensitive methods to measure the copy number and the expression level will be required when selecting pigs with low copy number and low expression of PERV as well as when inactivating PERVs using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated nuclease (CRISPR/Cas) technology. PMID:27527207

  3. Generation of neutralising antibodies against porcine endogenous retroviruses (PERVs)

    SciTech Connect

    Kaulitz, Danny; Fiebig, Uwe; Eschricht, Magdalena; Wurzbacher, Christian; Kurth, Reinhard; Denner, Joachim

    2011-03-01

    Antibodies neutralising porcine endogenous retroviruses (PERVs) were induced in different animal species by immunisation with the transmembrane envelope protein p15E. These antibodies recognised epitopes, designated E1, in the fusion peptide proximal region (FPPR) of p15E, and E2 in the membrane proximal external region (MPER). E2 is localised in a position similar to that of an epitope in the transmembrane envelope protein gp41 of the human immunodeficiency virus-1 (HIV-1), recognised by the monoclonal antibody 4E10 that is broadly neutralising. To detect neutralising antibodies specific for PERV, a novel assay was developed, which is based on quantification of provirus integration by real-time PCR. In addition, for the first time, highly effective neutralising antibodies were obtained by immunisation with the surface envelope protein of PERV. These data indicate that neutralising antibodies can be induced by immunisation with both envelope proteins.

  4. Functional hierarchy of two L domains in Porcine Endogenous Retrovirus (PERV) that influence release and infectivity

    PubMed Central

    Marcucci, Katherine T.; Martina, Yuri; Harrison, Frank; Wilson, Carolyn A.

    2008-01-01

    The Porcine Endogenous Retrovirus (PERV) Gag protein contains two late (L) domain motifs, PPPY and P(F/S)AP. Using viral release assays we demonstrate that PPPY is the dominant L domain involved in PERV release. PFAP represents a novel retroviral L domain variant and is defined by abnormal viral assembly phenotypes visualized by electron microscopy and attenuation of early PERV release as measured by viral genomes. PSAP is functionally dominant over PFAP in early PERV release. PSAP virions are 3.5-fold more infectious in vitro by TCID50 and in vivo results in more RNA positive tissues and higher levels of proviral DNA using our human PERV-A receptor (HuPAR-2) transgenic mouse model (Martina et al., 2006. Journal of Virology. 80: 3135-46). The functional hierarchies displayed by PERV L domains, demonstrates that L domain selection in viral evolution exists to promote efficient viral assembly, release and infectivity in the virus-host context. PMID:18355887

  5. Identification of receptors for pig endogenous retrovirus.

    PubMed

    Ericsson, Thomas A; Takeuchi, Yasuhiro; Templin, Christian; Quinn, Gary; Farhadian, Shelli F; Wood, James C; Oldmixon, Beth A; Suling, Kristen M; Ishii, Jennifer K; Kitagawa, Yoshinori; Miyazawa, Takayuki; Salomon, Daniel R; Weiss, Robin A; Patience, Clive

    2003-05-27

    Xenotransplantation of porcine tissues has the potential to treat a wide variety of major health problems including organ failure and diabetes. Balanced against the potential benefits of xenotransplantation, however, is the risk of human infection with a porcine microorganism. In particular, the transmission of porcine endogenous retrovirus (PERV) is a major concern [Chapman, L. E. & Bloom, E. T. (2001) J. Am. Med. Assoc. 285, 2304-2306]. Here we report the identification of two, sequence-related, human proteins that act as receptors for PERV-A, encoded by genes located on chromosomes 8 and 17. We also describe homologs from baboon and porcine cells that also are active as receptors. Conversely, activity could not be demonstrated with a syntenic murine receptor homolog. Sequence analysis indicates that PERV-A receptors [human PERV-A receptor (HuPAR)-1, HuPAR-2, baboon PERV-A receptor 2, and porcine PERV-A receptor] are multiple membrane-spanning proteins similar to receptors for other gammaretroviruses. Expression is widespread in human tissues including peripheral blood mononuclear cells, but their biological functions are unknown. The identification of the PERV-A receptors opens avenues of research necessary for a more complete assessment of the retroviral risks of pig to human xenotransplantation.

  6. Susceptibility of porcine endogenous retrovirus to anti-retroviral inhibitors

    PubMed Central

    Argaw, Takele; Colon-Moran, Winston; Wilson, Carolyn

    2016-01-01

    Background Porcine endogenous retrovirus (PERV) is an endogenous retrovirus that poses a risk of iatrogenic transmission in the context of pig-to-human xenotransplantation. The lack of a means to control PERV infection in the context of pig-to-human xenotransplantation is a major concern in the field. In this study, we set out to evaluate the ability of currently licensed anti-HIV drugs, and other types of anti-retroviral compounds, to inhibit PERV infection in vitro. Methods We used target cells stably expressing one of the known PERV viral receptors, an infectious molecular clone, PERV-A 14/220, and at least one drug from each class of anti-retroviral inhibitors as well as off-label drugs shown to have anti-viral activities. The susceptibility of PERV-A 14/220 LacZ to the anti-retroviral drugs was determined from infected cells by histochemical staining. Results We extend the results of previous studies by showing that, in addition to raltegravir, dolutegravir is found to have a potent inhibitory activity against PERV replication (IC50 8.634 ±0.336 and IC50 3.06 ± 0.844 nm, respectively). The anti-HIV drug zidovudine (AZT) showed considerable anti-PERV activity with IC50 of 1.923 ±0.691 μm as well. Conclusions The study results indicate that some of the licensed antiretroviral drugs may be useful for controlling PERV infection. However, the efficacy at nanomolar concentrations put forward integrase inhibitors as a drug that has the potential to be useful in the event that xenotransplantation recipients have evidence of PERV transmission and replication. PMID:27028725

  7. 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…

  8. Transient transmission of porcine endogenous retrovirus to fetal lambs after pig islet tissue xenotransplantation.

    PubMed

    Popp, Sarah K; Mann, David A; Milburn, Peter J; Gibbs, Adrian J; McCullagh, Peter J; Wilson, James Dennis; Tönjes, Ralf R; Simeonovic, Charmaine J

    2007-01-01

    Evidence for the in vivo transmission of porcine endogenous retrovirus (PERV) from porcine xenografts to various recipient animals has been inconsistent. To characterize the contribution of the host immune system to the potential for PERV transmission from pig islet tissue xenografts to host tissues, we examined two immunoincompetent animal models, thymectomizsed fetal lambs and NODscid mice. Pig proislets were grafted into fetal lambs or adult NODscid mice. Conventional, nested and real-time PCR/RT-PCR tests were used to search for PERV and pig cell-specific sequences (porcine mitochondrial cytochrome oxidase II (COII) or mitochondrial ribosomal 12S) in pig proislets, host liver and spleen at 5-84 days (lambs) or 96 days (mice) after transplantation. Xenografts were harvested at the same time points. The copy number of PERV sequences and host cell-specific nuclear (palmitoylcarnitine transferase) sequences was assessed by real-time PCR to estimate the proportion of PERV-infected host cells. Pig proislets were shown to be PERV+ve by PCR and immunohistochemistry (PERV B env protein p15E). PERV transmission (PERV A, B or C DNA in the absence of porcine COII or 12S sequences) was detected by nested PCR and real-time PCR in 4/12 fetal lamb liver samples 5-23 days after transplantation; the maximum copy number of PERV B env sequences was found at day 5 (700 copies/1 x 10(6) lamb cells). A total of 4/12 fetal lambs demonstrated both PERV and 12S porcine sequences in liver samples (days 5-84) by real-time PCR, suggesting that pig cells had migrated to those tissues and established microchimerism; nested PCR showed evidence for microchimerism (porcine COII sequences alone) in 2/12 lambs (day 5). The incidence of PERV transmission and frequency of microchimerism was similar in host spleen analysed by real-time PCR. Histological examination showed complete xenograft rejection by 23 days after transplantation to fetal lambs. In contrast, pig proislet xenografts survived long

  9. Reappraisal of biosafety risks posed by PERVs in xenotransplantation.

    PubMed

    Louz, Derrick; Bergmans, Hans E; Loos, Birgit P; Hoeben, Rob C

    2008-01-01

    Donor materials of porcine origin could potentially provide an alternative source of cells, tissues or whole organs for transplantation to humans, but is hampered by the health risk posed by infection with porcine viruses. Although pigs can be bred in such a way that all known exogenous microorganisms are eliminated, this is not feasible for all endogenous pathogens, such as the porcine endogenous retroviruses (PERVs) which are present in the germline of pigs as proviruses. Upon transplantation, PERV proviruses would be transferred to the human recipient along with the xenograft. If xenotransplantation stimulates or facilitates replication of PERVs in the new hosts, a risk exists for adaptation of the virus to humans and subsequent spread of these viruses. In a worst-case scenario, this might result in the emergence of a new viral disease. Although the concerns for disease potential of PERVs are easing, only limited pre-clinical and clinical data are available. Small-scale, well-designed and carefully controlled clinical trials would provide more evidence on the safety of this approach and allow a better appreciation of the risks involved. It is therefore important to have a framework of protective measures and monitoring protocols in place to facilitate such initially small scale clinical trials. This framework will raise ethical and social considerations regarding acceptability.

  10. [Inhibiting GDF-8 expression by retrovirus-based RNAi stably].

    PubMed

    Liu, Chaowu; Yang, Zhuo; Zhao, Bin; Liu, Changmei

    2008-02-01

    We cloned human U6 promoter from pAVU6 + 27 vector into pXSN to transcripe small RNA. Meanwhile, a shRNA targeting GDF-8 was cloned down-stream of the hU6 promoter to construct recombinant vector. Then the packing cell GP-293 was co-transfected the recombinant with pVSV-G to gernarate virus particle. Resistant C2C12 cell pools were screened using G418. Levels of mRNA and protein of GDF-8 were tested by Real-Time PCR and western blotting. Cell proliferation and cell cycle were analyzed using MTT and FACS. The expression of GDF-8 was dramatically decreased by the retrovirus-based system in C2C12 cells. Cells proliferated effectively after integrating the recombinant. The cells in G0/G1 phase decreased by 13.7%, while cells in S phase increased by 14.9%. In conclusion, the retrovirus-based RNAi could be used to stably silence GDF-8. It can be a powerful tool in curing muscle atrophy.

  11. Transmission of Porcine Endogenous Retrovirus Produced from Different Recipient Cells In Vivo

    PubMed Central

    Kim, Sehyun; Gwon, Yong-Dae; Cho, Yeondong; Yang, Jae Myung; Oh, Yu-Kyoung; Kim, Young bong

    2016-01-01

    Humanized pigs have been developed to reduce the incidence of immune rejection in xenotransplantation, but significant concerns remain, such as transmission of viral zoonosis. Porcine endogenous retroviruses (PERV), which exist in the genome of pigs, are produced as infectious virions from all porcine cells and cause zoonosis. Here, we examined the possibility of zoonosis of hosts under conditions of immune suppression or xenotransplantation of cells producing host-adapted viruses. Upon transplantation of PERV-producing porcine cells into mice, no transmission of PERV was detected, whereas, transmission of PERV from mice transplanted with mouse-adapted PERV-producing cells was detected. In addition, the frequency of PERV transmission was increased in CsA treated mice transplanted with PERV-producing murine cells, compared with PERV-producing porcine cells. Transmission of PERV to host animals did not affect weight but immune responses, in particular, the number of T cells from PERV-transmitted mice, were notably reduced. The observed risk of PERV zoonosis highlights the requirement for thorough evaluation of viral zoonosis under particular host conditions, such as immunosuppressive treatment and transplantation with host-adapted virus-producing cells. PMID:27832080

  12. Xenotransplantation and pig endogenous retroviruses.

    PubMed

    Magre, Saema; Takeuchi, Yasuhiro; Bartosch, Birke

    2003-01-01

    Xenotransplantation, in particular transplantation of pig cells, tissues and organs into human patients, may alleviate the current shortage of suitable allografts available for human transplantation. This overview addresses the physiological, immunological and virological factors considered with regard to xenotransplantation. Among the issues reviewed are the merits of using pigs as xenograft source species, the compatibility of pig and human organ physiology and the immunological hindrances with regard to the various types of rejection and attempts at abrogating rejection. Advances in the prevention of pig organ rejection by creating genetically modified pigs that are more suited to the human microenvironment are also discussed. Finally, with regard to virology, possible zoonotic infections emanating from pigs are reviewed, with special emphasis on the pig endogenous retrovirus (PERV). An in depth account of PERV studies, comprising their discovery as well as recent knowledge of the virus, is given. To date, all retrospective studies on patients with pig xenografts have shown no evidence of PERV transmission, however, many factors make us interpret these results with caution. Although the lack of PERV infection in xenograft recipients up to now is encouraging, more basic research and controlled animal studies that mimic the pig to human xenotransplantation setting more closely are required for safety assessment.

  13. Restriction of Porcine Endogenous Retrovirus by Porcine APOBEC3 Cytidine Deaminases ▿

    PubMed Central

    Dörrschuck, Eva; Fischer, Nicole; Bravo, Ignacio G.; Hanschmann, Kay-Martin; Kuiper, Heidi; Spötter, Andreas; Möller, Ronny; Cichutek, Klaus; Münk, Carsten; Tönjes, Ralf R.

    2011-01-01

    Xenotransplantation of porcine cells, tissues, and organs shows promise to surmount the shortage of human donor materials. Among the barriers to pig-to-human xenotransplantation are porcine endogenous retroviruses (PERV) since functional representatives of the two polytropic classes, PERV-A and PERV-B, are able to infect human embryonic kidney cells in vitro, suggesting that a xenozoonosis in vivo could occur. To assess the capacity of human and porcine cells to counteract PERV infections, we analyzed human and porcine APOBEC3 (A3) proteins. This multigene family of cytidine deaminases contributes to the cellular intrinsic immunity and act as potent inhibitors of retroviruses and retrotransposons. Our data show that the porcine A3 gene locus on chromosome 5 consists of the two single-domain genes A3Z2 and A3Z3. The evolutionary relationships of the A3Z3 genes reflect the evolutionary history of mammals. The two A3 genes encode at least four different mRNAs: A3Z2, A3Z3, A3Z2-Z3, and A3Z2-Z3 splice variant A (SVA). Porcine and human A3s have been tested toward their antiretroviral activity against PERV and murine leukemia virus (MuLV) using novel single-round reporter viruses. The porcine A3Z2, A3Z3 and A3Z2-Z3 were packaged into PERV particles and inhibited PERV replication in a dose-dependent manner. The antiretroviral effect correlated with editing by the porcine A3s with a trinucleotide preference for 5′ TGC for A3Z2 and A3Z2-Z3 and 5′ CAC for A3Z3. These results strongly imply that human and porcine A3s could inhibit PERV replication in vivo, thereby reducing the risk of infection of human cells by PERV in the context of pig-to-human xenotransplantation. PMID:21307203

  14. Repression of retrovirus-mediated transgene expression by interferons: implications for gene therapy.

    PubMed Central

    Ghazizadeh, S; Carroll, J M; Taichman, L B

    1997-01-01

    Retrovirus-mediated gene transfer is commonly used in gene therapy protocols and has the potential to provide long-term expression of the transgene. Although expression of a retrovirus-delivered transgene is satisfactory in cultured cells, it has been difficult to achieve consistent and high-level expression in vivo. In this investigation, we explored the possibility of modulating transgene expression by host-derived cytokines. Normal human keratinocytes and dermal fibroblasts were transduced with recombinant retroviruses expressing a reporter gene (lacZ). Treatment of transduced cells with a proinflammatory cytokine, gamma interferon (IFN-gamma), significantly reduced lacZ expression to less than 25% of that of nontreated cells. The inhibition was concentration dependent (peak at 5 ng/ml) and time dependent (maximal at 16 h for transcript and 24 h for protein); expression remained repressed in the continued presence of IFN-gamma but returned to normal levels 24 h after IFN-gamma withdrawal. The decrease in beta-galactosidase activity appeared to result from decrease in steady-state lacZ mRNA levels. Inhibitors of transcription and translation blocked IFN-gamma-induced repression, suggesting involvement of newly synthesized protein intermediates. Similar results were obtained by treatment of transduced cells with IFN-alpha but not with other proinflammatory cytokines, including tumor necrosis factor alpha, interleukin-2 (IL-1), IL-4, and granulocyte colony-stimulating factor. Although the level of lacZ mRNA was reduced by >70% following IFN treatment, the rate of lacZ transcription was not significantly different from that for nontreated cells. These results suggest that IFN-mediated regulation of transgene expression is at a posttranscriptional level. Interestingly, IFN-gamma also suppressed transgene expression driven by a cellular promoter (involucrin) inserted in an internal position in the retroviral vector. The presence of the overlapping 3' untranslated

  15. Expression of human endogenous retrovirus K and W in babies.

    PubMed

    Nali, L H S; Oliveira, A C S; Alves, D O; Caleiro, G S; Nunes, C F; Gerhardt, D; Succi, R C M; Romano, Camila M; Machado, D M

    2017-03-01

    Here we determined the relative expression of HERV-K and W proviruses in HIV infected and non-infected mothers as well as their respective babies up to 1 year-old. HIV-infected mothers, their babies and uninfected control groups presented expression of both HERV-K and HERV-W with relatively high frequency. While the level of HERV-K expression was similar among groups, the level of HERV-W expression in HIV-infected mothers was four-fold higher than the uninfected mothers from the control group (p < 0.01). HERV-W was down regulated in HIV-exposed babies in comparison to non-exposed babies. To our knowledge, this is the first report of HERV transcriptional activity in babies from 0-1 year-old.

  16. Protection against Friend retrovirus-induced leukemia by recombinant vaccinia viruses expressing the gag gene.

    PubMed Central

    Miyazawa, M; Nishio, J; Chesebro, B

    1992-01-01

    High sequence variability in the envelope gene of human immunodeficiency virus has provoked interest in nonenvelope antigens as potential immunogens against retrovirus infection. However, the role of core protein antigens encoded by the gag gene in protective immunity against retroviruses is unclear. By using recombinant vaccinia viruses expressing the Friend murine leukemia helper virus (F-MuLV) gag gene, we could prime CD4+ T-helper cells and protectively immunize susceptible strains of mice against Friend retrovirus infection. Recovery from leukemic splenomegaly developed more slowly after immunization with vaccinia virus-F-MuLV gag than with vaccinia virus-F-MuLV env; however, genetic nonresponders to the envelope protein could be partially protected with Gag vaccines. Class switching of F-MuLV-neutralizing antibodies from immunoglobulin M to immunoglobulin G after challenge with Friend virus complex was facilitated in mice immunized with the Gag antigen. Sequential deletion of the gag gene revealed that the major protective epitope was located on the N-terminal hydrophobic protein p15. Images PMID:1534853

  17. Epigenetic regulation on the 5'-proximal CpG island of human porcine endogenous retrovirus subgroup A receptor 2/GPR172B.

    PubMed

    Nakaya, Yuki; Shojima, Takayuki; Yasuda, Jiro; Imakawa, Kazuhiko; Miyazawa, Takayuki

    2011-01-01

    Porcine endogenous retroviruses (PERVs) have been considered one of the major risks of xenotransplantation from pigs to humans. PERV-A efficiently utilizes human PERV-A receptor 2 (HuPAR-2)/GPR172B to infect human cells; however, there has been no study on the regulation mechanisms of HuPAR-2/GPR172B expression. In this study, we examined the expression of HuPAR-2/GPR172B from the standpoint of epigenetic regulation and discussed the risks of PERV-A infection in xenotransplantation. Quantitative real-time RT-PCR revealed that HuPAR-2 mRNA was preferentially expressed in placental tissue, whereas it was highly suppressed in BeWo cells (a human choriocarcinoma cell line) and HEK293 cells. A CpG island containing the HuPAR-2 transcription starting site was identified by in silico analysis. The DNA methylation ratio (the relative quantity of methylcytosine to total cytosine) and histone modification (H3K9me3) levels in the CpG island measured by bisulfite genomic sequencing and ChIP assay, respectively, were inversely correlated with the mRNA levels. Both HuPAR-2 mRNA and HuPAR-2 protein were up-regulated in HEK293 cells by inhibiting DNA methylation and histone deacetylation. Additionally, promoter/enhancer activities within the CpG island were suppressed by in vitro DNA methylation. Our results demonstrated that epigenetic modification regulates HuPAR-2 expression.

  18. Expression of human endogenous retrovirus type K envelope glycoprotein in insect and mammalian cells.

    PubMed Central

    Tönjes, R R; Limbach, C; Löwer, R; Kurth, R

    1997-01-01

    The human endogenous retrovirus type K (HERV-K) family codes for the human teratocarcinoma-derived retrovirus (HTDV) particles. The existence of the envelope protein (ENV) of HERV-K encoded by the subgenomic env mRNA has not yet been demonstrated. To study the genetic requirements for successful expression of ENV, we have constructed a series of recombinant HERV-K env expression vectors for infection and transfection experiments in insect cells and mammalian cells, respectively. Six baculovirus constructs bearing full-length or truncated HERV-K env with or without homologous or heterologous signal peptides were used for infections of insect cells. All recombinant baculoviruses yielded ENV proteins with the expected molecular masses. The full-length 80- to 90-kDa HERV-K ENV protein including the cORF leader sequence was glycosylated in insect cells. In addition, the 14-kDa cORF protein was expressed due to splicing of the full-length env mRNA. The ENV precursor protein is not cleaved to the surface (SU) and transmembrane (TM) glycoproteins; it does not appear on the surface of infected insect cells and is not secreted into the medium. For ENV expression in COS cells, plasmid vectors harboring the cytomegalovirus immediate-early promoter/intron A element and the tissue plasminogen activator (t-PA) signal peptide or the homologous HERV-K signal peptide upstream of the env gene were employed. Glycosylated and uncleaved ENV was expressed as in GH teratocarcinoma cells but at higher levels. The heterologous t-PA signal sequence was instrumental for expression of HERV-K ENV on the cell surface. Hence, we have shown for the first time that the HERV-K env gene has the potential to be expressed as a full-length envelope protein with appropriate glycosylation. In addition, our data provide explanations for the lack of infectivity of HERV-K/HTDV particles. PMID:9060628

  19. Regulated expression of a complete human beta-globin gene encoded by a transmissible retrovirus vector.

    PubMed Central

    Cone, R D; Weber-Benarous, A; Baorto, D; Mulligan, R C

    1987-01-01

    We introduced a human beta-globin gene into murine erythroleukemia (MEL) cells by infection with recombinant retroviruses containing the complete genomic globin sequence. The beta-globin gene was correctly regulated during differentiation, steady-state mRNA levels being induced 5- to 30-fold after treatment of the cells with the chemical inducer dimethyl sulfoxide. Studies using vectors which yield integrated proviruses lacking transcriptional enhancer sequences indicated that neither retroviral transcription nor the retroviral enhancer sequences themselves had any obvious effect on expression of the globin gene. Viral RNA expression also appeared inducible, being considerably depressed in uninduced MEL cells but approaching normal wild-type levels after dimethyl sulfoxide treatment. We provide data which suggest that the control point for both repression and subsequent activation of virus expression in MEL cells lies in the viral enhancer element. Images PMID:3029570

  20. Expression of human adenosine deaminase in mice transplanted with hemopoietic stem cells infected with amphotropic retroviruses

    PubMed Central

    1990-01-01

    Amphotropic recombinant retroviruses were generated carrying sequences encoding human adenosine deaminase (ADA). Transcription of the human ADA gene was under control of a hybrid long terminal repeat in which the enhancer from the Moloney murine leukemia virus was replaced by an enhancer from the F101 host-range mutant of polyoma virus. Hemopoietic stem cells in murine bone marrow were infected with this virus under defined culture conditions. As a result, 59% of day-12 colony forming unit spleen (CFU-S) stem cells became infected without any in vitro selection. Infected CFU-S were shown to express human ADA before transplantation and this expression sustained upon in vivo maturation. Mice transplanted with infected bone marrow exhibited human ADA expression in lymphoid, myeloid, and erythroid cell types. Moreover, human ADA expression persisted in secondary and tertiary transplanted recipients showing that human ADA-expressing cells were derived from pluripotent stem cells. These characteristics of our amphotropic viruses make them promising tools in gene therapy protocols for the treatment of severe combined immunodeficiency caused by ADA deficiency. In this respect it is also relevant that the viral vector that served as backbone for the ADA vector was previously shown to be nonleukemogenic. PMID:1974914

  1. Radiation-induced human endogenous retrovirus (HERV)-R env gene expression by epigenetic control.

    PubMed

    Lee, Ja-Rang; Ahn, Kung; Kim, Yun-Ji; Jung, Yi-Deun; Kim, Heui-Soo

    2012-11-01

    It is commonly accepted that ionizing radiation induces genomic instability by changes in genomic structure, epigenetic regulation and gene expression. Human endogenous retroviruses (HERV)-R also are often differentially expressed between normal and disease tissues under unstable genomic conditions and are implicated in the pathogenesis of several human diseases. To understand the influence of ionizing radiation on HERV-R expression, we performed quantitative reverse transcription-polymerase chain reaction (RT-PCR) analyses using γ-irradiated normal human cells. Compared to nonirradiated cells, HERV-R expression was up-regulated in γ-irradiated cells. The regulatory mechanism of HERV-R expression in irradiated cells was investigated by methylation analyses of HERV-R 5'LTRs and treatment with garcinol. These data indicated that the up-regulated transcription of HERV-R may be regulated by radiation-induced epigenetic changes induced by histone modification, and thus could be of great importance for understanding the relationship between radiation-induced biological effects and transposable elements.

  2. Long-term IgG response to porcine Neu5Gc-antigens without transmission of PERV in burn patients treated with porcine skin xenografts

    PubMed Central

    Scobie, Linda; Padler-Karavani, Vered; Le Bas-Bernardet, Stephanie; Crossan, Claire; Blaha, Josef; Matouskova, Magda; Hector, Ralph D; Cozzi, Emanuele; Vanhove, Bernard; Charreau, Beatrice; Blancho, Gilles; Bourdais, Ludovic; Tallacchini, Mariachiara; Ribes, Juan M; Yu, Hai; Chen, Xi; Kracikova, Jitka; Broz, Ludomir; Hejnar, Jiri; Vesely, Pavel; Takeuchi, Yasuhiro; Varki, Ajit; Soulillou, Jean-Paul

    2013-01-01

    Acellular materials of xenogenic origin are used worldwide as xenografts and Phase I trials of viable pig pancreatic islets are currently being performed. However, limited information is available on transmission of porcine endogenous retrovirus (PERV) after xenotransplantation and on the long-term immune response of recipients to xenoantigens. We analyzed the blood of burn patients who had received living pig skin dressings for up to 8 weeks for the presence of PERV as well as for the level and nature of their long term (maximum 34 years) immune response against pig antigens. Whilst no evidence of PERV genomic material or anti PERV antibody response was found, we observed a moderate increase in anti αGal antibodies and a high and sustained anti non-αGal IgG response in those patients. Antibodies against the non-human sialic acid Neu5Gc constituted the anti non-αGal response with the recognition pattern on a sialogly can array differing from that of burn patients treated without pig skin. These data suggest that anti-Neu5Gc antibodies may represent a barrier for long-term acceptance of porcine xenografts. As anti-Neu5Gc antibodies can promote chronic inflammation, the long-term safety of living and acellular pig tissue implants in recipients warrants further evaluation. PMID:23945141

  3. Long-term IgG response to porcine Neu5Gc antigens without transmission of PERV in burn patients treated with porcine skin xenografts.

    PubMed

    Scobie, Linda; Padler-Karavani, Vered; Le Bas-Bernardet, Stephanie; Crossan, Claire; Blaha, Josef; Matouskova, Magda; Hector, Ralph D; Cozzi, Emanuele; Vanhove, Bernard; Charreau, Beatrice; Blancho, Gilles; Bourdais, Ludovic; Tallacchini, Mariachiara; Ribes, Juan M; Yu, Hai; Chen, Xi; Kracikova, Jitka; Broz, Ludomir; Hejnar, Jiri; Vesely, Pavel; Takeuchi, Yasuhiro; Varki, Ajit; Soulillou, Jean-Paul

    2013-09-15

    Acellular materials of xenogenic origin are used worldwide as xenografts, and phase I trials of viable pig pancreatic islets are currently being performed. However, limited information is available on transmission of porcine endogenous retrovirus (PERV) after xenotransplantation and on the long-term immune response of recipients to xenoantigens. We analyzed the blood of burn patients who had received living pig-skin dressings for up to 8 wk for the presence of PERV as well as for the level and nature of their long term (maximum, 34 y) immune response against pig Ags. Although no evidence of PERV genomic material or anti-PERV Ab response was found, we observed a moderate increase in anti-αGal Abs and a high and sustained anti-non-αGal IgG response in those patients. Abs against the nonhuman sialic acid Neu5Gc constituted the anti-non-αGal response with the recognition pattern on a sialoglycan array differing from that of burn patients treated without pig skin. These data suggest that anti-Neu5Gc Abs represent a barrier for long-term acceptance of porcine xenografts. Because anti-Neu5Gc Abs can promote chronic inflammation, the long-term safety of living and acellular pig tissue implants in recipients warrants further evaluation.

  4. Copy Number Variation and Differential Expression of a Protective Endogenous Retrovirus in Sheep

    PubMed Central

    Viginier, Barbara; Dolmazon, Christine; Lantier, Isabelle; Lantier, Frédéric; Archer, Fabienne; Leroux, Caroline; Terzian, Christophe

    2012-01-01

    The Jaagsiekte sheep retrovirus exJSRV and its endogenous counterpart enJSRV co-exist in sheep. exJSRV, a betaretrovirus, is the etiological agent of ovine pulmonary adenocarcinoma, and it has been demonstrated in vitro that an enJSRV Gag variant bearing the R-to-W amino acid change at position 21 was able to block exJSRV budding from the cells, providing a potential protective role for the host. In this work, we developed a fast mutation detection assay based on the oligo ligation assay (OLA) that permits the quantification of the relative proportions of the R21 and W21 Gag variants present in individual genomes and in cDNA obtained from normal and exJSRV-induced lung tumors. We have shown that the W21/R21 ratio is variable within and between breeds. We also describe for the first time that putative protecting enJSRV variants were expressed in alveolar type II cells (AECII), the major target of exJSRV. PMID:22911867

  5. Koala retroviruses: characterization and impact on the life of koalas.

    PubMed

    Denner, Joachim; Young, Paul R

    2013-10-23

    Koala retroviruses (KoRV) have been isolated from wild and captive koalas in Australia as well as from koala populations held in zoos in other countries. They are members of the genus Gammaretrovirus, are most closely related to gibbon ape leukemia virus (GaLV), feline leukemia virus (FeLV) and porcine endogenous retrovirus (PERV) and are likely the result of a relatively recent trans-species transmission from rodents or bats. The first KoRV to be isolated, KoRV-A, is widely distributed in the koala population in both integrated endogenous and infectious exogenous forms with evidence from museum specimens older than 150 years, indicating a relatively long engagement with the koala population. More recently, additional subtypes of KoRV that are not endogenized have been identified based on sequence differences and host cell receptor specificity (KoRV-B and KoRV-J). A specific association with fatal lymphoma and leukemia has been recently suggested for KoRV-B. In addition, it has been proposed that the high viral loads found in many animals may lead to immunomodulation resulting in a higher incidence of diseases such as chlamydiosis. Although the molecular basis of this immunomodulation is still unclear, purified KoRV particles and a peptide corresponding to a highly conserved domain in the envelope protein have been shown to modulate cytokine expression in vitro, similar to that induced by other gammaretroviruses. While much is still to be learned, KoRV induced lymphoma/leukemia and opportunistic disease arising as a consequence of immunomodulation are likely to play an important role in the stability of koala populations both in the wild and in captivity.

  6. Porcine Endogenous Retrovirus Infects but Does Not Replicate in Nonhuman Primate Primary Cells and Cell Lines

    PubMed Central

    Ritzhaupt, Armin; van der Laan, Luc J. W.; Salomon, Daniel R.; Wilson, Carolyn A.

    2002-01-01

    Porcine endogenous retroviruses (PERV) can infect human cell lines in vitro; hence, there is a presumed risk of viral exposure to a recipient when pig cells are transplanted into humans (xenotransplantation). Nonhuman primates (NHP) are considered a potential permissive animal model to study the risk of in vivo infection of PERV after xenotransplantation. We set out to determine whether PERV can infect and replicate in NHP primary cells or established cell lines from African green monkey, rhesus macaque, and baboon. We confirm that the NHP cell lines under investigation were infected with PERV as measured by detection of viral DNA and RNA by PCR and reverse transcription (RT)-PCR, respectively, indicating that a functional receptor must be present on the cell surface. However, the load of detectable viral DNA in infected NHP cells declined over time, and the cells never had detectable reverse transcriptase activity. Utilizing quantitative real-time TaqMan PCR we found detectable levels of unintegrated DNA intermediates, but the levels were approximately 100-fold lower compared to HEK 293 cells infected with PERV. Virions released from infected NHP cells could productively infect naïve human cell lines, HEK 293 and HeLa, as shown by RT-PCR and RT assay. However, naïve NHP cells remained negative in RT-PCR and RT assay after exposure to virions from infected NHP cells. Together our data demonstrate that NHP cells are not permissive to productive replication by PERV, presumably due to inefficient cell entry and replication. In light of these observations, the appropriateness of NHP as suitable animal models to study PERV infection in vivo needs to be reevaluated. PMID:12388691

  7. 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.

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

    SciTech Connect

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

    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.

  9. Expression of human endogenous retrovirus-K is strongly associated with the basal-like breast cancer phenotype

    PubMed Central

    Johanning, Gary L.; Malouf, Gabriel G.; Zheng, Xiaofeng; Esteva, Francisco J.; Weinstein, John N.; Wang-Johanning, Feng; Su, Xiaoping

    2017-01-01

    Human endogenous retroviruses (HERVs), which make up approximately 8% of the human genome, are overexpressed in some breast cancer cells and tissues but without regard to cancer subtype. We, therefore, analyzed TCGA RNA-Seq data to evaluate differences in expression of the HERV-K family in breast cancers of the various subtypes. Four HERV-K loci on different chromosomes were analyzed in basal, Her2E, LumA, and LumB breast cancer subtypes of 512 breast cancer patients with invasive ductal carcinoma (IDC). The results for all four loci showed higher HERV-K expression in the basal subtype, suggesting similar mechanisms of regulation regardless of locus. Expression of the HERV-K envelope gene (env) was highly significantly increased in basal tumors in comparison with the also-upregulated expression of other HERV-K genes. Analysis of reverse-phase protein array data indicated that increased expression of HERV-K is associated with decreased mutation of H-Ras (wild-type). Our results show elevation of HERV-K expression exclusively in the basal subtype of IDC breast cancer (as opposed to the other subtypes) and suggest HERV-K as a possible target for cancer vaccines or immunotherapy against this highly aggressive form of breast cancer. PMID:28165048

  10. Intracisternal A-type particles express their proteinase in a separate reading frame by translational frameshifting, similar to D-type retroviruses.

    PubMed

    Fehrmann, F; Welker, R; Kräusslich, H G

    1997-09-01

    Intracisternal A-type particles (IAP) are defective endogenous retroviruses that accumulate in the endoplasmic reticulum of rodent cells. IAP genomes share extensive sequence homologies with D-type retroviruses, but were presumed to express the viral proteinase (PR) as part of the gag open reading frame (ORF) while D-type retroviruses express PR in a separate ORF. Here we show that expression of the murine IAP element MIA14 yields three major translation products, corresponding to the Gag, Gag-PR, and Gag-PR-Pol polyproteins. Sequence analysis revealed that MIA14 PR is encoded in its own reading frame, separate from gag and pol. Frameshifting occurred with an efficiency of approximately 25% between the gag and pro ORFs and 35% between pro and pol. The region containing the putative gag-pro frameshift signal consists of a heptanucleotide slippery sequence (A6C) and a stem-loop structure probably forming a pseudoknot. Deletion of this structure element almost completely abolished frameshifting. Insertion of an additional base next to the frameshift signal placed gag and pro in the same ORF and resulted in predominant formation of Gag-PR and Gag-PR-Pol polyproteins which were not processed following in vitro translation. Expression of a similar construct in tissue culture cells, on the other hand, led to efficient intracellular processing of the mutant polyproteins.

  11. Human Endogenous Retroviruses-K (HML-2) Expression Is Correlated with Prognosis and Progress of Hepatocellular Carcinoma

    PubMed Central

    Ma, Weijie; Hong, Zhenfei; Liu, Hailing; Chen, Xi; Ding, Lu; Liu, Zhisu

    2016-01-01

    Background. The association between human endogenous retroviruses-K (HERV-K) (HML-2) and human disease, including a variety of cancers, has been indicated. However, the function of HERV-K (HML-2) in the progression of hepatocellular carcinoma (HCC) still remains largely unclear. Methods. We detected the expression of HERV-K (HML-2) in 84 HCC tissues and adjacent nontumor tissues by quantitative real-time PCR (qRT-PCR) and analyzed its correlation with the clinical parameters. Result. The HEVR-K level was significantly increased in HCC compared with adjacent normal tissues (P < 0.01) which was proved to be significantly associated with cirrhosis (P < 0.05), tumor differentiation (P < 0.05), and TNM stage (P < 0.05). Moreover, the high expression of HERV-K (HML-2) had a poorer overall survival than patients with lower expression by a Kaplan-Meier survival analysis (P < 0.01). The multivariate Cox regression analysis indicated that the level of HERV-K (HML-2) was an independent prognostic factor for the overall survival rate of HCC patients. Receiver operating characteristic (ROC) curves demonstrated the diagnostic accuracy of HERV-K (HML-2) expression in HCC (AUC = 0.729, 74.7% sensitivity, and 67.8% specificity). Conclusions. Our results suggested that upregulation of HERV-K (HML-2) in HCC patients was significantly related to cancer progression and poor outcome, indicating that HERV-K (HML-2) might be a novel candidate prognostic biomarker for HCC. PMID:28070518

  12. 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)

  13. Endogenous retroviruses in mammals: an emerging picture of how ERVs modify expression of adjacent genes.

    PubMed

    Isbel, Luke; Whitelaw, Emma

    2012-09-01

    Endogenous retrovirsuses (ERVs) have long been known to influence gene expression in plants in important ways, but what of their roles in mammals? Our relatively sparse knowledge in that area was recently increased with the finding that ERVs can influence the expression of mammalian resident genes by disrupting transcriptional termination. For many mammalian biologists, retrotransposition is considered unimportant except when it disrupts the reading frame of a gene, but this view continues to be challenged. It has been known for some time that integration into an intron can create novel transcripts and integration upstream of a gene can alter the expression of the transcript, in many cases producing phenotypic consequences and disease. The new findings on transcriptional termination extend the opportunities for retrotransposons to play a role in human disease.

  14. Tolerance and immune response to the porcine endogenous retrovirus in German landrace pigs immunised with viral proteins.

    PubMed

    Denner, Joachim; Petersen, Björn; Niemann, Heiner

    2015-10-02

    Immunisation of goats, mice, rats, rabbits, guinea pigs, and hamsters with the recombinant ectodomain of the porcine endogenous retrovirus (PERV) transmembrane envelope (TM) protein (p15E) induced binding and neutralising immune antibodies in all animals. In contrast, no antibodies were induced when pigs were immunised with p15E, indicating that pigs are tolerant to their endogenous retroviruses, at least to the TM protein. To answer the question of whether pigs are tolerant to other structural proteins of PERV, we immunised German landrace pigs with p15E, this time in conjunction with the surface envelope proteins gp70 and the core capsid Gag protein p27CA. To ensure that the pigs were immunocompetent and that immunisation was successful, all animals also received an injection of an unrelated protein, keyhole limpet hemocyanin (KLH). Whereas all animals produced antibodies against KLH, no animals produced antibodies against the viral envelope proteins, thus confirming previous results for p15E and extending them to the other envelope protein, gp70. However, the pigs did produce antibodies against p27CA, indicating that there is no tolerance to the core capsid protein of PERV.

  15. Retrovirus-Mediated Expression of E2A-PBX1 Blocks Lymphoid Fate but Permits Retention of Myeloid Potential in Early Hematopoietic Progenitors

    PubMed Central

    Woodcroft, Mark W.; Nanan, Kyster; Thompson, Patrick; Tyryshkin, Kathrin; Smith, Steven P.; Slany, Robert K.; LeBrun, David P.

    2015-01-01

    The oncogenic transcription factor E2A-PBX1 is expressed consequent to chromosomal translocation 1;19 and is an important oncogenic driver in cases of pre-B-cell acute lymphoblastic leukemia (ALL). Elucidating the mechanism by which E2A-PBX1 induces lymphoid leukemia would be expedited by the availability of a tractable experimental model in which enforced expression of E2A-PBX1 in hematopoietic progenitors induces pre-B-cell ALL. However, hematopoietic reconstitution of irradiated mice with bone marrow infected with E2A-PBX1-expressing retroviruses consistently gives rise to myeloid, not lymphoid, leukemia. Here, we elucidate the hematopoietic consequences of forced E2A-PBX1 expression in primary murine hematopoietic progenitors. We show that introducing E2A-PBX1 into multipotent progenitors permits the retention of myeloid potential but imposes a dense barrier to lymphoid development prior to the common lymphoid progenitor stage, thus helping to explain the eventual development of myeloid, and not lymphoid, leukemia in transplanted mice. Our findings also indicate that E2A-PBX1 enforces the aberrant, persistent expression of some genes that would normally have been down-regulated in the subsequent course of hematopoietic maturation. We show that enforced expression of one such gene, Hoxa9, a proto-oncogene associated with myeloid leukemia, partially reproduces the phenotype produced by E2A-PBX1 itself. Existing evidence suggests that the 1;19 translocation event takes place in committed B-lymphoid progenitors. However, we find that retrovirus-enforced expression of E2A-PBX1 in committed pro-B-cells results in cell cycle arrest and apoptosis. Our findings indicate that the neoplastic phenotype induced by E2A-PBX1 is determined by the developmental stage of the cell into which the oncoprotein is introduced. PMID:26098938

  16. Expression of human. alpha. -globin and mouse/human hybrid. beta. -globin genes in murine hemopoietic stem cells transduced by recombinant retroviruses

    SciTech Connect

    Li, C.L.; Dwarki, V.J.; Verma, I.M. )

    1990-06-01

    Murine cell lines releasing helper-free recombinant retroviruses containing human {alpha}-globin and mouse/human hybrid {beta}-globin genes were generated. The expression of the hybrid {beta}-globin gene but not the human {alpha}-globin gene was regulated appropriately in infected mouse erythroid leukemia (MEL) cells. Murine bone marrow cells were infected by coculture with virus-producing cells and transplanted into lethally irradiated syngeneic recipients. Greater than 90% of the spleen colonies (12-15 days), which are derived from hemopoietic multipotential stem cells, showed proviral integration. Various levels of expression of the transduced globin genes were detected in all of the provirus-positive spleen colonies. Proviral sequences and transcripts from the transduced globin genes could also be detected in a few long-term reconstituted recipients in an observation period of 10 months after transplantation.

  17. Human endogenous retroviruses and cancer

    PubMed Central

    Gonzalez-Cao, María; Iduma, Paola; Karachaliou, Niki; Santarpia, Mariacarmela; Blanco, Julià; Rosell, Rafael

    2016-01-01

    Human endogenous retroviruses (HERVs) are retroviruses that infected human genome millions of years ago and have persisted throughout human evolution. About 8% of our genome is composed of HERVs, most of which are nonfunctional because of epigenetic control or deactivating mutations. However, a correlation between HERVs and human cancer has been described and many tumors, such as melanoma, breast cancer, germ cell tumors, renal cancer or ovarian cancer, express HERV proteins, mainly HERV-K (HML6) and HERV-K (HML2). Although the causative role of HERVs in cancer is controversial, data from animal models demonstrated that endogenous retroviruses are potentially oncogenic. HERV protein expression in human cells generates an immune response by activating innate and adaptive immunities. Some HERV-derived peptides have antigenic properties. For example, HERV-K (HML-6) encodes the HER-K MEL peptide recognized by CD8+ lymphocytes. In addition, HERVs are two-edged immunomodulators. HERVs show immunosuppressive activity. The presence of genomic retroviral elements in host-cell cytosol may activate an interferon type I response. Therefore, targeting HERVs through cellular vaccines or immunomodulatory drugs combined with checkpoint inhibitors is attracting interest because they could be active in human tumors. PMID:28154780

  18. Macroevolution of complex retroviruses.

    PubMed

    Katzourakis, Aris; Gifford, Robert J; Tristem, Michael; Gilbert, M Thomas P; Pybus, Oliver G

    2009-09-18

    Retroviruses can leave a "fossil record" in their hosts' genomes in the form of endogenous retroviruses. Foamy viruses, complex retroviruses that infect mammals, have been notably absent from this record. We have found an endogenous foamy virus within the genomes of sloths and show that foamy viruses were infecting mammals more than 100 million years ago and codiverged with their hosts across an entire geological era. Our analysis highlights the role of evolutionary constraint in maintaining viral genome structure and indicates that accessory genes and mammalian mechanisms of innate immunity are the products of macroevolutionary conflict played out over a geological time scale.

  19. Transspecies Transmission of Gammaretroviruses and the Origin of the Gibbon Ape Leukaemia Virus (GaLV) and the Koala Retrovirus (KoRV)

    PubMed Central

    Denner, Joachim

    2016-01-01

    Transspecies transmission of retroviruses is a frequent event, and the human immunodeficiency virus-1 (HIV-1) is a well-known example. The gibbon ape leukaemia virus (GaLV) and koala retrovirus (KoRV), two gammaretroviruses, are also the result of a transspecies transmission, however from a still unknown host. Related retroviruses have been found in Southeast Asian mice although the sequence similarity was limited. Viruses with a higher sequence homology were isolated from Melomys burtoni, the Australian and Indonesian grassland melomys. However, only the habitats of the koalas and the grassland melomys in Australia are overlapping, indicating that the melomys virus may not be the precursor of the GaLV. Viruses closely related to GaLV/KoRV were also detected in bats. Therefore, given the fact that the habitats of the gibbons in Thailand and the koalas in Australia are far away, and that bats are able to fly over long distances, the hypothesis that retroviruses of bats are the origin of GaLV and KoRV deserves consideration. Analysis of previous transspecies transmissions of retroviruses may help to evaluate the potential of transmission of related retroviruses in the future, e.g., that of porcine endogenous retroviruses (PERVs) during xenotransplantation using pig cells, tissues or organs. PMID:27999419

  20. Transspecies Transmission of Gammaretroviruses and the Origin of the Gibbon Ape Leukaemia Virus (GaLV) and the Koala Retrovirus (KoRV).

    PubMed

    Denner, Joachim

    2016-12-20

    Transspecies transmission of retroviruses is a frequent event, and the human immunodeficiency virus-1 (HIV-1) is a well-known example. The gibbon ape leukaemia virus (GaLV) and koala retrovirus (KoRV), two gammaretroviruses, are also the result of a transspecies transmission, however from a still unknown host. Related retroviruses have been found in Southeast Asian mice although the sequence similarity was limited. Viruses with a higher sequence homology were isolated from Melomys burtoni, the Australian and Indonesian grassland melomys. However, only the habitats of the koalas and the grassland melomys in Australia are overlapping, indicating that the melomys virus may not be the precursor of the GaLV. Viruses closely related to GaLV/KoRV were also detected in bats. Therefore, given the fact that the habitats of the gibbons in Thailand and the koalas in Australia are far away, and that bats are able to fly over long distances, the hypothesis that retroviruses of bats are the origin of GaLV and KoRV deserves consideration. Analysis of previous transspecies transmissions of retroviruses may help to evaluate the potential of transmission of related retroviruses in the future, e.g., that of porcine endogenous retroviruses (PERVs) during xenotransplantation using pig cells, tissues or organs.

  1. Monosodium luminol upregulates the expression of Bcl-2 and VEGF in retrovirus-infected mice through downregulation of corresponding miRNAs.

    PubMed

    Lungu, G; Kuang, X; Stoica, G; Wong, P K Y

    2010-01-01

    The retrovirus ts1 is a mutant of Moloney murine leukemia virus (MoMuLV) that causes neurodegeneration (ND) in susceptible mice. Our previous studies showed that the antioxidant drug monosodium luminol (GVT) prevented the development of ND in ts1-infected mice. In this study, we analyzed effect of GVT on the expression of B-cell lymphoma-2 protein (Bcl-2) and vascular endothelial growth factor (VEGF) in central nervous system (CNS) tissues of these animals. Our data showed that GVT treatment of ts1-infected mice significantly increased their expression of Bcl-2 and VEGF in brainstem compared with ts1-infected untreated mice. We also studied the expression of specific microRNAs (miRNAs) such as miRNA-15 and -16 (targeting Bcl-2), and miRNA-20 (targeting VEGF). We found that the expression of miRNAs inversely correlated with the upregulation of their target proteins in ts1-infected untreated as well as in GVT-treated-ts1-infected mice. The data showed that GVT treatment prevented ts1-induced ND at least in part by upregulating Bcl-2 and VEGF expression, what likely occurred as a consequence of downregulation of their corresponding miRNAs.

  2. Reduced expression of human endogenous retrovirus (HERV)-W GAG protein in the cingulate gyrus and hippocampus in schizophrenia, bipolar disorder, and depression.

    PubMed

    Weis, S; Llenos, I C; Sabunciyan, S; Dulay, J R; Isler, L; Yolken, R; Perron, H

    2007-01-01

    The human endogenous retrovirus (HERV)-W multicopy family was identified in human DNA from the previously characterized multiple sclerosis associated retroviral element (MSRV). Upregulation of the HERV-W POL has been reported in cerebrospinal fluid of patients with schizophrenia. The expression of capsid (GAG) protein of HERV-W was studied by immunohistochemistry and western blotting in postmortem brain tissue of the anterior cingulate cortex and hippocampal formation of normal controls and of patients with schizophrenia, bipolar disorder and major depression. A physiological expression of GAG protein was detected in neurons as well as astroglial cells in normal brain both in the anterior cingulate cortex and in the hippocampal formation. There was a statistically significant reduction of this expression in neurons and astroglial cells in brains from individuals with schizophrenia, major depression, and bipolar disorder. The results from the present study confirm that GAG protein encoded by the HERV-W multicopy gene family is expressed in cells of the central nervous system under normal conditions. Our findings of a cell type-, brain region- and disease-specific reduced expression in schizophrenia, major depression, and bipolar disorder are compatible with a pathophysiological role of HERVs in human brain disorders. The causes and biological consequences of this differential regulation will be the subject of further investigations.

  3. APOBEC3 proteins expressed in mammary epithelial cells are packaged into retroviruses and can restrict transmission of milk-borne virions.

    PubMed

    Okeoma, Chioma M; Huegel, Alyssa L; Lingappa, Jaisri; Feldman, Michael D; Ross, Susan R

    2010-12-16

    Viruses, including retroviruses like human immunodeficiency virus (HIV) and mouse mammary tumor virus (MMTV), are transmitted from mother to infants through milk. Lymphoid cells and antibodies are thought to provide mammary gland and milk-borne immunity. In contrast, little is known about the role of mammary epithelial cells (MECs). The APOBEC3 family of retroviral restriction factors is highly expressed in macrophages and lymphoid and dendritic cells. We now show that APOBEC3 proteins are also expressed in mouse and human MECs. Lymphoid cell-expressed APOBEC3 restricts in vivo spread of MMTV to lymphoid and mammary tissue. In contrast, mammary gland-expressed APOBEC3 is packaged into MMTV virions and decreases the infectivity of milk-borne viruses. Moreover, APOBEC3G and other APOBEC3 genes are expressed in human mammary cells and have the potential to restrict viruses produced in this cell type. These data point to a role for APOBEC3 proteins in limiting infectivity of milk-transmitted viruses.

  4. Murine myeloid cell lines derived by in vitro infection with recombinant c-myb retroviruses express myb from rearranged vector proviruses.

    PubMed Central

    Gonda, T J; Ramsay, R G; Johnson, G R

    1989-01-01

    To date, cellular transformation in vitro by the myb oncogene has been described for avian haemopoietic cells only. In order to exploit the well-characterized murine haemopoietic system to study transformation by myb, we have infected fetal liver cells with retroviral vectors carrying cDNAs that encode either complete or carboxy-terminally truncated c-myb proteins. We describe four cell lines which, despite our ability to efficiently infect haemopoietic target cells, were generated at low frequency. This was due, as least in part, to the requirement for a rearrangement within the vector that allowed expression of myb sequences. Three of the lines express a truncated myb protein while the fourth apparently expresses a normal c-myb protein, and thus constitutes an exception to the general association of truncation with transformation by myb. All four cell lines resemble immature cells of the myelomonocytic lineage and are dependent on colony-stimulating factors (CSFs) for their growth in vitro. One representative line could be converted to CSF-independence by infection with either Abelson murine leukaemia virus or a recombinant granulocyte-macrophage-CSF-encoding retrovirus; unlike the parental line, the resultant sublines were highly tumorigenic when injected into syngeneic mice. Images PMID:2670561

  5. Endogenous retroviruses in domestic animals.

    PubMed

    Garcia-Etxebarria, Koldo; Sistiaga-Poveda, Maialen; Jugo, Begoña Marina

    2014-08-01

    Endogenous retroviruses (ERVs) are genomic elements that are present in a wide range of vertebrates. Although the study of ERVs has been carried out mainly in humans and model organisms, recently, domestic animals have become important, and some species have begun to be analyzed to gain further insight into ERVs. Due to the availability of complete genomes and the development of new computer tools, ERVs can now be analyzed from a genome-wide viewpoint. In addition, more experimental work is being carried out to analyze the distribution, expression and interplay of ERVs within a host genome. Cats, cattle, chicken, dogs, horses, pigs and sheep have been scrutinized in this manner, all of which are interesting species in health and economic terms. Furthermore, several studies have noted differences in the number of endogenous retroviruses and in the variability of these elements among different breeds, as well as their expression in different tissues and the effects of their locations, which, in some cases, are near genes. These findings suggest a complex, intriguing relationship between ERVs and host genomes. In this review, we summarize the most important in silico and experimental findings, discuss their implications and attempt to predict future directions for the study of these genomic elements.

  6. 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.

  7. 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

  8. Long-term expression of human adenosine deaminase in mice transplanted with retrovirus-infected hematopoietic stem cells

    SciTech Connect

    Lim, B.; Apperley, J.F.; Orkin, S.H.; Williams, D.A. )

    1989-11-01

    Long-term stable expression of foreign genetic sequences transferred into hematopoietic stem cells by using retroviral vectors constitutes a relevant model for somatic gene therapy. Such stability of expression may depend on vector design, including the presence or absence of specific sequences within the vector, in combination with the nature and efficiency of infection of the hematopoietic target cells. The authors have previously reported successful transfer of human DNA encoding adenosine deaminase (ADA) into CFU-S (colony-forming unit-spleen) stem cells using simplified recombinant retroviral vectors. Human ADA was expressed in CFU-S-derived spleen colonies at levels near to endogenous enzyme. However, because of the lack of an efficient dominant selectable marker and low recombinant viral titers, stability of long-term expression of human ADA was not examined. They report here the development of an efficient method of infection of hematopoietic stem cells (HSC) without reliance on in vitro selection. Peripheral blood samples of 100% of mice transplanted with HSC infected by this protocol exhibit expression of human ADA 30 days after transplantation. Some mice (6 of 13) continue to express human ADA in all lineages after complete hematopoietic reconstitution (4 months). The use of recombinant retroviral vectors that efficiently transfer human ADA cDNA into HSC leading to stable expression of functional ADA in reconstituted mice, provides an experimental framework for future development of approaches to somatic gene therapy.

  9. Lack of retrovirus gene expression in somatic cell hybrids of friend cells and teratocarcinoma cells with a teratocarcinoma phenotype.

    PubMed Central

    Asche, W; Colletta, G; Warnecke, G; Nobis, P; Pennie, S; King, R M; Ostertag, W

    1984-01-01

    Two types of hybrids between cells with erythroid phenotype (Friend cells) and teratocarcinoma cells can be distinguished: cell hybrids with an erythroid phenotype, which release or can be induced to release large amounts of Friend spleen focus-forming virus (F-SFFV) on exposure to bromodeoxyuridine and cell hybrids with a teratocarcinoma phenotype, which do not release Friend virus and are not inducible for F-SFFV release. In this paper, we attempted to relate these differences to the expression of F-SFFV and Friend murine leukemia virus (F-MuLV) functions. Teratocarcinoma phenotype hybrids retained F-SFFV-and F-MuLV-related provirus sequences. They did not express F-SFFV- or F-MuLV-related RNA or proteins. The hybrids differentiated to endoderm-like cells on exposure to retinoic acid or hexamethylene-bis -acetamide. These cells, in contrast to the teratocarcinoma phenotype (uninduced) cells expressing SSEA-1-like antigens, did not express SSEA-1-like antigens; they formed typical, prekeratin-staining cytoskeletal structures and could be induced to release mouse interferon. The differentiating cells, but not the uninduced teratocarcinoma hybrids, were infected productively with F-MuLV or the F-MuLV--F-SFFV complex. They, however, did not express endogenous F-SFFV. Endogenous F-SFFV functions could not be rescued by infection with F-MuLV. Induction of teratocarcinoma hybrids with retinoic acid did not activate endogenous F-MuLV or F-SFFV transcription or protein synthesis. These data demonstrated two control mechanisms of Friend virus repression: one which acted trans during formation of the cell hybrids and was maintained only in teratocarcinoma phenotype cells and the other which acted cis and was still operative during induction of endodermal differentiation. Images PMID:6727874

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

    SciTech Connect

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

    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 vector 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.

  11. 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

  12. 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.

  13. 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

  14. Novel Endogenous Retrovirus in Rabbits Previously Reported as Human Retrovirus 5

    PubMed Central

    Griffiths, David J.; Voisset, Cécile; Venables, Patrick J. W.; Weiss, Robin A.

    2002-01-01

    Human retrovirus 5 (HRV-5) represented a fragment of a novel retrovirus sequence identified in human RNA and DNA preparations. In this study, the genome of HRV-5 was cloned and sequenced and integration sites were analyzed. Using PCR and Southern hybridization, we showed that HRV-5 is not integrated into human DNA. A survey of other species revealed that HRV-5 is present in the genomic DNA of the European rabbit (Oryctolagus cuniculus) and belongs to an endogenous retrovirus family found in rabbits. The presence of rabbit sequences flanking HRV-5 proviruses in human DNA extracts suggested that rabbit DNA was present in our human extracts, and this was confirmed by PCR analysis that revealed the presence of rabbit mitochondrial DNA sequences in four of five human DNA preparations tested. The origin of the rabbit DNA and HRV-5 in human DNA preparations remains unclear, but laboratory contamination cannot explain the preferential detection of HRV-5 in inflammatory diseases and lymphomas reported previously. This is the first description of a retrovirus genome in rabbits, and sequence analysis shows that it is related to but distinct from A-type retroelements of mice and other rodents. The species distribution of HRV-5 is restricted to rabbits; other species, including other members of the order Lagomorpha, do not contain this sequence. Analysis of HRV-5 expression by Northern hybridization and reverse transcriptase PCR indicates that the virus is transcribed at a low level in many rabbit tissues. In light of these findings we propose that the sequence previously designated HRV-5 should now be denoted RERV-H (for rabbit endogenous retrovirus H). PMID:12072509

  15. Analogy Between Lymphotropic Human Retroviruses and Large Animal Retroviruses

    PubMed Central

    Bouillant, Alain M.P.

    1986-01-01

    The family Retroviridae comprises some fifty viruses in three subfamilies: Oncoviridae, Lentiviridae and Spumaviridae. A better understanding of retroviral pathobiology has resulted from the rapid developments in knowledge of the molecular biology of normal and cancerous cells as well as retroviruses. Genomic relatedness was found between two human T cell leukemia viruses and bovine leukemia virus, similarly, some relatedness appears possible between human AIDS (acquired immunodeficiency syndrome) virus and lentiviruses of large animals. Because of their genomic relatedness, retroviruses from man and animals could theoretically form recombinants during in vitro manipulation. Therefore persons who work with retroviral materials should follow established laboratory practices to control infectious agents. PMID:17422719

  16. Biology and evolution of the endogenous koala retrovirus.

    PubMed

    Tarlinton, R; Meers, J; Young, P

    2008-11-01

    Although endogenous retroviruses are ubiquitous features of all mammalian genomes, the process of initial germ line invasion and subsequent inactivation from a pathogenic element has not yet been observed in a wild species. Koala retrovirus (KoRV) provides a unique opportunity to study this process of endogenisation in action as it still appears to be spreading through the koala population. Ongoing expression of the endogenous sequence and consequent high levels of viraemia have been linked to neoplasia and immunosuppression in koalas. This apparently recent invader of the koala genome shares a remarkably close sequence relationship with the pathogenic exogenous Gibbon ape leukaemia virus (GALV), and comparative analyses of KoRV and GALVare helping to shed light on how retroviruses in general adapt to a relatively benign or at least less pathogenic existence within a new host genome. (Part of a multi-author review).

  17. Novel Denisovan and Neanderthal retroviruses.

    PubMed

    Lee, Adam; Huntley, Derek; Aiewsakun, Pakorn; Kanda, Ravinder K; Lynn, Claire; Tristem, Michael

    2014-11-01

    Following the recent availability of high-coverage genomes for Denisovan and Neanderthal hominids, we conducted a screen for endogenized retroviruses, identifying six novel, previously unreported HERV-K(HML2) elements (HERV-K is human endogenous retrovirus K). These elements are absent from the human genome (hg38) and appear to be unique to archaic hominids. These findings provide further evidence supporting the recent activity of the HERV-K(HML2) group, which has been implicated in human disease. They will also provide insights into the evolution of archaic hominids.

  18. Human retroviruses and AIDS 1994

    SciTech Connect

    Myers, G.; Korber, B.; Wain-Hobson, S.; Jeang, Kuan-Teh; Henderson, L.E.; Pavlakis, G.N.

    1995-01-01

    This compendium, including accompanying floppy diskettes, is 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 it comprises: (I) Nucleic Acid Alignments and Sequences; (II) Amino Acid Alignments; (III) Analysis; (IV) Related Sequences; (V) Database communications.

  19. MuLV packaging systems as models for estimating/measuring retrovirus recombination frequency.

    PubMed

    Patience, C; Takeuch, Y; Cosset, F L; Weiss, R A

    2001-01-01

    Interaction of retrovirus vectors and endogenous retroviruses present in packaging cell lines and target cells may result in the formation of recombinant viruses. Using sensitive RT-PCR assays, we have investigated human and murine gene therapy packaging cell lines for the incorporation of endogenous retrovirus transcripts into murine leukaemia virus (MLV) vector particles and whether vector genomes are incorporated into human endogenous retrovirus (HERV) particles. VL30 endogenous retrovirus sequences were packaged in particles produced by the murine AM12 packaging system. For every seven MLV-derived -galactosidase beta-Gal vector genomes present in the particles, one copy of VL30 was also packaged. Although human FLY packaging cells expressed HERV transcripts (HERV-K, HuRT, type C, and RTVL-H), none was detectable in the MLV vector particles released from the cells. Non-specific packaging of the MLV gag-pol expression vector transcripts was detected in the FLY virions at a low level (one in 17,000 sequences). In other experiments, gag proteins produced by HERV-K particles present in human teratocarcinoma cells did not appear to package MLV-based vectors that expressed Gal transcripts. These findings indicate that retrovirus vectors interact with human packaging cells to produce retrovirus particles that are far less contaminated by endogenous viral sequences or other types of extraneous particles than murine packaging cells.

  20. Transspecies Transmission of the Endogenous Koala Retrovirus

    PubMed Central

    Fiebig, Uwe; Hartmann, Manuel Garcia; Bannert, Norbert; Kurth, Reinhard; Denner, Joachim

    2006-01-01

    The koala retrovirus (KoRV) is a gammaretrovirus closely related to the gibbon ape leukemia virus and induces leukemias and immune deficiencies associated with opportunistic infections, such as chlamydiosis. Here we characterize a KoRV newly isolated from an animal in a German zoo and show infection of human and rat cell lines in vitro and of rats in vivo, using immunological and PCR methods for virus detection. The KoRV transmembrane envelope protein (p15E) was cloned and expressed, and p15E-specific neutralizing antibodies able to prevent virus infection in vitro were developed. Finally, evidence for immunosuppressive properties of the KoRV was obtained. PMID:16699047

  1. Recent amplification of the kangaroo endogenous retrovirus, KERV, limited to the centromere.

    PubMed

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

    2011-05-01

    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.

  2. Heterogeneity of koala retrovirus isolates.

    PubMed

    Shimode, Sayumi; Nakagawa, So; Yoshikawa, Rokusuke; Shojima, Takayuki; Miyazawa, Takayuki

    2014-01-03

    Koala retrovirus (KoRV) is a gammaretrovirus which may induce immune suppression, leukemia and lymphoma in koalas. Currently three KoRV subgroups (A, B, and J) have been reported. Our phylogenetic analysis suggests that KoRV-B and KoRV-J should be classified as the same subgroup. In long terminal repeat (LTR), a KoRV-B isolate has four 17 bp tandem repeats named direct repeat (DR)-1, while a KoRV-J isolate (strain OJ-4) has three 37 bp tandem repeats named DR-2. We also found that the promoter activity of the KoRV-J strain OJ-4 is stronger than that of original KoRV-A, suggesting that KoRV-J may replicate more efficiently than KoRV-A.

  3. Involvement of Endogenous Retroviruses in Prion Diseases

    PubMed Central

    Lee, Yun-Jung; Jeong, Byung-Hoon; Choi, Eun-Kyung; Kim, Yong-Sun

    2013-01-01

    For millions of years, vertebrates have been continuously exposed to infection by retroviruses. Ancient retroviral infection of germline cells resulted in the formation and accumulation of inherited retrovirus sequences in host genomes. These inherited retroviruses are referred to as endogenous retroviruses (ERVs), and recent estimates have revealed that a significant portion of animal genomes is made up of ERVs. Although various host factors have suppressed ERV activation, both positive and negative functions have been reported for some ERVs in normal and abnormal physiological conditions, such as in disease states. Similar to other complex diseases, ERV activation has been observed in prion diseases, and this review will discuss the potential involvement of ERVs in prion diseases. PMID:25437206

  4. Expression of human factor IX in rabbit hepatocytes by retrovirus-mediated gene transfer: Potential for gene therapy of hemophilia B

    SciTech Connect

    Thompson, A.R. Puget Sound Blood Center, Seattle, WA ); Darlington, G. ); Armentano, D.; Woo, S.L.C.

    1990-08-01

    Hemophilia B (Christmas disease) is a chromosome X-linked blood clotting disorder which results when factor IX is deficient or functionally defective. The enzyme is synthesized in the liver, and the existence of animal models for this genetic disease will permit the development of somatic gene therapy protocols aimed at transfer of the functional gene into the liver. The authors report the construction of an N2-based recombinant retroviral vector, NCMVFIX, for efficient transfer and expression of human factor IX cDNA in primary rabbit hepatocytes. In this construct the human cytomegalovirus immediate early promoter directs the expression of factor IX. Hepatocytes were isolated from 3-week-old New Zealand White rabbits, infected with the recombinant virus, and analyzed for secretion of active factor IX. The infected rabbit hepatocytes produced human factor IX that is indistinguishable from enzyme derived from normal human plasma. The recombinant protein is sufficiently {gamma}-carboxylated and is functionally active in clotting assays. These results establish the feasibility of using infected hepatocytes for the expression of this protein and are a step toward the goal of correcting hemophilia B by hepatic gene transfer.

  5. Nonequilibirum assembly, retroviruses, and conical structures.

    PubMed

    Levandovsky, Artem; Zandi, Roya

    2009-05-15

    We study the spontaneous assembly of viral shells composed of several identical subunits under nonequilibrium conditions. We find that within the basic continuum elasticity framework, the nonequilibrium assembly process is able to predict the formation of structures pertinent to retroviruses. Our minimal model of assembly yields a unified one-dimensional phase diagram in which the appearance of spherical, irregular, conical and cylindrical structures of retroviruses is seen to be governed by the spontaneous curvature of protein subunits.

  6. World Reference Center for Arboviruses and Retroviruses

    DTIC Science & Technology

    1990-05-01

    GROUP SUD GROUP Arbo4irus, retrovirus, Cache Valley virus, phlebovirus, 󈧨 1 1 ELISA, yellow fever, Kaposi sarcoma , rapid diagnosis, 06 I 03 I I...aposi sarcoma patients was studied serologically. Reference reagents were distrihuted to scientists in over 2W) countries. 20 DISTRISUnON...demonstrate the origin of an introduced dengue virus. Retrovirus isolation capability was developed and a novel enveloped agent from tissues of Kaposi

  7. Transcription termination and polyadenylation in retroviruses.

    PubMed Central

    Guntaka, R V

    1993-01-01

    The provirus structure of retroviruses is bracketed by long terminal repeats (LTRs). The two LTRs (5' and 3') are identical in nucleotide sequence and organization. They contain signals for transcription initiation as well as termination and cleavage polyadenylation. As in eukaryotic pre-mRNAs, the two common signals, the polyadenylation signal, AAUAAA, or a variant AGUAAA, and the G+U-rich sequence are present in all retroviruses. However, the AAUAAA sequence is present in the U3 region in some retroviruses and in the R region in other retroviruses. As in animal cell RNAs, both AAUAAA and G+U-rich sequences apparently contribute to the 3'-end processing of retroviral RNAs. In addition, at least in a few cases examined, the sequences in the U3 region determine the efficiency of 3'-end processing. In retroviruses in which the AAUAAA is localized in the R region, the poly(A) signal in the 3' LTR but not the 5' LTR must be selectively used for the production of genomic RNA. It appears that the short distance between the 5' cap site and polyadenylation signal in the 5' LTR precludes premature termination and polyadenylation. Since 5' and 3' LTRs are identical in sequence and structural organization yet function differently, it is speculated that flanking cellular DNA sequences, chromatin structure, and binding of transcription factors may be involved in the functional divergence of 5' and 3' LTRs of retroviruses. PMID:7902524

  8. Perspectives on retroviruses and the etiologic agent of AIDS.

    PubMed Central

    Hsiung, G. D.

    1987-01-01

    In 1911, the first retrovirus was described: the Rous sarcoma virus, an avian retrovirus. Forty years later the murine leukemic virus, a mouse retrovirus, was reported. Although many other retroviruses from non-primate species were identified during the 1960s, the first primate retrovirus was not recognized until it was isolated from a monkey tumor in 1970. The search for human retroviruses in human leukemic cells remained unsuccessful at that time. Facilitated by the discovery of T-cell growth factor, a substance used for the propagation of human leukocytes in cultures, the first human retrovirus was discovered in 1980. Soon thereafter, in 1983, another human retrovirus, human immunodeficiency virus (HIV), was reported and implicated as the etiologic agent of AIDS. The isolation and identification of HIV has stimulated much interest in the study of human retroviruses and the control of this new viral disease. Images FIG. 2 FIG. 3 PMID:2829449

  9. 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

  10. Human retroviruses and neoplastic disease.

    PubMed

    Kaplan, M H

    1993-11-01

    Human retroviral infections result in significant neoplastic disease. Human T cell lymphotropic virus I (HTLV-I), the first human retrovirus to be discovered, is associated with the development of acute T cell leukemia with characteristic hypercalcemia and skin lesions after many years of chronic infection of CD4+ cells. HTLV-I also produces myelopathy. A minor T cell immunodeficiency occurs in HTLV-I acute T cell leukemia with associated strongyloidiasis and Pneumocystis carinii pneumonia. Human T cell lymphotropic virus II (HTLV-II) is found to be endemic in Amerindians and intravenous drug users (IVDUs) and has been linked to some cases of hairy-cell leukemia. HTLV-II infects the CD8+ population, with significant cell-associated viremia. Clinical neurological disease is rare, with one patient with myelopathy having been described. Immunodeficiency does not seem to occur. Human immunodeficiency virus 1 (HIV-1) produces aggressive large cell and Burkitt's lymphoma in as many as 10% of HIV-1-infected patients. More than 20% of homosexual men infected with HIV-1 develop Kaposi's sarcoma (KS). The pathogenesis of KS is better understood through studying KS-like cell lines that induce angiogenic factors. In some patients HIV-1 and HTLV-I or HTLV-II infections occur concomitantly. HIV-1 accelerates the tumorigenesis of HTLV-I and produces unusual skin diseases when combined with HTLV-II. Immunodeficiency occurs in all HIV-1-infected patients.

  11. Human retroviruses and aids, 1992

    SciTech Connect

    Myers, G.; Korber, B. ); Berzofsky, J.A.; Pavlakis, G.N. ); Smith, R.F. )

    1992-10-01

    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; (H) 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. 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. In addition to the general descriptions below of the parts of the compendium, the user should read the individual introductions for each part.

  12. Koala Retroviruses: Evolution and Disease Dynamics.

    PubMed

    Xu, Wenqin; Eiden, Maribeth V

    2015-11-01

    A retroviral etiology for malignant neoplasias in koalas has long been suspected. Evidence for retroviral involvement was bolstered in 2000 by the isolation of a koala retrovirus (KoRV), now termed KoRV-A. KoRV-A is an endogenous retrovirus-a retrovirus that infects germ cells-a feature that makes it a permanent resident of the koala genome. KoRV-A lacks the genetic diversity of an exogenous retrovirus, a quality associated with the ability of a retrovirus to cause neoplasias. In 2013, a second KoRV isolate, KoRV-B, was obtained from koalas with lymphomas in the Los Angeles Zoo. Unlike KoRV-A, which is present in the genomes of all koalas in the United States, KoRV-B is restricted in its distribution and is associated with host pathology (neoplastic disease). Here, our current understanding of the evolution of endogenous and exogenous KoRVs, and the relationship between them, is reviewed to build a perspective on the future impact of these viruses on koala sustainability.

  13. 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

    SciTech Connect

    Pech, M.; Gazit, A.; Arnstein, P.; Aaronson, S.A. )

    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 potent 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.

  14. Toward a More Accurate Quantitation of the Activity of Recombinant Retroviruses: Alternatives to Titer and Multiplicity of Infection

    PubMed Central

    Andreadis, Stylianos; Lavery, Thomas; Davis, Howard E.; Le Doux, Joseph M.; Yarmush, Martin L.; Morgan, Jeffrey R.

    2000-01-01

    In this paper, we present a mathematical model with experimental support of how several key parameters govern the adsorption of active retrovirus particles onto the surface of adherent cells. These parameters, including time of adsorption, volume of virus, and the number, size, and type of target cells, as well as the intrinsic properties of the virus, diffusion coefficient, and half-life (t1/2), have been incorporated into a mathematical expression that describes the rate at which active virus particles adsorb to the cell surface. From this expression, we have obtained estimates of Cvo, the starting concentration of active retrovirus particles. In contrast to titer, Cvo is independent of the specific conditions of the assay. The relatively slow diffusion (D = 2 × 10−8 cm2/s) and rapid decay (t1/2 = 6 to 7 h) of retrovirus particles explain why Cvo values are significantly higher than titer values. Values of Cvo also indicate that the number of defective particles in a retrovirus stock is much lower than previously thought, which has implications especially for the use of retroviruses for in vivo gene therapy. With this expression, we have also computed AVC (active viruses/cell), the number of active retrovirus particles that would adsorb per cell during a given adsorption time. In contrast to multiplicity of infection, which is based on titer and is subject to the same inaccuracies, AVC is based on the physicochemical parameters of the transduction assay and so is a more reliable alternative. PMID:10627536

  15. Human retroviruses and AIDS 1997

    SciTech Connect

    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 our 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.

  16. Genetic and molecular interactions of the Erv41p-Erv46p complex involved in transport between the endoplasmic reticulum and Golgi complex.

    PubMed

    Welsh, Leah M; Tong, Amy Hin Yan; Boone, Charles; Jensen, Ole N; Otte, Stefan

    2006-11-15

    Erv41p and Erv46p are integral membrane proteins conserved across species. They were originally identified as abundant constituents of COPII-coated vesicles, and form a complex which cycles between the endoplasmic reticulum and Golgi complex. Yeast strains lacking these proteins are viable but display subtle secretory phenotypes. In order to obtain information about possible biological roles of this protein complex in endoplasmic reticulum to Golgi transport, we employed the Synthetic Genetic Array approach to screen for synthetic genetic interactions with the erv46 null mutation. We identified synthetic interactions with vma12, vma21, vma22 and vps1 deletion mutations. The vma21Delta mutation exacerbates transport defects caused by the erv46Delta mutation. Unexpectedly, yeast strains lacking Vma21p fail to sort the endoplasmic reticulum to Golgi v-SNARE, Bos1p, efficiently into COPII vesicles, yet these vesicles are fully fusion competent. In addition, we set out to identify, by a biochemical approach, proteins interacting with the Erv41p-Erv46p complex. We report a strong interaction between the Erv41p-Erv46p complex and endoplasmic reticulum glucosidase II. Strains lacking a cycling Erv41p-Erv46p complex display a mild glycoprotein processing defect.

  17. 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

  18. COM, a heterochromatic locus governing the control of independent endogenous retroviruses from Drosophila melanogaster.

    PubMed

    Desset, Sophie; Meignin, Carine; Dastugue, Bernard; Vaury, Chantal

    2003-06-01

    ZAM and Idefix are two endogenous retroviruses whose expression is tightly controlled in Drosophila melanogaster. However, a line exists in which this control has been perturbed, resulting in a high mobilization rate for both retroviruses. This line is called the U (unstable) line as opposed to the other S (stable) lines. In the process of analyzing this control and tracing the genetic determinant involved, we found that ZAM and Idefix expression responded to two types of controls: one restricting their expression to specific somatic cells in the ovaries and the other silencing their expression in S lines but permitting it in U lines. While studying this second control in the U or S backgrounds, we found that the heterochromatic locus 20A2-3 on the X chromosome, previously implicated in the regulation of a third retroelement, gypsy, also controlled both ZAM and Idefix. We report here that genetic determinants necessary for endogenous retrovirus silencing occur at the 20A2-3 locus, which we call COM, for centre organisateur de mobilisation. We propose that if this point of control becomes mutated during the life of the fly, it may trigger processes reactivating dormant endogenous retroviruses and thus bring about sudden bursts of mobilization.

  19. Endogenous retroviruses regulate periimplantation placental growth and differentiation

    PubMed Central

    Dunlap, Kathrin A.; Palmarini, Massimo; Varela, Mariana; Burghardt, Robert C.; Hayashi, Kanako; Farmer, Jennifer L.; Spencer, Thomas E.

    2006-01-01

    Endogenous retroviruses (ERVs) are fixed and abundant in the genomes of vertebrates. Circumstantial evidence suggests that ERVs play a role in mammalian reproduction, particularly placental morphogenesis, because intact ERV envelope genes were found to be expressed in the syncytiotrophoblasts of human and mouse placenta and to elicit fusion of cells in vitro. We report here in vivo and in vitro experiments finding that the envelope of a particular class of ERVs of sheep, endogenous Jaagsiekte sheep retroviruses (enJSRVs), regulates trophectoderm growth and differentiation in the periimplantation conceptus (embryo/fetus and associated extraembryonic membranes). The enJSRV envelope gene is expressed in the trophectoderm of the elongating ovine conceptus after day 12 of pregnancy. Loss-of-function experiments were conducted in utero by injecting morpholino antisense oligonucleotides on day 8 of pregnancy that blocked enJSRV envelope protein production in the conceptus trophectoderm. This approach retarded trophectoderm outgrowth during conceptus elongation and inhibited trophoblast giant binucleate cell differentiation as observed on day 16. Pregnancy loss was observed by day 20 in sheep receiving morpholino antisense oligonucleotides. In vitro inhibition of the enJSRV envelope reduced the proliferation of mononuclear trophectoderm cells isolated from day 15 conceptuses. Consequently, these results demonstrate that the enJSRV envelope regulates trophectoderm growth and differentiation in the periimplantation ovine conceptus. This work supports the hypothesis that ERVs play fundamental roles in placental morphogenesis and mammalian reproduction. PMID:16980413

  20. 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

  1. 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

    2017-04-07

    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-based libraries, which takes advantage of retroviral superinfection resistance. However, the dynamic features of retrovirus-mediated transduction are not well-characterized. Here, we engineered two MSCV-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 harvested virion supernatant decreased by >60% after three days in storage. We found that retrovirus 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 retrovirus 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.Gene Therapy accepted article preview online, 07 April 2017. doi:10.1038/gt.2017.24.

  2. Rabbit endogenous retrovirus-H encodes a functional protease.

    PubMed

    Voisset, Cécile; Myers, Richard E; Carne, Alex; Kellam, Paul; Griffiths, David J

    2003-01-01

    Recent studies have revealed that 'human retrovirus-5' sequences found in human samples belong to a rabbit endogenous retrovirus family named RERV-H. A part of the gag-pro region of the RERV-H genome was amplified by PCR from DNA in human samples and several forms of RERV-H protease were expressed in bacteria. The RERV-H protease was able to cleave itself from a precursor protein and was also able to cleave the RERV-H Gag polyprotein precursor in vitro whereas a form of the protease with a mutation engineered into the active site was inactive. Potential N- and C-terminal autocleavage sites were characterized. The RERV-H protease was sensitive to pepstatin A, showing it to be an aspartic protease. Moreover, it was strongly inhibited by PYVPheStaAMT, a pseudopeptide inhibitor specific for Mason-Pfizer monkey virus and avian myeloblastosis-associated virus. A structural model of the RERV-H protease was constructed that, together with the activity data, confirms that this is a retroviral aspartic protease.

  3. Strategies of retrovirus survival in the cat.

    PubMed

    Jarrett, O

    1999-09-01

    Retroviruses establish persistent infections in their hosts which often lead to serious and fatal diseases after a long incubation period. The molecular basis of this persistence is the integration of a copy of the viral genome into cellular chromosomal DNA. At the level of the whole animal, however, each retrovirus genus has evolved a different strategy to ensure its survival. This variety is well illustrated in the cat. Feline leukaemia virus, an oncovirus, has a simple genomic structure and survives in its host by suppressing the immune response to the virus. As a result, this virus is antigenically highly conserved. By contrast, feline immunodeficiency virus and feline foamy virus, representatives of the lentiviruses and spumaviruses, respectively, have more complex genomes which include genes responsible for maintaining the virus in a latent state thereby avoiding elimination in the face of a powerful antiviral immune response. In the lentiviruses, this response drives the selection of viruses exhibiting variation in antigenicity and pathogenicity.

  4. [Endogenous retroviruses are associated with autoimmune diseases].

    PubMed

    Nexø, Bjørn A; Jensen, Sara B; Hansen, Bettina; Laska, Magdalena J

    2016-06-13

    Retroviruses can be transmitted in two fundamentally different ways: 1) They can be horizontally transmitted as infectious virus, or 2) they can integrate in the germ line and be transmitted to offspring and the offsprings' offspring as DNA. The latter is called endogenous viruses. The mode of transmission is called vertical. Viral variants of importance for development of disease must be more frequent among diseased persons than among healthy individuals. Multiple sclerosis, diabetes and rheumatoid arthritis are all associated with sets of endogenouos retroviruses but not the same sets. If a virus grows and this contributes to disease, one should be able to alleviate disease with antiretroviral drugs. We call for clinical trials to elucidate this issue.

  5. 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

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

    PubMed Central

    Gilroy, Kathryn L.; Terry, Anne; Naseer, Asif; de Ridder, Jeroen; Wang, Weiwei; Carpenter, Eric; Mason, Andrew; Wong, Gane K-S.; 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. PMID:27097319

  7. Human retroviruses and AIDS, 1991. [CONTAINS GLOSSARY

    SciTech Connect

    Myers, G.; Korber, B. ); Berzofsky, J.A.; Pavlakis, G.N. ); Smith, R.F. )

    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.

  8. Distribution of endogenous retroviruses in crocodilians.

    PubMed

    Jaratlerdsiri, Weerachai; Rodríguez-Zárate, Clara J; Isberg, Sally R; Damayanti, Chandramaya Siska; Miles, Lee G; Chansue, Nantarika; Moran, Chris; Melville, Lorna; Gongora, Jaime

    2009-10-01

    Knowledge of endogenous retroviruses (ERVs) in crocodilians (Crocodylia) is limited, and their distribution among extant species is unclear. Here we analyzed the phylogenetic relationships of these retroelements in 20 species of crocodilians by studying the pro-pol gene. The results showed that crocodilian ERVs (CERVs) cluster into two major clades (CERV 1 and CERV 2). CERV 1 clustered as a sister group of the genus Gammaretrovirus, while CERV 2 clustered distantly with respect to all known ERVs. Interestingly, CERV 1 was found only in crocodiles (Crocodylidae). The data generated here could assist future studies aimed at identifying orthologous and paralogous ERVs among crocodilians.

  9. Biochemical and proteomic characterization of retrovirus Gag based microparticles carrying melanoma antigens

    PubMed Central

    Kurg, Reet; Reinsalu, Olavi; Jagur, Sergei; Õunap, Kadri; Võsa, Liisi; Kasvandik, Sergo; Padari, Kärt; Gildemann, Kiira; Ustav, Mart

    2016-01-01

    Extracellular vesicles are membraneous particles released by a variety of cells into the extracellular microenvironment. Retroviruses utilize the cellular vesiculation pathway for virus budding/assembly and the retrovirus Gag protein induces the spontaneous formation of microvesicles or virus-like particles (VLPs) when expressed in the mammalian cells. In this study, five different melanoma antigens, MAGEA4, MAGEA10, MART1, TRP1 and MCAM, were incorporated into the VLPs and their localization within the particles was determined. Our data show that the MAGEA4 and MAGEA10 proteins as well as MCAM are expressed on the surface of VLPs. The compartmentalization of exogenously expressed cancer antigens within the VLPs did not depend on the localization of the protein within the cell. Comparison of the protein content of VLPs by LC-MS/MS-based label-free quantitative proteomics showed that VLPs carrying different cancer antigens are very similar to each other, but differ to some extent from VLPs without recombinant antigen. We suggest that retrovirus Gag based virus-like particles carrying recombinant antigens have a potential to be used in cancer immunotherapy. PMID:27403717

  10. Immunological relationships of an endogenous guinea pig retrovirus with prototype mammalian type B and type D retroviruses.

    PubMed Central

    Dahlberg, J E; Tronick, S R; Aaronson, S A

    1980-01-01

    A retrovirus endogenous to guinea pig cells was earlier shown to be morphologically similar to type B and type D prototype retroviruses. Molecular hybridization techniques were used to show that guinea pig virus nucleotide sequences are endogenous to both domestic (Cavia porcellus) and indigenous (Cavia aperea) guinea pigs, but cannot be detected in the DNA of either other hystricomorph rodents or other mammals tested. Using radioimmunological techniques designed to detect interspecies relationships, the major internal polypeptide of guinea pig virus (p26) was shown to share three different sets of interspecies antigenic determinants with squirrel monkey retrovirus, viper retrovirus, and mouse mammary tumor virus. Thus, guinea pig virus appears to provide an evolutionary link between type B and D retroviruses. Images PMID:6154150

  11. Conserved Region of Mammalian Retrovirus RNA

    PubMed Central

    Kominami, R.; Hatanaka, M.

    1979-01-01

    The viral RNAs of various mammalian retroviruses contain highly conserved sequences close to their 3′ ends. This was demonstrated by interviral molecular hybridization between fractionated viral complementary DNA (cDNA) and RNA. cDNA near the 3′ end (cDNA3′) from a rat virus (RPL strain) was fractionated by size and mixed with mouse virus RNA (Rauscher leukemia virus). No hybridization occurred with total cDNA (cDNAtotal), in agreement with previous results, but a cross-reacting sequence was found with the fractionated cDNA3′. The sequences between 50 to 400 nucleotides from the 3′ terminus of heteropolymeric RNA were most hybridizable. The rat viral cDNA3′ hybridized with mouse virus RNA more extensively than with RNA of remotely related retroviruses. The related viral sequence of the rodent viruses (mouse and rat) showed as much divergence in heteroduplex thermal denaturation profiles as did the unique sequence DNA of these two rodents. This suggests that over a period of time, rodent viruses have preserved a sequence with changes correlated to phylogenetic distance of hosts. The cross-reacting sequence of replication-competent retroviruses was conserved even in the genome of the replication-defective sarcoma virus and was also located in these genomes near the 3′ end of 30S RNA. A fraction of RD114 cDNA3′, corresponding to the conserved region, cross-hybridized extensively with RNA of a baboon endogenous virus (M7). Fractions of similar size prepared from cDNA3′ of MPMV, a primate type D virus, hybridized with M7 RNA to a lesser extent. Hybridization was not observed between Mason-Pfizer monkey virus and M7 if total cDNA's were incubated with viral RNAs. The degree of cross-reaction of the shared sequence appeared to be influenced by viral ancestral relatedness and host cell phylogenetic relationships. Thus, the strikingly high extent of cross-reaction at the conserved region between rodent viruses and simian sarcoma virus and between baboon

  12. Endogenous retrovirus and radiation-induced leukemia in the RMF mouse

    SciTech Connect

    Tennant, R.W.; Boone, L.R.; Lalley, P.; Yang, W.K.

    1982-01-01

    The induction of myeloid leukemia in irradiated RFM/Un mice has been associated with retrovirus infection. However, two characteristics of this strain complicate efforts to define the role of the virus. This strain possesses only one inducible host range class of endogenous virus and a unique gene, in addition to the Fv-1/sup n/ locus, which specifically restricts exogenous infection by endogenous viruses. These characteristics possibly account for absence of recombinant viruses in this strain, even though virus is amply expressed during most of the animal's life span. We have examined further the distribution of retrovirus sequences and the chromosomal locus of the inducible virus in this strain. This report describes evidence for additional viral sequences in cells of a radiation-induced myeloid leukemia line and discusses the possible origin of these added copies.

  13. Wolbachia Influences the Maternal Transmission of the gypsy Endogenous Retrovirus in Drosophila melanogaster

    PubMed Central

    Touret, Franck; Guiguen, François

    2014-01-01

    ABSTRACT 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. PMID:25182324

  14. Human endogenous retroviruses and chosen disease parameters in morphea

    PubMed Central

    Dańczak-Pazdrowska, Aleksandra; Szramka-Pawlak, Beata; Żaba, Ryszard; Osmola-Mańkowska, Agnieszka; Silny, Wojciech

    2017-01-01

    Introduction Morphea (localized scleroderma) is a relatively rare disease characterized by excessive skin fibrosis. Human endogenous retroviruses (HERV) are largely distributed within the human genome with hundreds of thousands of elements. The HERV have been widely studied in autoimmune disorders, yet hardly ever assessed in diseases with a good prognosis such as morphea. Aim In this study we focus on the possible relations between the expression of chosen HERV and factors influencing the pathomechanism of the disease, such as age, sex, titres of anti-nuclear antibodies, as well as duration, activity, and severity of the disease (LoSSI index). Material and methods Real-time polymerase chain reaction (PCR) targeting six HERV sequences of interest were performed on samples derived from peripheral blood mononuclear cells (PBMC) and skin biopsies. Results In PBMC we found a statistically significant negative correlation between HERV-W env expression and LoSSI index (p = 0.01). Additionally, HERV-W env was downregulated in patients with the active form of morphea. In all other cases we found no correlation whatsoever nor statistically significant differences below the p = 0.05 threshold. Conclusions Morphea seems to be an autoimmune disease where the impact of HERV is not so apparent. It seems that probing many patients for the expression of just a few sequences is not as effective as previously expected. For initial studies of HERV in other diseases we recommend high throughput techniques such as HERV-dedicated DNA microarrays or massive parallel sequencing. PMID:28261031

  15. A Modified γ-Retrovirus Vector for X-Linked Severe Combined Immunodeficiency

    PubMed Central

    Hacein-Bey-Abina, S.; Pai, S.-Y.; Gaspar, H.B.; Armant, M.; Berry, C.C.; Blanche, S.; Bleesing, J.; Blondeau, J.; de Boer, H.; Buckland, K.F.; Caccavelli, L.; Cros, G.; De Oliveira, S.; Fernández, K.S.; Guo, D.; Harris, C.E.; Hopkins, G.; Lehmann, L.E.; Lim, A.; London, W.B.; van der Loo, J.C.M.; Malani, N.; Male, F.; Malik, P.; Marinovic, M.A.; McNicol, A.-M.; Moshous, D.; Neven, B.; Oleastro, M.; Picard, C.; Ritz, J.; Rivat, C.; Schambach, A.; Shaw, K.L.; Sherman, E.A.; Silberstein, L.E.; Six, E.; Touzot, F.; Tsytsykova, A.; Xu-Bayford, J.; Baum, C.; Bushman, F.D.; Fischer, A.; Kohn, D.B.; Filipovich, A.H.; Notarangelo, L.D.; Cavazzana, M.; Williams, D.A.; Thrasher, A.J.

    2014-01-01

    BACKGROUND In previous clinical trials involving children with X-linked severe combined immunodeficiency (SCID-X1), a Moloney murine leukemia virus–based γ-retrovirus vector expressing interleukin-2 receptor γ-chain (γc) complementary DNA successfully restored immunity in most patients but resulted in vector-induced leukemia through enhancer-mediated mutagenesis in 25% of patients. We assessed the efficacy and safety of a self-inactivating retrovirus for the treatment of SCID-X1. METHODS We enrolled nine boys with SCID-X1 in parallel trials in Europe and the United States to evaluate treatment with a self-inactivating (SIN) γ-retrovirus vector containing deletions in viral enhancer sequences expressing γc (SIN-γc). RESULTS All patients received bone marrow–derived CD34+ cells transduced with the SIN-γc vector, without preparative conditioning. After 12.1 to 38.7 months of follow-up, eight of the nine children were still alive. One patient died from an overwhelming adenoviral infection before reconstitution with genetically modified T cells. Of the remaining eight patients, seven had recovery of peripheral-blood T cells that were functional and led to resolution of infections. The patients remained healthy thereafter. The kinetics of CD3+ T-cell recovery was not significantly different from that observed in previous trials. Assessment of insertion sites in peripheral blood from patients in the current trial as compared with those in previous trials revealed significantly less clustering of insertion sites within LMO2 , MECOM, and other lymphoid proto-oncogenes in our patients. CONCLUSIONS This modified γ-retrovirus vector was found to retain efficacy in the treatment of SCID-X1. The long-term effect of this therapy on leukemogenesis remains unknown. (Funded by the National Institutes of Health and others; ClinicalTrials.gov numbers, NCT01410019, NCT01175239, and NCT01129544.) PMID:25295500

  16. tirant, a Newly Discovered Active Endogenous Retrovirus in Drosophila simulans

    PubMed Central

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

    2012-01-01

    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. PMID:22278247

  17. Interferon but not MxB inhibits foamy retroviruses.

    PubMed

    Bähr, Ariane; Singer, Anna; Hain, Anika; Vasudevan, Ananda Ayyappan Jaguva; Schilling, Mirjam; Reh, Juliane; Riess, Maximilian; Panitz, Sylvia; Serrano, Vanessa; Schweizer, Matthias; König, Renate; Chanda, Sumit; Häussinger, Dieter; Kochs, Georg; Lindemann, Dirk; Münk, Carsten

    2016-01-15

    Foamy viruses (FV) are retroviruses that are widely distributed in primate and non-primate animal species. We tested here FV with capsids of simian and non-simian origin for sensitivity to interferon-β (IFN-β). Our data show significant inhibition of FV by IFN-β early in infection of human HOS and THP-1 but not of HEK293T cells. The post-entry restriction of FV was not mediated by the interferon-induced MxB protein that was recently identified as a capsid-interacting restriction factor targeting Human immunodeficiency virus (HIV) before integration. Neither the ectopic expression of MxA or MxB in HEK293T cells nor the lack of MxB expression in CRISPR/CAS MxB THP-1 knockout cells impacted the infection of the tested FV. IFN-β treated THP-1 and THP-1 KO MxB cells showed the same extend of restriction to FV. Together, the data demonstrate that IFN-β inhibits FV early in infection and that MxB is not a restriction factor of FV.

  18. Efficient intracellular retrotransposition of an exogenous primate retrovirus genome

    PubMed Central

    Heinkelein, Martin; Pietschmann, Thomas; Jármy, Gergely; Dressler, Marco; Imrich, Horst; Thurow, Jana; Lindemann, Dirk; Bock, Michael; Moebes, Astrid; Roy, Jacqueline; Herchenröder, Ottmar; Rethwilm, Axel

    2000-01-01

    The foamy virus (FV) subgroup of Retroviridae reverse transcribe their RNA (pre-)genome late in the replication cycle before leaving an infected cell. We studied whether a marker gene-transducing FV vector is able to shuttle to the nucleus and integrate into host cell genomic DNA. While a potential intracellular retrotransposition of vectors derived from other retroviruses was below the detection limit of our assay, we found that up to 5% of cells transfected with the FV vector were stably transduced, harboring 1 to ∼10 vector integrants. Generation of the integrants depended on expression of functional capsid, reverse transcriptase and integrase proteins, and did not involve an extracellular step. PCR analysis of the U3 region of the 5′ long terminal repeat and determination of proviral integration sites showed that a reverse transcription step had taken place to generate the integrants. Co-expression of a mutated envelope allowing particle egress and avoiding extracellular infection resulted in a significantly increased rescue of cells harboring integrants, suggesting that accumulation of proviruses via intracellular retrotransposition represents an integral part of the FV replication strategy. PMID:10880456

  19. A novel exogenous retrovirus sequence identified in humans.

    PubMed Central

    Griffiths, D J; Venables, P J; Weiss, R A; Boyd, M T

    1997-01-01

    A 932-bp retrovirus sequence was cloned by reverse transcriptase PCR from salivary gland tissue of a patient with Sjögren's syndrome. The sequence is related to that of type B and type D retroviruses and was present in a sucrose density gradient fraction corresponding to that of an enveloped retrovirus particle. Sequences amplified from tissues of eight individuals with or without Sjögren's syndrome had over 90% similarity and were present at a level of less than one copy per 10(3) cells. The sequence was not detectable in human genomic DNA by PCR or by Southern hybridization. These data indicate that the sequence represents an infectiously acquired genome, provisionally called human retrovirus 5. PMID:9060643

  20. Searching for Common Mammalian Retroviruses in Pediatric Idiopathic Diseases

    PubMed Central

    Jeziorski, Eric; Foulongne, Vincent; Ludwig, Catherine; Louhaem, Djamel; Rodiere, Michel; Sitbon, Marc; Courgnaud, Valérie

    2016-01-01

    Mammalian retroviruses cause a variety of diseases in their hosts, including hematological and immunodeficiency disorders. Both human T-cell leukemia (HTLV) and human immunodeficiency (HIV) viruses originated from several independent zoonotic transmissions, indicating that cross-species transmissions from animal to humans may still occur. Thus, as the risk for retroviral transmissions from animals to humans increase, we investigated whether mammalian retroviruses are involved in selected pediatric idiopathic diseases whose symptoms evoke retroviral infections. Blood samples, sera, and synovial fluids, or bone marrow cells were collected from pediatric patients under 18 years of age with different autoimmune idiopathic diseases. Overall, we screened clinical samples from 110 children using sensitive nested and semi-nested PCR strategies targeting env genes, and a C-type retrovirus reverse transcriptase (RT) activity kit. All clinical samples were free of retroviral signatures, indicating the unlikelihood of an etiological role of the retroviruses we assessed in the pediatric diseases we tested. PMID:27102168

  1. Embryonic stem cell potency fluctuates with endogenous retrovirus activity.

    PubMed

    Macfarlan, Todd S; Gifford, Wesley D; Driscoll, Shawn; Lettieri, Karen; Rowe, Helen M; Bonanomi, Dario; Firth, Amy; Singer, Oded; Trono, Didier; Pfaff, Samuel L

    2012-07-05

    Embryonic stem (ES) cells are derived from blastocyst-stage embryos and are thought to be functionally equivalent to the inner cell mass, which lacks the ability to produce all extraembryonic tissues. Here we identify a rare transient cell population within mouse ES and induced pluripotent stem (iPS) cell cultures that expresses high levels of transcripts found in two-cell (2C) embryos in which the blastomeres are totipotent. We genetically tagged these 2C-like ES cells and show that they lack the inner cell mass pluripotency proteins Oct4 (also known as Pou5f1), Sox2 and Nanog, and have acquired the ability to contribute to both embryonic and extraembryonic tissues. We show that nearly all ES cells cycle in and out of this privileged state, which is partially controlled by histone-modifying enzymes. Transcriptome sequencing and bioinformatic analyses showed that many 2C transcripts are initiated from long terminal repeats derived from endogenous retroviruses, suggesting this foreign sequence has helped to drive cell-fate regulation in placental mammals.

  2. Multiple invasions of an infectious retrovirus in cat genomes.

    PubMed

    Shimode, Sayumi; Nakagawa, So; Miyazawa, Takayuki

    2015-02-02

    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.

  3. Discovery of unfixed endogenous retrovirus insertions in diverse human populations

    PubMed Central

    Wildschutte, Julia Halo; Williams, Zachary H.; Montesion, Meagan; Subramanian, Ravi P.; Kidd, Jeffrey M.; Coffin, John M.

    2016-01-01

    Endogenous retroviruses (ERVs) have contributed to more than 8% of the human genome. The majority of these elements lack function due to accumulated mutations or internal recombination resulting in a solitary (solo) LTR, although members of one group of human ERVs (HERVs), HERV-K, were recently active with members that remain nearly intact, a subset of which is present as insertionally polymorphic loci that include approximately full-length (2-LTR) and solo-LTR alleles in addition to the unoccupied site. Several 2-LTR insertions have intact reading frames in some or all genes that are expressed as functional proteins. These properties reflect the activity of HERV-K and suggest the existence of additional unique loci within humans. We sought to determine the extent to which other polymorphic insertions are present in humans, using sequenced genomes from the 1000 Genomes Project and a subset of the Human Genome Diversity Project panel. We report analysis of a total of 36 nonreference polymorphic HERV-K proviruses, including 19 newly reported loci, with insertion frequencies ranging from <0.0005 to >0.75 that varied by population. Targeted screening of individual loci identified three new unfixed 2-LTR proviruses within our set, including an intact provirus present at Xq21.33 in some individuals, with the potential for retained infectivity. PMID:27001843

  4. 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.

  5. Clinical Aspects of Feline Retroviruses: A Review

    PubMed Central

    Hartmann, Katrin

    2012-01-01

    Feline leukemia virus (FeLV) and feline immunodeficiency virus (FIV) are retroviruses with global impact on the health of domestic cats. The two viruses differ in their potential to cause disease. FeLV is more pathogenic, and was long considered to be responsible for more clinical syndromes than any other agent in cats. FeLV can cause tumors (mainly lymphoma), bone marrow suppression syndromes (mainly anemia), and lead to secondary infectious diseases caused by suppressive effects of the virus on bone marrow and the immune system. Today, FeLV is less commonly diagnosed than in the previous 20 years; prevalence has been decreasing in most countries. However, FeLV importance may be underestimated as it has been shown that regressively infected cats (that are negative in routinely used FeLV tests) also can develop clinical signs. FIV can cause an acquired immunodeficiency syndrome that increases the risk of opportunistic infections, neurological diseases, and tumors. In most naturally infected cats, however, FIV itself does not cause severe clinical signs, and FIV-infected cats may live many years without any health problems. This article provides a review of clinical syndromes in progressively and regressively FeLV-infected cats as well as in FIV-infected cats. PMID:23202500

  6. Recombinant Origin of the Retrovirus XMRV

    PubMed Central

    Paprotka, Tobias; Delviks-Frankenberry, Krista A.; Cingöz, Oya; Martinez, Anthony; Kung, Hsing-Jien; Tepper, Clifford G.; Hu, Wei-Shau; Fivash, Matthew J.; Coffin, John M.; Pathak, Vinay K.

    2012-01-01

    The retrovirus XMRV (xenotropic murine leukemia virus-related virus) has been detected in human prostate tumors and in blood samples from patients with chronic fatigue syndrome, but these findings have not been replicated. We hypothesized that an understanding of when and how XMRV first arose might help explain the discrepant results. We studied human prostate cancer cell lines CWR22Rv1 and CWR-R1, which produce XMRV virtually identical to the viruses recently found in patient samples, as well as their progenitor human prostate tumor xenograft (CWR22) that had been passaged in mice. We detected XMRV infection in the two cell lines and in the later passage xenografts, but not in the early passages. Importantly, we found that the host mice contained two proviruses, PreXMRV-1 and PreXMRV-2, which share 99.92% identity with XMRV over >3.2-kb stretches of their genomes. We conclude that XMRV was not present in the original CWR22 tumor but was generated by recombination of two proviruses during tumor passaging in mice. The probability that an identical recombinant was generated independently is negligible (~10-12); our results suggest that the association of XMRV with human disease is due to contamination of human samples with virus originating from this recombination event. PMID:21628392

  7. 75 FR 79006 - Submission for OMB Review; Comment Request; Transfusion-Transmitted Retrovirus and Hepatitis...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-12-17

    ...- Transmitted Retrovirus and Hepatitis Virus Rates and Risk Factors: Improving the Safety of the U.S. Blood...: Transfusion-transmitted retrovirus and hepatitis virus rates and risk factors: Improving the safety of the...

  8. 75 FR 59724 - Proposed Collection; Comment Request; Transfusion-Transmitted Retrovirus and Hepatitis Virus...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-09-28

    ...-Transmitted Retrovirus and Hepatitis Virus Rates and Risk Factors: Improving the Safety of the U.S. Blood... and approval. Proposed Collection: Title: Transfusion-transmitted retrovirus and hepatitis virus...

  9. 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.

  10. Marine origin of retroviruses in the early Palaeozoic Era.

    PubMed

    Aiewsakun, Pakorn; Katzourakis, Aris

    2017-01-10

    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.

  11. Human APOBEC3 proteins, retrovirus restriction, and HIV drug resistance.

    PubMed

    Haché, Guylaine; Mansky, Louis M; Harris, Reuben S

    2006-01-01

    Over 40 million people worldwide currently have HIV/AIDS. Many antiretroviral drugs have proven effective, but drug-resistant HIV variants frequently emerge to thwart treatment efforts. Reverse transcription errors undoubtedly contribute to drug resistance, but additional significant sources of viral genetic variation are debatable. The human APOBEC3F and APOBEC3G proteins can potently inhibit retrovirus infection by a mechanism that involves retroviral cDNA cytosine deamination. Here we review the current knowledge on the mechanism of APOBEC3-dependent retrovirus restriction and discuss whether this innate host-defense system actively contributes to HIV genetic variation.

  12. Retrovirus delivered neurotrophin-3 promotes survival, proliferation and neuronal differentiation of human fetal neural stem cells in vitro.

    PubMed

    Lu, Haixia; Li, Minjie; Song, Tusheng; Qian, Yihua; Xiao, Xinli; Chen, Xinlin; Zhang, Pengbo; Feng, Xinshun; Parker, Terence; Liu, Yong

    2008-10-22

    Poor survival and insufficient neuronal differentiation are the main obstacles to neural stem cell (NSC) transplantation therapy. Genetic modification of NSCs with neurotrophins is considered a promising approach to overcome these difficulties. In this study, the effects on survival, proliferation and neuronal differentiation of human fetal NSCs (hfNSCs) were observed after infection by a neurotrophin-3 (NT-3) recombinant retrovirus. The hfNSCs, from 12-week human fetal brains formed neurospheres, expressed the stem cell marker nestin and differentiated into the three main cell types of the nervous system. NT-3 recombinant retrovirus (Retro-NT-3) infected hfNSCs efficiently expressed NT-3 gene for at least 8 weeks, presented an accelerated proliferation, and therefore produced an increased number of neurospheres and after differentiation in vitro, contained a higher percentage of neuronal cells. Eight weeks after infection, 37.9+/-4.2% of hfNSCs in the Retro-NT-3 infection group expressed the neuronal marker, this was significantly higher than the control and mock infection groups. NT-3 transduced hfNSCs also displayed longer protruding neurites compared with other groups. Combined these results demonstrate that NT-3 modification promote the survival/proliferation, neuronal differentiation and growth of neurites of hfNSCs in vitro. This study proposes recombinant retrovirus mediated NT-3 modification may provide a promising means to resolve the poor survival and insufficient neuronal differentiation of NSCs.

  13. Modulating Drug Resistance by Targeting BCRP/ABCG2 Using Retrovirus-Mediated RNA Interference

    PubMed Central

    Yuan, Jianhui; Liu, Wenlan; Deng, Tingting; Li, Zigang; Jin, Yi; Hu, Zhangli

    2014-01-01

    Background 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. Methodology/Principal Findings 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. Conclusions/Significance 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. PMID:25076217

  14. Multiple sclerosis retrovirus-like envelope gene: Role of the chromosome 20 insertion

    PubMed Central

    Varadé, Jezabel; García-Montojo, Marta; de la Hera, Belén; Camacho, Iris; García-Martínez, Mª. Ángel; Arroyo, Rafael; Álvarez-Lafuente, Roberto; Urcelay, Elena

    2015-01-01

    Background The genetic basis involved in multiple sclerosis (MS) susceptibility was not completely revealed by genome-wide association studies. Part of it could lie in repetitive sequences, as those corresponding to human Endogenous Retroviruses (HERVs). Retrovirus-like particles were isolated from MS patients and the genome of the MS-associated retrovirus (MSRV) was the founder of the HERV-W family. We aimed to ascertain which chromosomal origin encodes the pathogenic ENV protein by genomic analysis of the HERV-W insertions. Methods/results In silico analyses allowed to uncover putative open reading frames containing the specific sequence previously reported for MSRV-like envelope (env) detection. Out of the 261 genomic insertions of HERV-W env, only 9 copies harbor the specific primers and probe featuring MSRV-like env. The copy from chromosome 20 was further studied considering its size, a truncated homologue of the functional HERV-W env sequence encoding syncytin. High Resolution Melting analysis of this sequence identified two single nucleotide polymorphisms, subsequently genotyped by Taqman chemistry in 668 MS patients and 678 healthy controls. No significant association of these polymorphisms with MS risk was evidenced. Transcriptional activity of this MSRV-like env copy was detected in peripheral blood mononuclear cells from patients and controls. RNA expression levels of chromosome 20-specific MSRV-like env did not show significant differences between MS patients and controls, neither were related to genotypes of the two mentioned polymorphisms. Conclusions The lack of association with MS risk of the identified polymorphisms together with the transcription results discard chromosome 20 as genomic origin of MSRV-like env. PMID:26675450

  15. Pathology of tumors in fish associated with retroviruses: a review.

    PubMed

    Coffee, L L; Casey, J W; Bowser, P R

    2013-05-01

    Thirteen proliferative diseases in fish have been associated in the literature with 1 or more retroviruses. Typically, these occur as seasonal epizootics affecting farmed and wild fish, and most lesions resolve spontaneously. Spontaneous resolution and lifelong resistance to reinfection are 2 features of some piscine retrovirus-induced tumors that have stimulated research interest in this field. The purpose of this review is to present the reader with the epidemiological and morphological features of proliferative diseases in fish that have been associated with retroviruses by 1 or more of the following methods: detection of C-type retrovirus-like particles or reverse transcriptase activity in tumor tissues; successful tumor transmission trials using well-characterized, tumor-derived, cell-free inocula; or molecular characterization of the virus from spontaneous and experimentally induced tumors. Two of the diseases included in this review, European smelt spawning papillomatosis and bicolor damselfish neurofibromatosis, at one time were attributed to a retroviral etiology, but both are now believed to involve additional viral agents based on more recent investigations. We include the latter 2 entities to update the reader about these developments.

  16. Human retroviruses, cancer, and AIDS: Approaches to prevention and therapy

    SciTech Connect

    Bolognesi, D.

    1988-01-01

    This book contains eight sections, each consisting of several papers. The section titles are: New Human Retroviruses and Their Properties; Biology and Genetics of HIV; Products of the HIV Genome; Viral Pathogenesis; HIV and Its Receptor; Vaccine Approaches Against HIV; Treatment Approaches Against HIV; and Closing Remarks.

  17. Simian Retrovirus 4 Induces Lethal Acute Thrombocytopenia in Japanese Macaques

    PubMed Central

    Yoshikawa, Rokusuke; Sakaguchi, Shoichi; Nakagawa, So; Miura, Tomoyuki; Hirai, Hirohisa

    2015-01-01

    ABSTRACT In 2001-2002, six of seven Japanese macaques (Macaca fuscata) died after developing hemorrhagic syndrome at the Kyoto University Primate Research Institute (KUPRI). While the cause of death was unknown at the time, we detected simian retrovirus 4 (SRV-4) in samples obtained from a similar outbreak in 2008-2011, during which 42 of 43 Japanese macaques died after exhibiting hemorrhagic syndrome. In this study, we isolated SRV-4 strain PRI-172 from a Japanese macaque showing severe thrombocytopenia. When inoculated into four Japanese macaques, the isolate induced severe thrombocytopenia in all within 37 days. We then constructed an infectious molecular clone of strain PRI-172, termed pSR415, and inoculated the clone-derived virus into two Japanese macaques. These animals also developed severe thrombocytopenia in just 31 days after inoculation, and the virus was reisolated from blood, bone marrow, and stool. At necropsy, we observed bleeding from the gingivae and subcutaneous bleeding in all animals. SRV-4 infected a variety of tissues, especially in digestive organs, including colon and stomach, as determined by real-time reverse transcription-PCR (RT-PCR) and immunohistochemical staining. Furthermore, we identified the SRV-4 receptor as ASCT2, a neutral amino acid transporter. ASCT2 mRNA was expressed in a variety of tissues, and the distribution of SRV-4 proviruses in infected Japanese macaques correlated well with the expression levels of ASCT2 mRNA. From these results, we conclude that the causative agent of hemorrhagic syndrome in KUPRI Japanese macaques was SRV-4, and its receptor is ASCT2. IMPORTANCE During two separate outbreaks at the KUPRI, in 2001-2002 and 2008-2011, 96% of Japanese macaques (JM) that developed an unknown hemorrhagic syndrome died. Here, we isolated SRV-4 from a JM developing thrombocytopenia. The SRV-4 isolate and a molecularly cloned SRV-4 induced severe thrombocytopenia in virus-inoculated JMs within 37 days. At necropsy, we

  18. 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.

  19. Inhibiting DNA methylation causes an interferon response in cancer via dsRNA including endogenous retroviruses

    PubMed Central

    Chiappinelli, Katherine B.; Strissel, Pamela L.; Desrichard, Alexis; Li, Huili; Henke, Christine; Akman, Benjamin; Hein, Alexander; Rote, Neal S.; Cope, Leslie M.; Snyder, Alexandra; Makarov, Vladimir; Buhu, Sadna; Slamon, Dennis J.; Wolchok, Jedd D.; Pardoll, Drew M.; Beckmann, Matthias W.; Zahnow, Cynthia A.; Mergoub, Taha; Chan, Timothy A.; Baylin, Stephen B.; Strick, Reiner

    2015-01-01

    Summary We show that DNA methyltransferase inhibitors (DNMTis) upregulate immune signaling in cancer through the viral defense pathway. In ovarian cancer (OC), DNMTis trigger cytosolic sensing of double-stranded RNA (dsRNA) causing a Type I Interferon response and apoptosis. Knocking down dsRNA sensors TLR3 and MAVS reduces this response twofold, and blocking interferon beta or its receptor abrogates it. Upregulation of hypermethylated endogenous retrovirus (ERV) genes accompanies the response and ERV overexpression activates the response. Basal levels of ERV and viral defense gene expression significantly correlate in primary OC and the latter signature separates primary samples for multiple tumor types from The Cancer Genome Atlas into low versus high expression groups. In melanoma patients treated with an immune checkpoint therapy, high viral defense signature expression in tumors significantly associates with durable clinical response and DNMTi treatment sensitizes to anti-CTLA4 therapy in a pre-clinical melanoma model. PMID:26317466

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

    SciTech Connect

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

    2011-04-10

    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 of 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.

  1. Retroviruses 2004: review of the 2004 Cold Spring Harbor Retroviruses Conference.

    PubMed

    Freed, Eric O; Ross, Susan R

    2004-09-09

    For the past several decades, retrovirologists from around the world have gathered in late May at the Cold Spring Harbor Laboratories in New York to present their studies in formal talks and posters, and to discuss their ongoing research informally at the bar or on the beach. As organizers of the 2004 Cold Spring Harbor Retroviruses Conference, we have been asked by the editors of Retrovirology to prepare a review of the meeting for publication on-line. Our goal in this review is not to provide a detailed description of data presented at the meeting but rather to highlight some of the significant developments reported this year. The review is structured in a manner that parallels the organization of the meeting; beginning with the entry phase of the replication cycle, proceeding with post-entry events, assembly and release, integration, reverse transcription, pathogenesis/host factors, RNA-related events (transcription, processing, export, and packaging) and finishing with antivirals. While the most striking developments this year involved post-entry events and assembly/release, significant progress was made towards elucidating a number of aspects of the retroviral replication cycle.

  2. Lack of antiviral antibody response in koalas infected with koala retroviruses (KoRV).

    PubMed

    Fiebig, Uwe; Keller, Martina; Möller, Annekatrin; Timms, Peter; Denner, Joachim

    2015-02-16

    Many wild koalas are infected with the koala retrovirus, KoRV, some of which suffer from lymphoma and chlamydial disease. Three subgroups, KoRV-A, KoRV-B and KoRV-J, have so far been described. It is well known that other closely related gammaretroviruses can induce tumours and severe immunodeficiencies in their respective hosts and a possible role for KoRV infection in lymphoma and chlamydial disease in koalas has been suggested. In many wild koalas, KoRV-A has become endogenised, i.e., it is integrated in the germ-line and is passed on with normal cellular genes. In this study, sera from koalas in European zoos and from wild animals in Australia were screened for antibodies against KoRV-A. These naturally infected animals all carry endogenous KoRV-A and two zoo animals are also infected with KoRV-B. The antibody response is generally an important diagnostic tool for detecting retrovirus infections. However, when Western blot analyses were performed using purified virus or recombinant proteins corresponding to KoRV-A, none of the koalas tested positive for specific antibodies, suggesting a state of tolerance. These results have implications for koala vaccination, as they suggest that therapeutic immunisation of animals carrying and expressing endogenous KoRV-A will not be successful. However, it remains unclear whether these animals can be immunised against KoRV-B and immunisation of uninfected koalas could still be worthwhile.

  3. Association of Endogenous Retroviruses and Long Terminal Repeats with Human Disorders

    PubMed Central

    Katoh, Iyoko; Kurata, Shun-ichi

    2013-01-01

    Since the human genome sequences became available in 2001, our knowledge about the human transposable elements which comprise ∼40% of the total nucleotides has been expanding. Non-long terminal repeat (non-LTR) retrotransposons are actively transposing in the present-day human genome, and have been found to cause ∼100 identified clinical cases of varied disorders. In contrast, almost all of the human endogenous retroviruses (HERVs) originating from ancient infectious retroviruses lost their infectivity and transposing activity at various times before the human-chimpanzee speciation (∼6 million years ago), and no known HERV is presently infectious. Insertion of HERVs and mammalian apparent LTR retrotransposons (MaLRs) into the chromosomal DNA influenced a number of host genes in various modes during human evolution. Apart from the aspect of genome evolution, HERVs and solitary LTRs being suppressed in normal biological processes can potentially act as extra transcriptional apparatuses of cellular genes by re-activation in individuals. There has been a reasonable prediction that aberrant LTR activation could trigger malignant disorders and autoimmune responses if epigenetic changes including DNA hypomethylation occur in somatic cells. Evidence supporting this hypothesis has begun to emerge only recently: a MaLR family LTR activation in the pathogenesis of Hodgkin’s lymphoma and a HERV-E antigen expression in an anti-renal cell carcinoma immune response. This mini review addresses the impacts of the remnant-form LTR retrotransposons on human pathogenesis. PMID:24062987

  4. The association between human endogenous retroviruses and multiple sclerosis: A systematic review and meta-analysis

    PubMed Central

    Morandi, Elena; Tanasescu, Radu; Tarlinton, Rachael E.; Constantinescu, Cris S.; Zhang, Weiya; Tench, Christopher

    2017-01-01

    Background The interaction between genetic and environmental factors is crucial to multiple sclerosis (MS) pathogenesis. Human Endogenous Retroviruses (HERVs) are endogenous viral elements of the human genome whose expression is associated with MS. Objective To perform a systematic review and meta-analysis and to assess qualitative and quantitative evidence on the expression of HERV families in MS patients. Methods Medline, Embase and the Cochrane Library were searched for published studies on the association of HERVs and MS. Meta-analysis was performed on the HERV-W family. Odds Ratio (OR) and 95% confidence interval (CI) were calculated for association. Results 43 reports were extracted (25 related to HERV-W, 13 to HERV-H, 9 to HERV-K, 5 to HRES-1 and 1 to HER-15 family). The analysis showed an association between expression of all HERV families and MS. For HERV-W, adequate data was available for meta-analysis. Results from meta-analyses of HERV-W were OR = 22.66 (95%CI 6.32 to 81.20) from 4 studies investigating MSRV/HERV-W (MS-associated retrovirus) envelope mRNA in peripheral blood mononuclear cells, OR = 44.11 (95%CI 12.95 to 150.30) from 6 studies of MSRV/HERV-W polymerase mRNA in serum/plasma and OR = 6.00 (95%CI 3.35 to 10.74) from 4 studies of MSRV/HERV-W polymerase mRNA in CSF. Conclusions This systematic review and meta-analysis shows an association between expression of HERVs, and in particular the HERV-W family, and MS. PMID:28207850

  5. Role of endogenous cat retrovirus in cell differentiation.

    PubMed Central

    Rasheed, S

    1982-01-01

    Several long-term cultures were established from a spontaneous melanoma of a cat. Cells were rounded or spindle shaped and exhibited black/brown pigmentation in the cytoplasm. No virus was released from these cells spontaneously or after treatment with chemicals. However, exogenous infection of the cat melanoma cells with the endogenous cat virus RD114 resulted in remarkable morphological and functional changes. Most of the RD114 virus-infected cells exhibited multiple neuritic extensions and about 1-2% of the population showed characteristics of neuronal cells. Because human, mouse, and hamster melanoma cultures infected with various mammalian retroviruses, including the RD114 virus, did not display any morphological alteration, it is concluded that the neuronal cell differentiation in the cat melanoma cells is a consequence of its specific interaction with the endogenous cat retrovirus. Images PMID:6961415

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

    PubMed Central

    Martin, Jessica L.; Cao, Sheng; Maldonado, Jose O.

    2016-01-01

    ABSTRACT 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. IMPORTANCE Comparative analysis among retroviruses has been critically important in

  7. On board a raft or boat in the retrovirus sea

    PubMed Central

    Svoboda, Jan

    2016-01-01

    This article summarizes the essential steps in understanding the chicken Rous sarcoma virus (RSV) genome association with a nonpermissive rodent host cell genome. This insight was made possible by in-depth study of RSV-transformed rat XC cells, which were called virogenic because they indefinitely carry virus genetic information in the absence of any infectious virus production. However, the virus was rescued by association of XC cells with chicken fibroblasts, allowing cell fusion between both partners. This and additional studies led to the interpretation that the RSV genome gets integrated into the host cell genome as a provirus. Study of additional rodent virogenic cell lines provided evidence that the transcript of oncogene v-src can be transmitted to other retroviruses and produce cell transformation by itself. As discussed in the text, two main questions related to nonpermissiveness to retrovirus infection remain to be solved. The first is changes in the retrovirus envelope gene allowing virus entry into a nonpermissive cell. The second is the nature of the permissive cell functions required by the nonpermissive cell to ensure infectious virus production. Both lines of investigation are being pursued. PMID:27035994

  8. Retroviruses integrate into a shared, non-palindromic DNA motif.

    PubMed

    Kirk, Paul D W; Huvet, Maxime; Melamed, Anat; Maertens, Goedele N; Bangham, Charles R M

    2016-11-14

    Many DNA-binding factors, such as transcription factors, form oligomeric complexes with structural symmetry that bind to palindromic DNA sequences(1). Palindromic consensus nucleotide sequences are also found at the genomic integration sites of retroviruses(2-6) and other transposable elements(7-9), and it has been suggested that this palindromic consensus arises as a consequence of the structural symmetry in the integrase complex(2,3). However, we show here that the palindromic consensus sequence is not present in individual integration sites of human T-cell lymphotropic virus type 1 (HTLV-1) and human immunodeficiency virus type 1 (HIV-1), but arises in the population average as a consequence of the existence of a non-palindromic nucleotide motif that occurs in approximately equal proportions on the plus strand and the minus strand of the host genome. We develop a generally applicable algorithm to sort the individual integration site sequences into plus-strand and minus-strand subpopulations, and use this to identify the integration site nucleotide motifs of five retroviruses of different genera: HTLV-1, HIV-1, murine leukaemia virus (MLV), avian sarcoma leucosis virus (ASLV) and prototype foamy virus (PFV). The results reveal a non-palindromic motif that is shared between these retroviruses.

  9. Review of the twelfth West Coast Retrovirus Meeting.

    PubMed

    Barry, Sheila M; Melar, Marta; Gallay, Philippe; Hope, Thomas J

    2005-11-17

    Every year the Cancer Research Institute from University of California at Irvine organizes the West Coast Retrovirus Meeting where participants have a chance to discuss the latest progress in understanding the pathology of retroviruses. The 12th meeting was held at the Hyatt Regency Suites in Palm Springs, California from October 6th to October 9th 2005, with the major focus on human immunodeficiency virus (HIV) pathogenesis. Philippe Gallay from The Scripps Research Institute and Thomas J. Hope from Northwestern University organized the meeting, which covered all the steps involved in the lifecycle of retroviruses with an emphasis on virus:host interactions. The trend in research appeared to be on the restriction of viral infection, both by the endogenous, cellular restriction factors, as well as by the potential antimicrobial compounds of known or unknown mechanisms. Additionally, new stories on the inevitable feedback from the host immune system were presented as well. HIV still represents a challenge that an army of motivated people has been working on for over 20 years. And yet, the field has not reached the plateau in knowledge nor enthusiasm, which was proven again in October 2005 in Palm Springs.

  10. Adeno-associated virus type 2-mediated transfer of ecotropic retrovirus receptor cDNA allows ecotropic retroviral transduction of established and primary human cells.

    PubMed

    Qing, K; Bachelot, T; Mukherjee, P; Wang, X S; Peng, L; Yoder, M C; Leboulch, P; Srivastava, A

    1997-07-01

    The cellular receptors that mediate binding and internalization of retroviruses have recently been identified. The concentration and accessibility of these receptors are critical determinants in accomplishing successful gene transfer with retrovirus-based vectors. Murine retroviruses containing ecotropic glycoproteins do not infect human cells since human cells do not express the receptor that binds the ecotropic glycoproteins. To enable human cells to become permissive for ecotropic retrovirus-mediated gene transfer, we have developed a recombinant adeno-associated virus type 2 (AAV) vector containing ecotropic retroviral receptor (ecoR) cDNA under the control of the Rous sarcoma virus (RSV) long terminal repeat (LTR) promoter (vRSVp-ecoR). Established human cell lines, such as HeLa and KB, known to be nonpermissive for murine ecotropic retroviruses, became permissive for infection by a retroviral vector containing a bacterial gene for resistance to neomycin (RV-Neo(r)), with a transduction efficiency of up to 47%, following transduction with vRSVp-ecoR, as determined by the development of colonies that were resistant to the drug G418, a neomycin analog. No G418-resistant colonies were present in cultures infected with either vRSVp-ecoR or RV-Neo(r) alone. Southern and Northern blot analyses revealed stable integration and long-term expression, respectively, of the transduced murine ecoR gene in clonal isolates of HeLa and KB cells. Similarly, ecotropic retrovirus-mediated Neo(r) transduction of primary human CD34+ hematopoietic progenitor cells from normal bone marrow was also documented, but only following infection with vRSVp-ecoR. The retroviral transduction efficiency was approximately 7% without prestimulation and approximately 14% with prestimulation of CD34+ cells with cytokines, as determined by hematopoietic clonogenic assays. No G418-resistant progenitor cell colonies were present in cultures infected with either vRSVp-ecoR or RV-Neo(r) alone. These

  11. HIV-1 Vpu Promotes Release and Prevents Endocytosis of Nascent Retrovirus Particles from the Plasma Membrane

    PubMed Central

    Neil, Stuart J. D; Eastman, Scott W; Jouvenet, Nolwenn; Bieniasz, Paul D

    2006-01-01

    The human immunodeficiency virus (HIV) type-1 viral protein U (Vpu) protein enhances the release of diverse retroviruses from human, but not monkey, cells and is thought to do so by ablating a dominant restriction to particle release. Here, we determined how Vpu expression affects the subcellular distribution of HIV-1 and murine leukemia virus (MLV) Gag proteins in human cells where Vpu is, or is not, required for efficient particle release. In HeLa cells, where Vpu enhances HIV-1 and MLV release approximately 10-fold, concentrations of HIV-1 Gag and MLV Gag fused to cyan fluorescent protein (CFP) were initially detected at the plasma membrane, but then accumulated over time in early and late endosomes. Endosomal accumulation of Gag-CFP was prevented by Vpu expression and, importantly, inhibition of plasma membrane to early endosome transport by dominant negative mutants of Rab5a, dynamin, and EPS-15. Additionally, accumulation of both HIV and MLV Gag in endosomes required a functional late-budding domain. In human HOS cells, where HIV-1 and MLV release was efficient even in the absence of Vpu, Gag proteins were localized predominantly at the plasma membrane, irrespective of Vpu expression or manipulation of endocytic transport. While these data indicated that Vpu inhibits nascent virion endocytosis, Vpu did not affect transferrin endocytosis. Moreover, inhibition of endocytosis did not restore Vpu-defective HIV-1 release in HeLa cells, but instead resulted in accumulation of mature virions that could be released from the cell surface by protease treatment. Thus, these findings suggest that a specific activity that is present in HeLa cells, but not in HOS cells, and is counteracted by Vpu, traps assembled retrovirus particles at the cell surface. This entrapment leads to subsequent endocytosis by a Rab5a- and clathrin-dependent mechanism and intracellular sequestration of virions in endosomes. PMID:16699598

  12. Structural characterization of reverse transcriptase and endonuclease polypeptides of the acquired immunodeficiency syndrome retrovirus.

    PubMed Central

    Lightfoote, M M; Coligan, J E; Folks, T M; Fauci, A S; Martin, M A; Venkatesan, S

    1986-01-01

    Automated N-terminal microsequencing of immune affinity-purified acquired immunodeficiency syndrome retrovirus polypeptides from infected cells was used to locate the N termini of 64-, 51-, and 34-kilodalton (kDa) polypeptides within the pol open reading frame (ORF) of the proviral DNA. The 64- and 51-kDa proteins had identical N termini (Pro-Ile-Ser-Pro-IIe-Glu-Thr-Val-) positioned 156 residues from the beginning of the pol ORF. The N terminus of the 34-kDa pol gene product, Phe-Leu-Asp-Gly-Ile-Asp-Lys-, mapped 716 residues into the pol ORF. These polypeptides were absent in an RT-negative, CD4-negative, persistently infected cell line (8E5) carrying a single defective copy of a constitutively expressed, integrated proviral DNA. Images PMID:2430111

  13. Insertional polymorphisms: a new lease of life for endogenous retroviruses in human disease.

    PubMed

    Moyes, David; Griffiths, David J; Venables, Patrick J

    2007-07-01

    Human endogenous retroviruses (HERVs) result from ancestral infection by infectious viruses over millions of years of primate evolution. Some are transcriptionally active, express proteins and therefore have the potential to cause disease. Here we review the controversial attempts to link them with cancer and autoimmunity. The main difficulty is that most HERVs investigated to date are present at the same locus in 100% of the population. However, a new class of insertionally polymorphic HERV-K family members, present in a minority of individuals, has recently been described. We propose that insertionally polymorphic HERVs could be novel genetic risk factors and hence provide a new lease of life for research into HERVs and disease.

  14. The use of retroviruses as pharmaceutical tools for target discovery and validation in the field of functional genomics.

    PubMed

    Lorens, J B; Sousa, C; Bennett, M K; Molineaux, S M; Payan, D G

    2001-12-01

    Retrovirally mediated functional genomics enables identification of physiologically relevant cellular therapeutic targets. Unique properties of retroviruses make them ideal tools for the introduction of large and diverse libraries of potential genetic effectors to a variety of cell types. The identification and recovery of intracellular library elements responsible for altered disease responses establishes a direct basis for pharmaceutical development. Recent innovations in retroviral infection efficiency and expression control have broadened application of the methodology to include libraries of mutagenized cDNAs, peptides and ribozyme genetic effectors.

  15. A paradigm for virus-host coevolution: Sequential counter-adaptations between endogenous and exogenous retroviruses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Endogenous retroviruses (ERVs) are remnants of ancient retroviral infections of the host germline transmitted vertically from generation to generation. It is hypothesized that during evolution some ERVs were used by the host to drive extinction of exogenous horizontally-transmitted retroviruses. Se...

  16. The aliens inside human DNA: HERV-W/MSRV/syncytin-1 endogenous retroviruses and neurodegeneration.

    PubMed

    Dolei, Antonina; Uleri, Elena; Ibba, Gabriele; Caocci, Maurizio; Piu, Claudia; Serra, Caterina

    2015-07-04

    The human genome contains remnants of ancestral retroviruses now endogenously transmitted, called human endogenous retroviruses (HERVs). HERVs can be variably expressed, and both beneficial and detrimental effects have described. This review focuses on the MSRV and syncytin-1 HERV-W elements in relationship to neurodegeneration in view of their neuro-pathogenic and immune-pathogenic properties. Multiple sclerosis (MS) and a neurodegenerative disease (neuroAIDS) are reported in this review. In vivo studies in patients and controls for molecular epidemiology and follow-up studies are reviewed, along with in vitro cellular studies of the effects of treatments and of molecular mechanisms. HERV-W/MSRV has been repeatedly found in MS patients (in blood, spinal fluid, and brain samples), and MRSV presence/load strikingly parallels MS stages and active/remission phases, as well as therapy outcome. The DNA of MS patients has increased MSRVenv copies, while syncytin-1 copies are unchanged in controls. Presence of MSRV in the spinal fluid predicted the worst MS progression, ten years in advance. The Epstein-Barr virus (EBV) activates HERV-W/MSRV both in vitro and in vivo. With respect to neuroAIDS, the HIV transactivator of transcription (Tat) protein activates HERV-W/MSRV in monocytes/macrophages and astrocytes indirectly by interaction with TLR4 and induction of TNFa. HERV-W/MSRV can be considered a biomarker for MS behavior and therapy outcome. Regarding MS pathogenesis, we postulate the possibility for EBV of an initial trigger of future MS, years later, and for MSRV of a direct role of effector of neuropathogenesis during MS. Additionally, HERV-W/MSR/syncytin-1 activation by HIV Tat could contribute to the HIV-related neurodegeneration.

  17. 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

  18. Hepadnaviruses and retroviruses share genome homology and features of replication.

    PubMed

    Robinson, W S; Miller, R H; Marion, P L

    1987-01-01

    The hepadnavirus family includes hepatitis B virus (HBV), woodchuck hepatitis virus (WHV), ground squirrel hepatitis virus (GSHV) and duck hepatitis B virus (DHBV). These viruses share unique ultrastructural, molecular and biological features. HBV has great medical importance in many parts of the world. More important numerically than acute hepatitis B in high prevalence geographic regions is the liver disease associated with chronic infection. There appear to be more than 200 million chronically infected humans in the world, and these HBV infections appear to be the single most common cause of chronic liver disease and liver cancer in man. All hepadnaviruses share the propensity for silent infection in early life leading to persistence of the virus, and hepatocellular carcinoma (HCC) is clearly associated with long-standing persistent infection in man, woodchucks and ground squirrels. Although the viral DNA has been found to be integrated in cellular DNA of many HCC in man, woodchucks and ground squirrels, the precise role of the virus in tumor formation has not been defined. Hepadna viruses have an interesting molecular structure and mechanisms of replication, and they appear to share certain important features with retroviruses as reviewed here. It is of interest to define similarities and differences between hepadnaviruses and retroviruses in order to understand their evolutionary relationship and to determine whether they share a common oncogenic mechanism, since infection with members of both virus families is associated with neoplastic disease.

  19. Seroepidemiology of human retroviruses in Ogun State of Nigeria.

    PubMed

    Olusanya, O; Lawoko, A; Blomberg, J

    1990-01-01

    We analyzed sera collected during 1987 and 1988 from 385 healthy business employees of both sexes, of Ogun state in Nigeria, for antibodies to the 3 human retroviruses HIV-1, HIV-2 and HTLV-I. No serum was HIV-1 positive, 1 was HIV-2 positive and 2 were HTLV-I positive. A few sera were false-positive in the antibody screening tests which preceded the confirmatory antibody tests. In the confirmatory tests, we found that in the HIV-1 Western blot test 1 serum reacted only with the HIV-1 gag protein p17, and 2 sera reacted only with the HIV-1 pol proteins p64, p53 and p31. None of these reactivities fulfill internationally accepted criteria for HIV-1 seropositivity. We conclude that HIV-1 was rare in the study population and that HIV-2 and HTLV-I are present at a low frequency. The false positive serological reactions observed are similar to those described previously from Africa and elsewhere. The findings emphasize the importance of routinely testing blood donations for antibodies to these retroviruses in Nigeria.

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

    PubMed Central

    Coffin, John M.

    2015-01-01

    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. PMID:26696625

  1. Receptor co-operation in retrovirus entry: recruitment of an auxiliary entry mechanism after retargeted binding.

    PubMed Central

    Valsesia-Wittmann, S; Morling, F J; Hatziioannou, T; Russell, S J; Cosset, F L

    1997-01-01

    We have constructed Moloney murine leukemia virus (MoMLV)-derived envelope glycoproteins (AMO) displaying an amino-terminal Ram-1-binding domain in which a variety of different amino acid spacers have been inserted between the displayed domain and the MoMLV surface (SU) subunit. Titres of retroviruses generated with these chimeric envelopes were enhanced on cells expressing both Ram-1 and Rec-1 receptors compared with the titres on cells expressing only one or other receptor type. The absolute viral titres and the degree of titre enhancement due to receptor cooperativity were highly variable between the different chimeric envelopes and were determined primarily by the properties of the interdomain spacer. An extreme example of receptor co-operativity was encountered when testing Ram-1-targeted AMOPRO envelopes with specific proline-rich interdomain spacers. AMOPRO viruses could not enter cells expressing only Rec-1 or only Ram-1 but could efficiently infect cells co-expressing both receptors. The data are consistent with a model for receptor co-operativity in which binding to the targeted (Ram-1) receptor triggers conformational rearrangements of the envelope that lead to complete unmasking of the hidden Rec-1-binding domain, thereby facilitating its interaction with the viral (Rec-1) receptor which leads to optimal fusion triggering. PMID:9135138

  2. 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

  3. Foamy-like endogenous retroviruses are extensive and abundant in teleosts

    PubMed Central

    Ruboyianes, Ryan; Worobey, Michael

    2016-01-01

    Recent discoveries indicate that the foamy virus (FV) (Spumavirus) ancestor may have been among the first retroviruses to appear during the evolution of vertebrates, demonstrated by foamy endogenous retroviruses present within deeply divergent hosts including mammals, coelacanth, and ray-finned fish. If they indeed existed in ancient marine environments hundreds of millions of years ago, significant undiscovered diversity of foamy-like endogenous retroviruses might be present in fish genomes. By screening published genomes and by applying PCR-based assays of preserved tissues, we discovered 23 novel foamy-like elements in teleost hosts. These viruses form a robust, reciprocally monophyletic sister clade with sarcopterygian host FV, with class III mammal endogenous retroviruses being the sister group to both clades. Some of these foamy-like retroviruses have larger genomes than any known retrovirus, exogenous or endogenous, due to unusually long gag-like genes and numerous accessory genes. The presence of genetic features conserved between mammalian FV and these novel retroviruses attests to a foamy-like replication biology conserved for hundreds of millions of years. We estimate that some of these viruses integrated recently into host genomes; exogenous forms of these viruses may still circulate. PMID:28058112

  4. 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.

  5. Transcriptionally active genome regions are preferred targets for retrovirus integration.

    PubMed Central

    Scherdin, U; Rhodes, K; Breindl, M

    1990-01-01

    We have analyzed the transcriptional activity of cellular target sequences for Moloney murine leukemia virus integration in mouse fibroblasts. At least five of the nine random, unselected integration target sequences studied showed direct evidence for transcriptional activity by hybridization to nuclear run-on transcripts prepared from uninfected cells. At least four of the sequences contained multiple recognition sites for several restriction enzymes that cut preferentially in CpG-rich islands, indicating integration into 5' or 3' ends or flanking regions of genes. Assuming that only a minor fraction (less than 20%) of the genome is transcribed in mammalian cells, we calculated the probability that this association of retroviral integration sites with transcribed sequences is due to chance to be very low (1.6 x 10(-2]. Thus, our results strongly suggest that transcriptionally active genome regions are preferred targets for retrovirus integration. Images PMID:2296087

  6. The ability of multimerized cyclophilin A to restrict retrovirus infection

    SciTech Connect

    Javanbakht, Hassan; Diaz-Griffero, Felipe; Yuan Wen; Yeung, Darwin F.; Li Xing; Song Byeongwoon; Sodroski, Joseph

    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 and 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.

  7. 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.

  8. Cellular restriction of retrovirus particle-mediated mRNA transfer.

    PubMed

    Galla, Melanie; Schambach, Axel; Towers, Greg J; Baum, Christopher

    2008-03-01

    Analyzing cellular restriction mechanisms provides insight into viral replication strategies, identifies targets for antiviral drug design, and is crucial for the development of novel tools for experimental or therapeutic delivery of genetic information. We have previously shown that retroviral vector mutants that are unable to initiate reverse transcription mediate a transient expression of any sequence which replaces the gag-pol transcription unit, a process we call retrovirus particle-mediated mRNA transfer (RMT). Here, we further examined the mechanism of RMT by testing its sensitivity to cellular restriction factors and short hairpin RNAs (shRNAs). We found that both human TRIM5alpha and, to a lesser extent, Fv1 effectively restrict RMT if the RNA is delivered by a restriction-sensitive capsid. While TRIM5alpha restriction of RMT led to reduced levels of retroviral mRNA in target cells, restriction by Fv1 did not. Treatment with the proteasome inhibitor MG132 partially relieved TRIM5alpha-mediated restriction of RMT. Finally, cells expressing shRNAs specifically targeting the retroviral mRNA inhibited RMT particles, but not reverse-transcribing particles. Retroviral mRNA may thus serve as a translation template if not used as a template for reverse transcription. Our data imply that retroviral nucleic acids become accessible to host factors, including ribosomes, as a result of particle remodeling during cytoplasmic trafficking.

  9. 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.

  10. The role of human endogenous retroviruses in brain development and function.

    PubMed

    Mortelmans, Kristien; Wang-Johanning, Feng; Johanning, Gary L

    2016-01-01

    Endogenous retroviral sequences are spread throughout the genome of all humans, and make up about 8% of the genome. Despite their prevalence, the function of human endogenous retroviruses (HERVs) in humans is largely unknown. In this review we focus on the brain, and evaluate studies in animal models that address mechanisms of endogenous retrovirus activation in the brain and central nervous system (CNS). One such study in mice found that TRIM28, a protein critical for mouse early development, regulates transcription and silencing of endogenous retroviruses in neural progenitor cells. Another intriguing finding in human brain cells and mouse models was that endogenous retrovirus HERV-K appears to be protective against neurotoxins. We also report on studies that associate HERVs with human diseases of the brain and CNS. There is little doubt of an association between HERVs and a number of CNS diseases. However, a cause and effect relationship between HERVs and these diseases has not yet been established.

  11. Interleukin-7 retroviruses transform pre-B cells by an autocrine mechanism not evident in Abelson murine.

    PubMed Central

    Overell, R W; Clark, L; Lynch, D; Jerzy, R; Schmierer, A; Weisser, K E; Namen, A E; Goodwin, R G

    1991-01-01

    In this study, we have constructed retroviral vectors expressing the interleukin-7 (IL-7) cDNA and have used infection with these retroviruses to express this cytokine endogenously in an IL-7-dependent pre-B-cell line. Infection with IL-7 retroviruses, but not with a control retrovirus, resulted in the conversion of the cells to IL-7 independence. The frequency at which this occurred, together with data on vector expression levels, indicated that secondary events were required for factor independence in this system. Southern analysis showed that the IL-7-dependent clones harbored unrearranged copies of the vector proviruses. The factor-independent cells produced variable quantities of IL-7 as measured by an IL-7-specific bioassay, and their proliferation could be substantially inhibited by a neutralizing antibody directed against IL-7, indicating that a classical autocrine-mechanism was responsible for their transformation. These IL-7-independent cells were tumorigenic, in contrast to the parental IL-7-dependent cells or those infected with a control vector. These results showed that IL-7 could participate in the malignant transformation of pre-B cells. However, neither of two Abelson murine leukemia virus (A-MuLV)-transformed pre-B-cell lines expressed detectable IL-7 mRNA, at a level of sensitivity corresponding to less than one molecule of mRNA per cell. Moreover, the proliferation of the A-MuLV transformants was unaffected by addition of the IL-7 antisera under conditions in which parallel experiments with IL-7 virus-infected cells resulted in greater than 70% growth inhibition. Thus, transformation of pre-B cells by A-MuLV was not associated with a demonstrable autocrine loop of IL-7 synthesis. These results show that IL-7 can participate in the malignant transformation of pre-B cells and suggest studies aimed at assessing the role of autocrine production of IL-7 in the generation of human leukemias and lymphomas. Images PMID:1996110

  12. Envelope-binding domain in the cationic amino acid transporter determines the host range of ecotropic murine retroviruses.

    PubMed Central

    Albritton, L M; Kim, J W; Tseng, L; Cunningham, J M

    1993-01-01

    Infection of rodent cells by ecotropic type C retroviruses requires the expression of a cationic amino acid transporter composed of multiple membrane-spanning domains. By exchanging portions of cDNAs encoding the permissive mouse and nonpermissive human transporters and examining their abilities to specify virus infection upon expression in human 293 cells, we have identified the amino acid residues in the extracellular loop connecting the fifth and sixth membrane-spanning segments of the mouse transporter that are required for both envelope gp70 binding and infection. These findings strongly suggest that the role of the mouse transporter in determining infection is to provide an envelope-binding site. This role is analogous to those of host membrane proteins composed of a single membrane-spanning domain that serve as binding proteins or receptors for other enveloped viruses such as human immunodeficiency virus, Epstein-Barr virus, and murine and human coronaviruses. PMID:8445722

  13. Gene expression during normal and malignant differentiation

    SciTech Connect

    Andersson, L.C.; Gahmberg, C.G.; Ekblom, P.

    1985-01-01

    This book contains 18 selections. Some of the titles are: Exploring Carcinogenesis with Retroviral and Cellular Oncogenes; Retroviruses, Oncogenes and Evolution; HTLV and Human Neoplasi; Modes of Activation of cMyc Oncogene in B and T Lymphoid Tumors; The Structure and Function of the Epidermal Growth Factor Receptor: Its Relationship to the Protein Product of the V-ERB-B Oncogene; and Expression of Human Retrovirus Genes in Normal and Neoplastic Epithelial Cells.

  14. Purification and characterization of retrovirus vector particles by rate zonal ultracentrifugation.

    PubMed

    Segura, María de las Mercedes; Garnier, Alain; Kamen, Amine

    2006-04-01

    Sucrose equilibrium density ultracentrifugation remains the most widely used technique for retrovirus purification. However, purified virus preparations obtained by this routine method usually contain considerable amounts of contaminating cell membrane vesicles. In addition, sucrose solutions are highly viscous and hyperosmotic which jeopardizes the integrity and functionality of the retrovirus particle. In order to overcome these limitations, an alternative purification technique using rate zonal ultracentrifugation and iodixanol as gradient medium was developed. Recombinant retrovirus particles were produced by 293-GPG packaging cells grown in suspension in the presence of 10% FBS. Concentrated supernatants were purified by rate zonal sedimentation on a 10-30% continuous iodixanol gradient. Virus particles were recovered intact and active from the central fractions of the gradient. By using this strategy, high levels of purification were achieved, with no evident contamination with cell membrane vesicles as indicated by subtilisin treatment studies. The level of purity of the retrovirus preparation is over 95% as shown by SDS-PAGE analysis and size-exclusion chromatography. Purified particles appear homogenous in size and morphology according to negative stain electron microscopy. In addition, large amounts of defective retrovirus particles produced by 293-GPG packaging cells can be separated from functional retrovirus particles using this purification strategy.

  15. Endogenous retrovirus drives hitherto unknown proapoptotic p63 isoforms in the male germ line of humans and great apes.

    PubMed

    Beyer, Ulrike; Moll-Rocek, Julian; Moll, Ute M; Dobbelstein, Matthias

    2011-03-01

    TAp63, but not its homolog p53, eliminates oocytes that suffered DNA damage. An equivalent gene for guarding the male germ line is currently not known. Here we identify hitherto unknown human p63 transcripts with unique 5'-ends derived from incorporated exons upstream of the currently mapped TP63 gene. These unique p63 transcripts are highly and specifically expressed in testis. Their most upstream region corresponds to a LTR of the human endogenous retrovirus 9 (ERV9). The insertion of this LTR upstream of the TP63 locus occurred only recently in evolution and is unique to humans and great apes (Hominidae). A corresponding p63 protein is the sole p63 species in healthy human testis, and is strongly expressed in spermatogenic precursors but not in mature spermatozoa. In response to DNA damage, this human male germ-cell-encoded TAp63 protein (designated GTAp63) is activated by caspase cleavage near its carboxyterminal domain and induces apoptosis. Human testicular cancer tissues and cell lines largely lost p63 expression. However, pharmacological inhibition of histone deacetylases completely restores p63 expression in testicular cancer cells (>3,000-fold increase). Our data support a model whereby testis-specific GTAp63 protects the genomic integrity of the male germ line and acts as a tumor suppressor. In Hominidae, this guardian function was greatly enhanced by integration of an endogenous retrovirus upstream of the TP63 locus that occurred 15 million years ago. By providing increased germ-line stability, this event may have contributed to the evolution of hominids and enabled their long reproductive periods.

  16. HIV-1 Interacts with Human Endogenous Retrovirus K (HML-2) Envelopes Derived from Human Primary Lymphocytes

    PubMed Central

    Brinzevich, Daria; Young, George R.; Sebra, Robert; Ayllon, Juan; Maio, Susan M.; Deikus, Gintaras; Chen, Benjamin K.; Fernandez-Sesma, Ana; Simon, Viviana

    2014-01-01

    ABSTRACT Human endogenous retroviruses (HERVs) are viruses that have colonized the germ line and spread through vertical passage. Only the more recently acquired HERVs, such as the HERV-K (HML-2) group, maintain coding open reading frames. Expression of HERV-Ks has been linked to different pathological conditions, including HIV infection, but our knowledge on which specific HERV-Ks are expressed in primary lymphocytes currently is very limited. To identify the most expressed HERV-Ks in an unbiased manner, we analyzed their expression patterns in peripheral blood lymphocytes using Pacific Biosciences (PacBio) single-molecule real-time (SMRT) sequencing. We observe that three HERV-Ks (KII, K102, and K18) constitute over 90% of the total HERV-K expression in primary human lymphocytes of five different donors. We also show experimentally that two of these HERV-K env sequences (K18 and K102) retain their ability to produce full-length and posttranslationally processed envelope proteins in cell culture. We show that HERV-K18 Env can be incorporated into HIV-1 but not simian immunodeficiency virus (SIV) particles. Moreover, HERV-K18 Env incorporation into HIV-1 virions is dependent on HIV-1 matrix. Taken together, we generated high-resolution HERV-K expression profiles specific for activated human lymphocytes. We found that one of the most abundantly expressed HERV-K envelopes not only makes a full-length protein but also specifically interacts with HIV-1. Our findings raise the possibility that these endogenous retroviral Env proteins could directly influence HIV-1 replication. IMPORTANCE Here, we report the HERV-K expression profile of primary lymphocytes from 5 different healthy donors. We used a novel deep-sequencing technology (PacBio SMRT) that produces the long reads necessary to discriminate the complexity of HERV-K expression. We find that primary lymphocytes express up to 32 different HERV-K envelopes, and that at least two of the most expressed Env proteins

  17. Feline immunodeficiency virus and retrovirus-mediated adventitial ex vivo gene transfer to rabbit carotid artery using autologous vascular smooth muscle cells.

    PubMed

    Kankkonen, Hanna M; Turunen, Mikko P; Hiltunen, Mikko O; Lehtolainen, Pauliina; Koponen, Jonna; Leppänen, Pia; Turunen, Anna-Mari; Ylä-Herttuala, Seppo

    2004-03-01

    We have developed an ex vivo gene transfer technique to rabbit arterial wall using autologous smooth muscle cells (SMCs). SMCs were harvested from rabbit ear artery, transduced in vitro with vesicular stomatitis virus G-glycoprotein pseudotyped retrovirus or feline immunodeficiency virus (FIV) and returned to the adventitial surface of the carotid artery using a periadventitial silicone collar or collagen sheet placed around the artery. Beta-galactosidase (lacZ) and human apolipoprotein E3 (apoE3) cDNAs were used as transgenes. After retrovirus-mediated gene transfer of lacZ the selected cells implanted with high efficiency and expressed lacZ marker gene at a very high level 7 and 14 days after the operation. The level of lacZ expression decreased thereafter but was still detectable 12 weeks after the gene transfer, and was exclusively localized to the site of cell implantation inside the collar. Utilizing FIV vector expressing apoE3, low levels of apoE were measured from serum collected from a low-density lipoprotein receptor deficient Watanabe heritable hyperlipidemic rabbits 1 month after the gene transfer. The physiological effect of apoE expression was detected as transiently elevated serum cholesterol levels. The results indicate that the model can be used for high efficiency local gene transfer in arteries, e.g. during vascular surgery. The model is also valuable for studying expression, stability and safety of new gene transfer vectors and their expression products in vivo.

  18. 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

  19. Endogenous retroviruses function as species-specific enhancer elements in the placenta

    PubMed Central

    Chuong, Edward B.; Rumi, M. A. Karim; Soares, Michael J.; Baker, Julie C.

    2013-01-01

    The mammalian placenta is remarkably distinct between species, suggesting a history of rapid evolutionary diversification1. To gain insight into the molecular drivers of placental evolution, we compared biochemically predicted enhancers between mouse and rat trophoblast stem cells (TSCs) and find that species-specific enhancers are highly enriched for endogenous retroviruses (ERVs) on a genome-wide level. One of these ERV families, RLTR13D5, contributes hundreds of mouse-specific H3K4me1/H3K27ac-defined enhancers that functionally bind Cdx2, Eomes, and Elf5 - core factors that define the TSC regulatory network. Furthermore, we demonstrate that RLTR13D5 is capable of driving gene expression in rat placental cells. Comparison with other tissues revealed that species-specific ERV enhancer activity is generally restricted to hypomethylated tissues, suggesting that tissues permissive to ERV activity gain access to an otherwise silenced source of regulatory variation. Overall, our results implicate ERV enhancer cooption as a mechanism underlying the striking evolutionary diversification of placental development. PMID:23396136

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

    SciTech Connect

    Schneider, Irene C.; Eckhardt, Manon; Brynza, Julia; Collins, Mary K.; Cichutek, Klaus; Buchholz, Christian J.

    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 contrast 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.

  1. Retrovirus-induced insertional mutagenesis: mechanism of collagen mutation in Mov13 mice.

    PubMed Central

    Barker, D D; Wu, H; Hartung, S; Breindl, M; Jaenisch, R

    1991-01-01

    The Mov13 mouse strain carries a mutation in the alpha 1(I) procollagen gene which is due to the insertion of a Moloney murine leukemia provirus into the first intron. This insertion results in the de novo methylation of the provirus and flanking DNA, the alteration of chromatin structure, and the transcriptional inactivity of the collagen promoter. To address the mechanism of mutagenesis, we reintroduced a cloned and therefore demethylated version of the Mov13 mutant allele into mouse fibroblasts. The transfected gene was not transcribed, indicating that the transcriptional defect was not due to the hypermethylation. Rather, this result strongly suggests that the mutation is due to the displacement or disruption of cis-acting regulatory DNA sequences within the first intron. We also constructed a Mov13 variant allele containing a single long terminal repeat instead of the whole provirus. This construct also failed to express mRNA, indicating that the Mov13 mutation does not revert by provirus excision as has been observed for other retrovirus-induced mutations. Images PMID:1922037

  2. Comparative and functional studies of Drosophila species invasion by the gypsy endogenous retrovirus.

    PubMed Central

    Mejlumian, Lucine; Pélisson, Alain; Bucheton, Alain; Terzian, Christophe

    2002-01-01

    Gypsy is an endogenous retrovirus of Drosophila melanogaster. Phylogenetic studies suggest that occasional horizontal transfer events of gypsy occur between Drosophila species. gypsy possesses infective properties associated with the products of the envelope gene that might be at the origin of these interspecies transfers. We report here the existence of DNA sequences putatively encoding full-length Env proteins in the genomes of Drosophila species other than D. melanogaster, suggesting that potentially infective gypsy copies able to spread between sexually isolated species can occur. The ability of gypsy to invade the genome of a new species is conditioned by its capacity to be expressed in the naive genome. The genetic basis for the regulation of gypsy activity in D. melanogaster is now well known, and it has been assigned to an X-linked gene called flamenco. We established an experimental simulation of the invasion of the D. melanogaster genome by gypsy elements derived from other Drosophila species, which demonstrates that these non- D. melanogaster gypsy elements escape the repression exerted by the D. melanogaster flamenco gene. PMID:11805056

  3. Cooccurrences of Putative Endogenous Retrovirus-Associated Diseases

    PubMed Central

    Emmer, Alexander; Kornhuber, Malte E.

    2017-01-01

    At least 8% of the human genome is composed of endogenous retrovirus (ERV) sequences. ERVs play a role in placental morphogenesis and can sometimes protect the host against exogenous viruses. On the other hand, ERV reactivation has been found to be associated with different diseases, for example, multiple sclerosis (MS), schizophrenia, type 1 diabetes mellitus (T1D), or amyotrophic lateral sclerosis (ALS). Little is known about the cooccurrence of these diseases. If all these diseases are caused by ERV, antiretroviral therapy should perhaps also show some effects in the other diseases. Here, we summarize literature demonstrating that some ERV-associated diseases seem to appear together more often than expected, for example, MS and ALS, MS and T1D, MS and schizophrenia, or ALS and T1D. In contrast, some ERV-associated diseases seem to appear together less frequently than expected, for example, schizophrenia and T1D. Besides, some reports demonstrate amelioration of MS, ALS, or schizophrenia under antiretroviral therapy in human immunodeficiency virus-infected patients. If such results could be confirmed in larger studies, alternative therapy strategies for ERV-associated diseases like MS and schizophrenia might be possible. PMID:28326328

  4. 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.

  5. Isolation of koala retroviruses from koalas in Japan.

    PubMed

    Miyazawa, Takayuki; Shojima, Takayuki; Yoshikawa, Rokusuke; Ohata, Takuji

    2011-01-01

    Koala retrovirus (KoRV) is considered to be associated with leukemia, lymphoma and immunodeficiency-like diseases in koalas. We therefore conducted a pilot study of KoRV infection in five Queensland koalas in Kobe Municipal Oji Zoo. By polymerase chain reaction to detect partial env and pol genes of KoRV in genomic DNA isolated from whole blood and feces, all five koalas were found to be positive for KoRV proviruses. We succeeded in culturing koala lymphocytes from less than 1 ml blood for over 14 days in the presence of recombinant human interleukin-2. By coculturing the lymphocytes with human embryonic kidney (HEK) 293T cells, we isolated KoRVs from all five koalas. We designated these isolates as strains OJ-1 to OJ-5. By electron microscopy, we observed C-type retroviral particles in HEK 293T cells chronically infected with KoRV strain OJ-4. This is the first report on the isolation of KoRV from koalas in a Japanese zoo.

  6. Flamenco, a gene controlling the gypsy retrovirus of drosophila melanogaster

    SciTech Connect

    Prud`homme, N.; Gans, M.; Masson, M.; Terzian, C.; Bucheton, A.

    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, can 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.

  7. Dynamic regulation of human endogenous retroviruses mediates factor-induced reprogramming and differentiation potential

    PubMed Central

    Ohnuki, Mari; Tanabe, Koji; Sutou, Kenta; Teramoto, Ito; Sawamura, Yuka; Narita, Megumi; Nakamura, Michiko; Tokunaga, Yumie; Nakamura, Masahiro; Watanabe, Akira; Yamanaka, Shinya; Takahashi, Kazutoshi

    2014-01-01

    Pluripotency can be induced in somatic cells by overexpressing transcription factors, including POU class 5 homeobox 1 (OCT3/4), sex determining region Y-box 2 (SOX2), Krüppel-like factor 4 (KLF4), and myelocytomatosis oncogene (c-MYC). However, some induced pluripotent stem cells (iPSCs) exhibit defective differentiation and inappropriate maintenance of pluripotency features. Here we show that dynamic regulation of human endogenous retroviruses (HERVs) is important in the reprogramming process toward iPSCs, and in re-establishment of differentiation potential. During reprogramming, OCT3/4, SOX2, and KLF4 transiently hyperactivated LTR7s—the long-terminal repeats of HERV type-H (HERV-H)—to levels much higher than in embryonic stem cells by direct occupation of LTR7 sites genome-wide. Knocking down LTR7s or long intergenic non-protein coding RNA, regulator of reprogramming (lincRNA-RoR), a HERV-H–driven long noncoding RNA, early in reprogramming markedly reduced the efficiency of iPSC generation. KLF4 and LTR7 expression decreased to levels comparable with embryonic stem cells once reprogramming was complete, but failure to resuppress KLF4 and LTR7s resulted in defective differentiation. We also observed defective differentiation and LTR7 activation when iPSCs had forced expression of KLF4. However, when aberrantly expressed KLF4 or LTR7s were suppressed in defective iPSCs, normal differentiation was restored. Thus, a major mechanism by which OCT3/4, SOX2, and KLF4 promote human iPSC generation and reestablish potential for differentiation is by dynamically regulating HERV-H LTR7s. PMID:25097266

  8. 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.

  9. Retrovirus transduction: Segregation of the viral transforming function and the Herpes Simplex virus tk gene in infectious friend spleen focus-forming virus thymidine kinase vectors

    SciTech Connect

    Joyner, A.L.; Bernstein, A.

    1983-12-01

    A series of deletions and insertions utilizing the herpesvirus thymidine kinase gene (tk) were constructed in the murine retrovirus Friend spleen focus-forming virus (SFFV). In all cases, the coding region for the SFFV-specific glycoprotein (gp55), which is implicated in erythroleukemic transformation, was left intact. These SFFV-TK and SFFV deletion vectors were analyzed for expression of tk and gp55 after DNA-mediated gene transfer. In addition, virus rescued by cotranfection of these vectors with Moloney murine leukemia virus was analyzed for infectious TK-transducing virus, gp55 expression, and erythroleukemia-inducing ability. The experiments demonstrated that deletions or insertions within the intron for the gp55 env gene can interfere with expression of gp55 after both DNA-mediated gene transfer and virus infection. In contrast, the gene transfer efficiency of the tk gene was unaffected in the SFFV-TK vectors, and high-titer infectious TK virus could be recovered. Revertant viruses capable of inducing erythroleukemia and expressing gp55 were generated after cotranfection of the SFFV-TK vectors with murine leukemia virus. The revertant viruses lost both tk sequences and the ability to transduce TK/sup -/ fibroblasts to a TK/sup +/ phenotype. These experiments demonstrate that segregation of the TK and erythroleukemia functions can occur in retrovirus vectors which initially carry both markers.

  10. Retrovirus transduction: segregation of the viral transforming function and the herpes simplex virus tk gene in infectious Friend spleen focus-forming virus thymidine kinase vectors.

    PubMed Central

    Joyner, A L; Bernstein, A

    1983-01-01

    A series of deletions and insertions utilizing the herpesvirus thymidine kinase gene (tk) were constructed in the murine retrovirus Friend spleen focus-forming virus (SFFV). In all cases, the coding region for the SFFV-specific glycoprotein (gp55), which is implicated in erythroleukemic transformation, was left intact. These SFFV-TK and SFFV deletion vectors were analyzed for expression of tk and gp55 after DNA-mediated gene transfer. In addition, virus rescued by cotransfection of these vectors with Moloney murine leukemia virus was analyzed for infectious TK-transducing virus, gp55 expression, and erythroleukemia-inducing ability. The experiments demonstrated that deletions or insertions within the intron for the gp55 env gene can interfere with expression of gp55 after both DNA-mediated gene transfer and virus infection. In contrast, the gene transfer efficiency of the tk gene was unaffected in the SFFV-TK vectors, and high-titer infectious TK virus could be recovered. Revertant viruses capable of inducing erythroleukemia and expressing gp55 were generated after cotransfection of the SFFV-TK vectors with murine leukemia virus. The revertant viruses lost both tk sequences and the ability to transduce TK- fibroblasts to a TK+ phenotype. These experiments demonstrate that segregation of the TK and erythroleukemia functions can occur in retrovirus vectors which initially carry both markers. Images PMID:6318088

  11. Colostrum and milk can transmit jaagsiekte retrovirus to lambs.

    PubMed

    Grego, Elena; De Meneghi, Daniele; Alvarez, Vega; Benito, Alfredo A; Minguijón, Esmeralda; Ortín, Aurora; Mattoni, Mario; Moreno, Bernardino; Pérez de Villarreal, Maider; Alberti, Alberto; Capucchio, Maria Teresa; Caporale, Marco; Juste, Ramón; Rosati, Sergio; De las Heras, Marcelo

    2008-08-25

    Ovine pulmonary adenocarcinoma (OPA) is a contagious disease caused by jaagsiekte sheep retrovirus (JSRV). In the three studies performed, we have obtained data of the importance of colostrum/milk (C/M) in the transmission of JSRV. In the first study, a group of sheep from a flock with a long history of OPA, samples from colostrum and peripheral blood leucocytes (PBLs) were collected. Two specific PCRs (U3-LTR and env of the JSRV) were carried out. Using U3PCR 8/34 sheep were positive in colostrum whereas with envPCR 7/34 were positive. From these animals only one was positive with U3PCR in the PBLs. Evidence of the transmission of JSRV infection by C/M was obtained in two more separate studies. In the second study, PBLs from five lambs from JSRV+ ewes and two from JSRV-ewes were tested by the U3PCR. They were fed C/M by their mothers during 3 months and slaughtered 7 months after birth. Three out of five lambs from the JSRV+ sheep become PBL positive at 3-4 months old and the other two were also positive at 4-6 months of age. One lamb of the JSRV-sheep became also PBL positive at an age of 3 months. In the third study, a group of lambs from JSRV negative mothers were fed with C/M from JSRV+ sheep and housed in separate unit. For comparison, another group of the same origin and maintained in another different unit, were fed with C/M containing a JSRV virus preparation. All lambs were blood sampled monthly and JSRV infection was detected as early as 15 days and several times onwards in both groups. Control groups fed with C/M from JSRV free flock and JSRV blood test negative sheep were always negative. Together these results indicate that suckling is an important natural transmission route for JSRV.

  12. Evolutionary history of bovine endogenous retroviruses in the Bovidae family

    PubMed Central

    2013-01-01

    Background Endogenous retroviruses (ERVs) are genomic elements of retroviral origin that are present in the genomes of almost all vertebrates. In cattle, more than 13,000 elements related to ERVs have been detected, and based on the pol gene, 24 families or groups of bovine ERVs have been described. However, information about ERVs in other bovids and the presence of families of related bovine ERVs in different species of the Bovidae family is scarce. Results The 24 families of bovine ERVs previously detected in cattle (Bos taurus) were also detected in zebus (Bos indicus) and yaks (Bos grunniens). In addition, six new families, named BoERV25 to BoERV30, were detected in the three Bos species. Five more ruminant species were screened for related ERVs: 26 families were detected in these species, but four families (BoERV24, BoERV26, BoERV28 and BoERV29) were specific to cattle, zebus, yaks and buffalo. An analysis of the homology of the ERVs of cattle, zebus and yaks revealed that the level of LTR divergence was similar between ERVs from cattle and zebus but was less similar between with ERVs from cattle and yaks. In addition, purifying selection was detected in the genes and retroviral regions of clusters of ERVs of cattle, zebus and yaks. Conclusions In this work, the 24 ERV families previously identified in cattle were also found in two other species in the Bos genus. In addition, six new bovine ERV families were detected. Based on LTR divergence, the most recently inserted families are from Class II. The divergence of the LTR, used as an indirect estimate of the ERV insertion time, seemed to be influenced by the differences in genome evolution since the divergence of the species. In addition, purifying selection could be acting on clusters of ERVs from different species. PMID:24256121

  13. Evolution of broad host range in retroviruses leads to cell death mediated by highly cytopathic variants.

    PubMed

    Rainey, G Jonah A; Coffin, John M

    2006-01-01

    The ability of many retroviruses to cause disease can be correlated to their cytopathic effect (CPE) in tissue culture characterized by an acute period of cell death and viral DNA accumulation. Here, we show that mutants of a subgroup B avian retrovirus (Alpharetrovirus) cause a very dramatic CPE in certain susceptible avian cells that is coincident with elevated levels of apoptosis, as measured by nuclear morphology, and persistent viral DNA accumulation. These mutants also have a broadly extended host range that includes rodent, cat, dog, monkey, and human cells (31). Previously, we have shown that the mutants exhibit diminished resistance to superinfection. The results presented here have important implications for the process of evolution of retroviruses to use distinct cellular receptors.

  14. Endogenous Retroviruses in Fish Genomes: From Relics of Past Infections to Evolutionary Innovations?

    PubMed Central

    Naville, Magali; Volff, Jean-Nicolas

    2016-01-01

    The increasing availability of fish genome sequences has allowed to gain new insights into the diversity and host distribution of retroviruses in fish and other vertebrates. This distribution can be assessed through the identification and analysis of endogenous retroviruses, which are proviral remnants of past infections integrated in genomes. Retroviral sequences are probably important for evolution through their ability to induce rearrangements and to contribute regulatory and coding sequences; they may also protect their host against new infections. We argue that the current mass of genome sequences will soon strongly improve our understanding of retrovirus diversity and evolution in aquatic animals, with the identification of new/re-emerging elements and host resistance genes that restrict their infectivity. PMID:27555838

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

    PubMed Central

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

    2013-01-01

    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. PMID:23938752

  16. Cross-Species Transmission and Differential Fate of an Endogenous Retrovirus in Three Mammal Lineages

    PubMed Central

    Zhuo, Xiaoyu; Feschotte, Cédric

    2015-01-01

    Endogenous retroviruses (ERVs) arise from retroviruses chromosomally integrated in the host germline. ERVs are common in vertebrate genomes and provide a valuable fossil record of past retroviral infections to investigate the biology and evolution of retroviruses over a deep time scale, including cross-species transmission events. Here we took advantage of a catalog of ERVs we recently produced for the bat Myotis lucifugus to seek evidence for infiltration of these retroviruses in other mammalian species (>100) currently represented in the genome sequence database. We provide multiple lines of evidence for the cross-ordinal transmission of a gammaretrovirus endogenized independently in the lineages of vespertilionid bats, felid cats and pangolin ~13–25 million years ago. Following its initial introduction, the ERV amplified extensively in parallel in both bat and cat lineages, generating hundreds of species-specific insertions throughout evolution. However, despite being derived from the same viral species, phylogenetic and selection analyses suggest that the ERV experienced different amplification dynamics in the two mammalian lineages. In the cat lineage, the ERV appears to have expanded primarily by retrotransposition of a single proviral progenitor that lost infectious capacity shortly after endogenization. In the bat lineage, the ERV followed a more complex path of germline invasion characterized by both retrotransposition and multiple infection events. The results also suggest that some of the bat ERVs have maintained infectious capacity for extended period of time and may be still infectious today. This study provides one of the most rigorously documented cases of cross-ordinal transmission of a mammalian retrovirus. It also illustrates how the same retrovirus species has transitioned multiple times from an infectious pathogen to a genomic parasite (i.e. retrotransposon), yet experiencing different invasion dynamics in different mammalian hosts. PMID

  17. Cross-Species Transmission and Differential Fate of an Endogenous Retrovirus in Three Mammal Lineages.

    PubMed

    Zhuo, Xiaoyu; Feschotte, Cédric

    2015-01-01

    Endogenous retroviruses (ERVs) arise from retroviruses chromosomally integrated in the host germline. ERVs are common in vertebrate genomes and provide a valuable fossil record of past retroviral infections to investigate the biology and evolution of retroviruses over a deep time scale, including cross-species transmission events. Here we took advantage of a catalog of ERVs we recently produced for the bat Myotis lucifugus to seek evidence for infiltration of these retroviruses in other mammalian species (>100) currently represented in the genome sequence database. We provide multiple lines of evidence for the cross-ordinal transmission of a gammaretrovirus endogenized independently in the lineages of vespertilionid bats, felid cats and pangolin ~13-25 million years ago. Following its initial introduction, the ERV amplified extensively in parallel in both bat and cat lineages, generating hundreds of species-specific insertions throughout evolution. However, despite being derived from the same viral species, phylogenetic and selection analyses suggest that the ERV experienced different amplification dynamics in the two mammalian lineages. In the cat lineage, the ERV appears to have expanded primarily by retrotransposition of a single proviral progenitor that lost infectious capacity shortly after endogenization. In the bat lineage, the ERV followed a more complex path of germline invasion characterized by both retrotransposition and multiple infection events. The results also suggest that some of the bat ERVs have maintained infectious capacity for extended period of time and may be still infectious today. This study provides one of the most rigorously documented cases of cross-ordinal transmission of a mammalian retrovirus. It also illustrates how the same retrovirus species has transitioned multiple times from an infectious pathogen to a genomic parasite (i.e. retrotransposon), yet experiencing different invasion dynamics in different mammalian hosts.

  18. Molecular structure of a gypsy element of Drosophila subobscura (gypsyDs) constituting a degenerate form of insect retroviruses.

    PubMed Central

    Alberola, T M; de Frutos, R

    1996-01-01

    We have determined the nucleotide sequence of a 7.5 kb full-size gypsy element from Drosophila subobscura strain H-271. Comparative analyses were carried out on the sequence and molecular structure of gypsy elements of D.subobscura (gypsyDs), D.melanogaster (gypsyDm) and D.virilis (gypsyDv). The three elements show a structure that maintains a common mechanism of expression. ORF1 and ORF2 show typical motifs of gag and pol genes respectively in the three gypsy elements and could encode functional proteins necessary for intracellular expansion. In the three ORF1 proteins an arginine-rich region was found which could constitute a RNA binding motif. The main differences among the gypsy elements are found in ORF3 (env-like gene); gypsyDm encodes functional env proteins, whereas gypsyDs and gypsyDv ORF3s lack some motifs essential for functionality of this protein. On the basis of these results, while gypsyDm is the first insect retrovirus described, gypsyDs and gypsyDv could constitute degenerate forms of these retroviruses. In this context, we have found some evidence that gypsyDm could have recently infected some D.subobscura strains. Comparative analyses of divergence and phylogenetic relationships of gypsy elements indicate that the gypsy elements belonging to species of different subgenera (gypsyDs and gypsyDv) are closer than gypsy elements of species belonging to the same subgenus (gypsyDs and gypsyDm). These data are congruent with horizontal transfer of gypsy elements among different Drosophila spp. PMID:8600460

  19. Inflammatory response of endothelial cells to a human endogenous retrovirus associated with multiple sclerosis is mediated by TLR4.

    PubMed

    Duperray, Alain; Barbe, Delphin; Raguenez, Gilda; Weksler, Babette B; Romero, Ignacio A; Couraud, Pierre-Olivier; Perron, Hervé; Marche, Patrice N

    2015-11-01

    The MSRV (multiple sclerosis-associated retrovirus) belongs to the human endogenous retrovirus HERV-W family. The envelope protein originating from the MSRV has been found in most patients with multiple sclerosis (MS). This protein (Env-ms) has pro-inflammatory properties for several types of immune cells and could therefore play a role in MS pathogenesis by promoting the leukocyte diapedesis observed in the central nervous system of patients. Our study aims to analyze the effects of Env-ms on the blood-brain barrier (BBB) at a molecular and functional level. We demonstrate that the recombinant MSRV envelope is able to stimulate several inflammatory parameters in a human BBB in vitro model, the HCMEC/D3 brain endothelial cell line. Indeed, Env-ms induces over-expression of ICAM-1, a major mediator of leukocyte adhesion to endothelial cells, in a dose-dependent manner as well as a strong dose-dependent production of the pro-inflammatory cytokines IL-6 and IL-8. Furthermore, using a silencing approach with siRNAs, we show that Env-ms is recognized via the Toll-like receptor 4 receptor, a pattern recognition receptor of innate immunity present on endothelial cells. We also show, using functional assays, that treatment of brain endothelial cells with Env-ms significantly stimulated the adhesion and the transmigration of activated immune cells through a monolayer of endothelial cells. These findings support the hypothesis that MSRV could be involved in the pathogenesis of MS disease or at least in maintenance of inflammatory conditions, thus fueling the auto-immune disorder. MSRV could also play a role in other chronic inflammatory diseases.

  20. 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.

  1. 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

  2. Mechanism of inhibition of retrovirus release from cells by interferon-induced gene ISG15.

    PubMed

    Kuang, Zhizhou; Seo, Eun Joo; Leis, Jonathan

    2011-07-01

    Budding of retroviruses from cell membranes requires ubiquitination of Gag and recruitment of cellular proteins involved in endosome sorting, including endosome sorting complex required for transport III (ESCRT-III) protein complex and vacuolar protein sorting 4 (VPS4) and its ATPase. In response to infection, a cellular mechanism has evolved that blocks virus replication early and late in the budding process through expression of interferon-stimulated gene 15 (ISG15), a dimer homologue of ubiquitin. Interferon treatment of DF-1 cells blocks avian sarcoma/leukosis virus release, demonstrating that this mechanism is functional under physiological conditions. The late block to release is caused in part by a loss in interaction between VPS4 and its coactivator protein LIP5, which is required to promote the formation of the ESCRT III-VPS4 double-hexamer complex to activate its ATPase. ISG15 is conjugated to two different LIP5-ESCRT-III-binding charged multivesicular body proteins, CHMP2A and CHMP5. Upon ISGylation of each, interaction with LIP5 is no longer detected. Two other ESCRT-III proteins, CHMP4B and CHMP6, are also conjugated to ISG15. ISGylation of CHMP2A, CHMP4B, and CHMP6 weakens their binding directly to VPS4, thereby facilitating the release of this protein from the membrane into the cytosol. The remaining budding complex fails to release particles from the cell membrane. Introducing a mutant of ISG15 into cells that cannot be conjugated to proteins prevents the ISG15-dependent mechanism from blocking virus release. CHMP5 is the primary switch to initiate the antiviral mechanism, because removal of CHMP5 from cells prevents ISGylation of CHMP2A and CHMP6.

  3. 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.

  4. 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

  5. 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.

  6. Proviral-activated c-erbB is leukemogenic but not sarcomagenic: characterization of a replication-competent retrovirus containing the activated c-erbB.

    PubMed Central

    Pelley, R J; Moscovici, C; Hughes, S; Kung, H J

    1988-01-01

    Avian leukosis virus (ALV) induces erythroblastosis in chickens by integrating its DNA into the host c-erbB locus and by activating expression of truncated c-erbB transcripts. Although there is a 100% correlation of c-erbB activation with ALV-induced erythroblastosis, direct evidence that the activated c-erbB is oncogenic has not been established. We have constructed a replication-competent retrovirus containing the activated c-erbB to investigate its transforming potential. The Rous c-erbB virus (REB-c) was constructed by inserting the activated c-erbB cDNA into a Rous sarcoma virus vector in place of src. When transfected into transformed quail fibroblasts (QT6), the REB-c construct stably integrates and expresses c-erbB-specific transcripts and produces infectious virus. The REB-c retrovirus produces short-latency polyclonal erythroblastosis in chickens. However, in contrast to avian erythroblastosis virus which contains v-erbB, the REB-c construct does not transform chicken embryo fibroblasts in vitro, nor does the REB-c virus produce sarcomas when injected into the wing web of chickens. Our results provide the first direct evidence that the activated c-erbB which lacks the amino-terminal extracellular domain but which retains the entire carboxy-terminal sequences is leukemogenic but not sarcomagenic. Images PMID:2833627

  7. 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

  8. Host Control of Insect Endogenous Retroviruses: Small RNA Silencing and Immune Response

    PubMed Central

    Fablet, Marie

    2014-01-01

    Endogenous retroviruses are relics of ancient infections from retroviruses that managed to integrate into the genome of germline cells and remained vertically transmitted from parent to progeny. Subsequent to the endogenization process, these sequences can move and multiply in the host genome, which can have deleterious consequences and disturb genomic stability. Natural selection favored the establishment of silencing pathways that protect host genomes from the activity of endogenous retroviruses. RNA silencing mechanisms are involved, which utilize piRNAs. The response to exogenous viral infections uses siRNAs, a class of small RNAs that are generated via a distinct biogenesis pathway from piRNAs. However, interplay between both pathways has been identified, and interactions with anti-bacterial and anti-fungal immune responses are also suspected. This review focuses on Diptera (Arthropods) and intends to compile pieces of evidence showing that the RNA silencing pathway of endogenous retrovirus regulation is not independent from immunity and the response to infections. This review will consider the mechanisms that allow the lasting coexistence of viral sequences and host genomes from an evolutionary perspective. PMID:25412365

  9. Shell disorder, immune evasion and transmission behaviors among human and animal retroviruses.

    PubMed

    Goh, Gerard Kian-Meng; Dunker, A Keith; Uversky, Vladimir N

    2015-08-01

    This study involves measurements of percentages of intrinsic disorder (PIDs) in the GAG protein shells of various retroviruses. Unique patterns of shell protein disorder can be seen especially when GAG proteins (matrix M, capsid C, and nucleocapsid N) of primate and non-primate retroviruses are compared. HIV-1 presents the most unique pattern of disorder distribution with generally high levels of disorder in all three proteins, while EIAV (PIDs:: 26, 29, 13) is diametrically different from HIV-1 (N C M PIDs: 39.5 ± 3.0, 44.5 ± 2.6, 56.5 ± 10.8). The HTLV viruses (CPID: 32.8 ± 3.4) resemble HIV-2 (C PID: 26.6 ± 2.9) with a moderately disordered capsid. Totally distinct patterns, however, are seen for the non-primate retroviruses. They generally have highly disordered nucleocapsids (PID > 65%) and more ordered outer shells especially the matrix. These characteristics might be attributed to the differences in the way the retroviruses are transmitted, with non-primate viruses having greater non-sexual transmission components such as oral-fecal transmission. These differences are also evolutionarily related to the ways the viruses evade the host immune systems, and thus, have implications for oncolytic virotherapy and animal models in vaccine research. The importance of protein shell disorder in immune evasion, as related to the case of HIV-1, and the difficult search for its vaccines are highlighted.

  10. Host control of insect endogenous retroviruses: small RNA silencing and immune response.

    PubMed

    Fablet, Marie

    2014-11-18

    Endogenous retroviruses are relics of ancient infections from retroviruses that managed to integrate into the genome of germline cells and remained vertically transmitted from parent to progeny. Subsequent to the endogenization process, these sequences can move and multiply in the host genome, which can have deleterious consequences and disturb genomic stability. Natural selection favored the establishment of silencing pathways that protect host genomes from the activity of endogenous retroviruses. RNA silencing mechanisms are involved, which utilize piRNAs. The response to exogenous viral infections uses siRNAs, a class of small RNAs that are generated via a distinct biogenesis pathway from piRNAs. However, interplay between both pathways has been identified, and interactions with anti-bacterial and anti-fungal immune responses are also suspected. This review focuses on Diptera (Arthropods) and intends to compile pieces of evidence showing that the RNA silencing pathway of endogenous retrovirus regulation is not independent from immunity and the response to infections. This review will consider the mechanisms that allow the lasting coexistence of viral sequences and host genomes from an evolutionary perspective.

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

    SciTech Connect

    Serafino, A. Balestrieri, E.; Pierimarchi, P.; Matteucci, C.; Moroni, G.; Oricchio, E.; Rasi, G.; Mastino, A.; Spadafora, C.; Garaci, E.; Vallebona, P. Sinibaldi

    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-derived 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.

  12. Human RNA “Rumor” Viruses: the Search for Novel Human Retroviruses in Chronic Disease

    PubMed Central

    Voisset, Cécile; Weiss, Robin A.; Griffiths, David J.

    2008-01-01

    Summary: Retroviruses are an important group of pathogens that cause a variety of diseases in humans and animals. Four human retroviruses are currently known, including human immunodeficiency virus type 1, which causes AIDS, and human T-lymphotropic virus type 1, which causes cancer and inflammatory disease. For many years, there have been sporadic reports of additional human retroviral infections, particularly in cancer and other chronic diseases. Unfortunately, many of these putative viruses remain unproven and controversial, and some retrovirologists have dismissed them as merely “human rumor viruses.” Work in this field was last reviewed in depth in 1984, and since then, the molecular techniques available for identifying and characterizing retroviruses have improved enormously in sensitivity. The advent of PCR in particular has dramatically enhanced our ability to detect novel viral sequences in human tissues. However, DNA amplification techniques have also increased the potential for false-positive detection due to contamination. In addition, the presence of many families of human endogenous retroviruses (HERVs) within our DNA can obstruct attempts to identify and validate novel human retroviruses. Here, we aim to bring together the data on “novel” retroviral infections in humans by critically examining the evidence for those putative viruses that have been linked with disease and the likelihood that they represent genuine human infections. We provide a background to the field and a discussion of potential confounding factors along with some technical guidelines. In addition, some of the difficulties associated with obtaining formal proof of causation for common or ubiquitous agents such as HERVs are discussed. PMID:18322038

  13. Custom human endogenous retroviruses dedicated microarray identifies self-induced HERV-W family elements reactivated in testicular cancer upon methylation control

    PubMed Central

    Gimenez, Juliette; Montgiraud, Cécile; Pichon, Jean-Philippe; Bonnaud, Bertrand; Arsac, Maud; Ruel, Karine; Bouton, Olivier; Mallet, François

    2010-01-01

    Endogenous retroviruses (ERVs) are an inherited part of the eukaryotic genomes, and represent ∼400 000 loci in the human genome. Human endogenous retroviruses (HERVs) can be divided into distinct families, composed of phylogenetically related but structurally heterogeneous elements. The majority of HERVs are silent in most physiological contexts, whereas a significant expression is observed in pathological contexts, such as cancers. Owing to their repetitive nature, few of the active HERV elements have been accurately identified. In addition, there are no criteria defining the active promoters among HERV long-terminal repeats (LTRs). Hence, it is difficult to understand the HERV (de)regulation mechanisms and their implication on the physiopathology of the host. We developed a microarray to specifically detect the LTR-containing transcripts from the HERV-H, HERV-E, HERV-W and HERV-K(HML-2) families. HERV transcriptome was analyzed in the placenta and seven normal/tumoral match-pair samples. We identified six HERV-W loci overexpressed in testicular cancer, including a usually placenta-restricted transcript of ERVWE1. For each locus, specific overexpression was confirmed by quantitative RT-PCR, and comparison of the activity of U3 versus U5 regions suggested a U3-promoted transcription coupled with 5′R initiation. The analysis of DNA from tumoral versus normal tissue revealed that hypomethylation of U3 promoters in tumors is a prerequisite for their activation. PMID:20053729

  14. 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.

  15. The human endogenous retrovirus link between genes and environment in multiple sclerosis and in multifactorial diseases associating neuroinflammation.

    PubMed

    Perron, Hervé; Lang, Alois

    2010-08-01

    Endogenous retroviruses represent about 8% of the human genome and belong to the superfamily of transposable and retrotransposable genetic elements. Altogether, these mobile genetic elements and their numerous inactivated "junk" sequences represent nearly one half of the human DNA. Nonetheless, a significant part of this "non-conventional" genome has retained potential activity. Epigenetic control is notably involved in silencing most of these genetic elements but certain environmental factors such as viruses are known to dysregulate their expression in susceptible cells. More particularly, embryonal cells with limited gene methylation are most susceptible to uncontrolled activation of these mobile genetic elements by, e.g., viral infections. In particular, certain viruses transactivate promoters from endogenous retroviral family type W (HERV-W). HERV-W RNA was first isolated in circulating viral particles (Multiple Sclerosis-associated RetroViral element, MSRV) that have been associated with the evolution and prognosis of multiple sclerosis. HERV-W elements encode a powerful immunopathogenic envelope protein (ENV) that activates a pro-inflammatory and autoimmune cascade through interaction with Toll-like receptor 4 on immune cells. This ENV protein has repeatedly been detected in MS brain lesions and may be involved in other diseases. Epigenetic factors controlling HERV-W ENV protein expression then reveal critical. This review addresses the gene-environment epigenetic interface of such HERV-W elements and its potential involvement in disease.

  16. Infection with human retroviruses other than HIV-1: HIV-2, HTLV-1, HTLV-2, HTLV-3 and HTLV-4.

    PubMed

    Nicolás, David; Ambrosioni, Juan; Paredes, Roger; Marcos, M Ángeles; Manzardo, Christian; Moreno, Asunción; Miró, José M

    2015-08-01

    HIV-1 is the most prevalent retrovirus, with over 30 million people infected worldwide. Nevertheless, infection caused by other human retroviruses like HIV-2, HTLV-1, HTLV-2, HTLV-3 and HTLV-4 is gaining importance. Initially confined to specific geographical areas, HIV-2, HTLV-1 and HTLV-2 are becoming a major concern in non-endemic countries due to international migration flows. Clinical manifestations of retroviruses range from asymptomatic carriers to life-threatening conditions, such as AIDS in HIV-2 infection or adult T-cell lymphoma/leukemia or tropical spastic paraparesis in HTLV-1 infection. HIV-2 is naturally resistant to some antiretrovirals frequently used to treat HIV-1 infection, but it does have effective antiretroviral therapy options. Unfortunately, HTLV still has limited therapeutic options. In this article, we will review the epidemiological, clinical, diagnostic, pathogenic and therapeutic aspects of infections caused by these human retroviruses.

  17. SIRE-1, a putative plant retrovirus is closely related to a legume TY1-copia retrotransposon family.

    PubMed

    Pearce, Stephen R

    2007-01-01

    SIRE-1 is a potential soybean retrovirus which has a gene order similar to Ty1-copia retrotransposons but also contains an envelope-like open reading frame (ORF), which is characteristic of retroviruses. PCR and Southern analysis reveals that SIRE-1 is closely related to a legume-wide family of envelope-lacking Ty1-copia group retrotransposons which suggests that SIRE-1 was formed by the recent acquisition of an envelope gene by a Ty1-copia retrotransposon.

  18. Retroviruses use CD169-mediated trans-infection of permissive lymphocytes to establish infection.

    PubMed

    Sewald, Xaver; Ladinsky, Mark S; Uchil, Pradeep D; Beloor, Jagadish; Pi, Ruoxi; Herrmann, Christin; Motamedi, Nasim; Murooka, Thomas T; Brehm, Michael A; Greiner, Dale L; Shultz, Leonard D; Mempel, Thorsten R; Bjorkman, Pamela J; Kumar, Priti; Mothes, Walther

    2015-10-30

    Dendritic cells can capture and transfer retroviruses in vitro across synaptic cell-cell contacts to uninfected cells, a process called trans-infection. Whether trans-infection contributes to retroviral spread in vivo remains unknown. Here, we visualize how retroviruses disseminate in secondary lymphoid tissues of living mice. We demonstrate that murine leukemia virus (MLV) and human immunodeficiency virus (HIV) are first captured by sinus-lining macrophages. CD169/Siglec-1, an I-type lectin that recognizes gangliosides, captures the virus. MLV-laden macrophages then form long-lived synaptic contacts to trans-infect B-1 cells. Infected B-1 cells subsequently migrate into the lymph node to spread the infection through virological synapses. Robust infection in lymph nodes and spleen requires CD169, suggesting that a combination of fluid-based movement followed by CD169-dependent trans-infection can contribute to viral spread.

  19. 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

  20. [Additive effect of marihuana and retrovirus in the anergy of natural killer cells in mice].

    PubMed

    Ongrádi, J; Specter, S; Horváth, A; Friedman, H

    1999-01-10

    Among the immunosuppressive effects of marijuana, impairment of natural killer cell activity is significant. HIV also inhibits these cells. Friend leukemia virus complex and its helper component Rowson-Parr virus induce early immunosuppression in mice resembling human AIDS, and late leukemia, providing a small animal AIDS model. Leukemia susceptible BALB/c and resistant C57BL/6 mice were infected with these viruses. At different time points, their natural killer cells separated from spleens were treated with 0 to 10 micrograms/ml tetrahydrocannabinol, subsequently mixed with Yac-1 target cells for 4 and 18 h. The natural killer cell activity in both mouse strains infected by either virus complex or helper virus weakened on days 2 to 4 postinfection, normalized by day 8 and enhanced on days 11 to 14. Natural killer cell activity upon the effect of low concentration (1.0 to 2.5 micrograms/ml) of tetrahydrocannabinol slightly increased in BALB/c, was unaffected in C57BL/6, especially in 18 h assays. In the combined effects of marijuana and retrovirus, damages by marijuana dominated over those of retroviruses. Inhibition or reactive enhancement of natural killer cell activity on the effect of viruses are similar to those of infected but marijuana-free counterparts, but on the level of uninfected cells treated with marijuana. The effects of marijuana and retrovirus are additive resulting in anergy of natural killer cells.

  1. One hundred twenty years of koala retrovirus evolution determined from museum skins.

    PubMed

    Ávila-Arcos, María C; Ho, Simon Y W; Ishida, Yasuko; Nikolaidis, Nikolas; Tsangaras, Kyriakos; Hönig, Karin; Medina, Rebeca; Rasmussen, Morten; Fordyce, Sarah L; Calvignac-Spencer, Sébastien; Willerslev, Eske; Gilbert, M Thomas P; Helgen, Kristofer M; Roca, Alfred L; Greenwood, Alex D

    2013-02-01

    Although endogenous retroviruses are common across vertebrate genomes, the koala retrovirus (KoRV) is the only retrovirus known to be currently invading the germ line of its host. KoRV is believed to have first infected koalas in northern Australia less than two centuries ago. We examined KoRV in 28 koala museum skins collected in the late 19th and 20th centuries and deep sequenced the complete proviral envelope region from five northern Australian specimens. Strikingly, KoRV env sequences were conserved among koalas collected over the span of a century, and two functional motifs that affect viral infectivity were fixed across the museum koala specimens. We detected only 20 env polymorphisms among the koalas, likely representing derived mutations subject to purifying selection. Among northern Australian koalas, KoRV was already ubiquitous by the late 19th century, suggesting that KoRV evolved and spread among koala populations more slowly than previously believed. Given that museum and modern koalas share nearly identical KoRV sequences, it is likely that koala populations, for more than a century, have experienced increased susceptibility to diseases caused by viral pathogenesis.

  2. Adaptation of a retrovirus as a eucaryotic vector transmitting the herpes simplex virus thymidine kinase gene

    SciTech Connect

    Tabin, C.J.; Hoffman, J.W.; Goff, S.P.; Weinberg, R.A.

    1982-04-01

    The authors investigated the feasibility of using retroviruses as vectors for transferring DNA sequences into animal cells. The thymidine kinase (tk) gene of herpes simplex virus was chosen as a convenient model. The internal BamHI fragments of a DNA clone of Moloney leukemia virus (MLV) were replaced with a purified BamHI DNA segment containing the tk gene. Chimeric genomes were created carrying the tk insert on both orientations relative to the MLV sequence. Each was transfected into TK/sup -/ cells along with MLV helper virus, and TK/sup +/ colonies were obtained by selection in the presence of hypoxanthine, aminopterin, and thymidine (HAT). Virus collected from TK/sup +/-transformed, MLV producer cells passed the TK/sup +/ phenotype to TK/sup -/ cells. Nonproducer cells were isolated, and TK/sup +/ transducing virus was subsequently rescued from them. The chimeric virus showed single-hit kinetics in infections. Virion and cellular RNA and cellular DNA from infected cells were all shown to contain sequences which hybridized to both MLV- and tk-specific probes. The sizes of these sequences were consistent with those predicted for the chimeric virus. In all respects studied, the chimeric MLV-tk virus behaved like known replication-defective retroviruses. These experiments suggest great general applicability of retroviruses as eucaryotic vectors.

  3. Existence of Two Distinct Infectious Endogenous Retroviruses in Domestic Cats and Their Different Strategies for Adaptation to Transcriptional Regulation

    PubMed Central

    Kuse, Kyohei; Ito, Jumpei; Miyake, Ariko; Kawasaki, Junna; Watanabe, Shinya; Makundi, Isaac; Ngo, Minh Ha; Otoi, Takeshige

    2016-01-01

    ABSTRACT Endogenous retroviruses (ERVs) are the remnants of ancient retroviral infections of germ cells. Previous work identified one of the youngest feline ERV groups, ERV-DC, and reported that two ERV-DC loci, ERV-DC10 and ERV-DC18 (ERV-DC10/DC18), can replicate in cultured cells. Here, we identified another replication-competent provirus, ERV-DC14, on chromosome C1q32. ERV-DC14 differs from ERV-DC10/DC18 in its phylogeny, receptor usage, and, most notably, transcriptional activities; although ERV-DC14 can replicate in cultured cells, it cannot establish a persistent infection owing to its low transcriptional activity. Furthermore, we examined ERV-DC transcription and its regulation in feline tissues. Quantitative reverse transcription-PCR (RT-PCR) detected extremely low ERV-DC10 expression levels in feline tissues, and bisulfite sequencing showed that 5′ long terminal repeats (LTRs) of ERV-DC10/DC18 are significantly hypermethylated in feline blood cells. Reporter assays found that the 5′-LTR promoter activities of ERV-DC10/DC18 are high, whereas that of ERV-DC14 is low. This difference in promoter activity is due to a single substitution from A to T in the LTR, and reverse mutation at this nucleotide in ERV-DC14 enhanced its replication and enabled it to persistently infect cultured cells. Therefore, ERV-DC LTRs can be divided into two types based on this nucleotide, the A type or T type, which have strong or attenuated promoter activity, respectively. Notably, ERV-DCs with T-type LTRs, such as ERV-DC14, have expanded in the cat genome significantly more than A-type ERV-DCs, despite their low promoter activities. Our results provide insights into how the host controls potentially infectious ERVs and, conversely, how ERVs adapt to and invade the host genome. IMPORTANCE The domestic cat genome contains many endogenous retroviruses, including ERV-DCs. These ERV-DCs have been acquired through germ cell infections with exogenous retroviruses. Some of these ERV

  4. 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.

  5. The interferon-induced gene ISG15 blocks retrovirus release from cells late in the budding process.

    PubMed

    Pincetic, Andrew; Kuang, Zhizhou; Seo, Eun Joo; Leis, Jonathan

    2010-05-01

    The release of retroviruses from cells requires ubiquitination of Gag and recruitment of cellular proteins involved in endosome sorting, including the ESCRT-III proteins and the Vps4 ATPase. In response to infection, cells have evolved an interferon-induced mechanism to block virus replication through expression of the interferon-stimulated gene 15 (ISG15), a dimer homologue of ubiquitin, which interferes with ubiquitin pathways in cells. Previously, it has been reported that ISG15 expression inhibited the E3 ubiquitin ligase, Nedd4, and prevented association of the ESCRT-I protein Tsg101 with human immunodeficiency virus type 1 (HIV-1) Gag. The budding of avian sarcoma leukosis virus and HIV-1 Gag virus-like particles containing L-domain mutations can be rescued by fusion to ESCRT proteins, which cause entry into the budding pathway beyond these early steps. The release of these fusions from cells was susceptible to inhibition by ISG15, indicating that there was a block late in the budding process. We now demonstrate that the Vps4 protein does not associate with the avian sarcoma leukosis virus or the HIV-1 budding complexes when ISG15 is expressed. This is caused by a loss in interaction between Vps4 with its coactivator protein LIP5 needed to promote the formation of the ESCRT-III-Vps4 double-hexamer complex required for membrane scission and virus release. The inability of LIP5 to interact with Vps4 is the probable result of ISG15 conjugation to the ESCRT-III protein, CHMP5, which regulates the availability of LIP5. Thus, there appear to be multiple levels of ISG15-induced inhibition acting at different stages of the virus release process.

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

    PubMed Central

    Hu, Yani; O’Boyle, Kaitlin; Auer, Jim; You, Fuping; Wang, Penghua; Fikrig, Erol

    2017-01-01

    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. PMID:28152074

  7. An evaluation of the putative human mammary tumour retrovirus associated with peripheral blood monocytes.

    PubMed Central

    Kahl, L. P.; Carroll, A. R.; Rhodes, P.; Wood, J.; Read, N. G.

    1991-01-01

    The primary aims of this study were purification and molecular cloning of a putative retrovirus designated human mammary tumour virus (HMTV). However, our preliminary unpublished data of negative reverse transcriptase (RT) activity in ostensibly 'infected' cells led us to re-examine the evidence for this virus; namely multinucleate giant cell (MNGC) formation and RT activity in cultured blood monocytes from breast cancer patients versus benign breast tumour and normal control subjects. MNGCs from by fusion of monocytes and we estimated the total number of cell fusions which had occurred after 10 days of culture in vitro by counting cells with two, three, four and five or more nuclei (n) and by measuring the density of adherent mononuclear cells for each subject studied. We found no clear-cut difference in MNGC formation between the three subject groups. Moreover, a substantial number of cultures, encompassing the three groups, showed far more MNGCs per 10(5) monocytes than previously reported. Various parametric and nonparametric statistical analyses were performed on the multinucleate cell data and only one parametric test, which utilised the density of monolayers as a co-variate, showed a statistically significant difference at the 5% level between the breast cancer and the normal subject groups. We observed marked subject-to-subject variation in multinucleate cell formation and we suggest that the evidence for a difference between the breast cancer and the normal groups is marginal. Further, MNGC formation by breast cancer monocytes may not be attributed to the presence of a retrovirus since 5'-Azacytidine (AZA), an agent known to stimulate replication of latent retroviruses showed no effect on the MNGC formation. In addition, culture supernatants from the three groups were assayed for RT activity and no test sample gave a significant signal above background. Preliminary transmission electron microscopy analysis failed to identify viral particles in MNGCs

  8. Proliferation of endogenous retroviruses in the early stages of a host germ line invasion.

    PubMed

    Ishida, Yasuko; Zhao, Kai; Greenwood, Alex D; Roca, Alfred L

    2015-01-01

    Endogenous retroviruses (ERVs) comprise 8% of the human genome and are common in all vertebrate genomes. The only retrovirus known to be currently transitioning from exogenous to endogenous form is the koala retrovirus (KoRV), making koalas (Phascolarctos cinereus) ideal for examining the early stages of retroviral endogenization. To distinguish endogenous from exogenous KoRV proviruses, we isolated koala genomic regions flanking KoRV integration sites. In three wild southern Australian koalas, there were fewer KoRV loci than in three captive Queensland koalas, consistent with reports that southern Australian koalas carry fewer KoRVs. Of 39 distinct KoRV proviral loci examined in a sire-dam-progeny triad, all proved to be vertically transmitted and endogenous; none was exogenous. Of the 39 endogenous KoRVs (enKoRVs), only one was present in the genomes of both the sire and the dam, suggesting that, at this early stage in the retroviral invasion of a host germ line, very large numbers of ERVs have proliferated at very low frequencies in the koala population. Sequence divergence between the 5'- and 3'-long terminal repeats (LTRs) of a provirus can be used as a molecular clock. Within each of ten enKoRVs, the 5'-LTR sequence was identical to the 3'-LTR sequence, suggesting a maximum age for enKoRV invasion of the koala germ line of approximately 22,200-49,900 years ago, although a much younger age is possible. Across the ten proviruses, seven LTR haplotypes were detected, indicating that at least seven different retroviral sequences had entered the koala germ line.

  9. 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.

  10. The population history of endogenous retroviruses in mule deer (Odocoileus hemionus).

    PubMed

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

    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.

  11. Proliferation of Endogenous Retroviruses in the Early Stages of a Host Germ Line Invasion

    PubMed Central

    Ishida, Yasuko; Zhao, Kai; Greenwood, Alex D.; Roca, Alfred L.

    2015-01-01

    Endogenous retroviruses (ERVs) comprise 8% of the human genome and are common in all vertebrate genomes. The only retrovirus known to be currently transitioning from exogenous to endogenous form is the koala retrovirus (KoRV), making koalas (Phascolarctos cinereus) ideal for examining the early stages of retroviral endogenization. To distinguish endogenous from exogenous KoRV proviruses, we isolated koala genomic regions flanking KoRV integration sites. In three wild southern Australian koalas, there were fewer KoRV loci than in three captive Queensland koalas, consistent with reports that southern Australian koalas carry fewer KoRVs. Of 39 distinct KoRV proviral loci examined in a sire–dam–progeny triad, all proved to be vertically transmitted and endogenous; none was exogenous. Of the 39 endogenous KoRVs (enKoRVs), only one was present in the genomes of both the sire and the dam, suggesting that, at this early stage in the retroviral invasion of a host germ line, very large numbers of ERVs have proliferated at very low frequencies in the koala population. Sequence divergence between the 5′- and 3′-long terminal repeats (LTRs) of a provirus can be used as a molecular clock. Within each of ten enKoRVs, the 5′-LTR sequence was identical to the 3′-LTR sequence, suggesting a maximum age for enKoRV invasion of the koala germ line of approximately 22,200–49,900 years ago, although a much younger age is possible. Across the ten proviruses, seven LTR haplotypes were detected, indicating that at least seven different retroviral sequences had entered the koala germ line. PMID:25261407

  12. Molecular Mechanisms of Cytopathogenicity of Primate Lymphotropic Retroviruses: Relevance to Treatment and Vaccine for AIDS

    DTIC Science & Technology

    1989-03-10

    Covering the Period 9/29/87 to 9/28/88 by 0Mark M . Manak and Linda Jagodzinski March 10, 1989 Supported by: US ARMY MEDICAL RESEARCH AND DEVELOPMENT...of Cytopathogenicity of Primate Lymphotropic Retroviruses: Relevance to Treatment and Vaccine for AIDS 12. PERSONAL AUTHOR(S) Manak, Mark M . and...1 hour. The mixture was adjusted to a pH >8.5 by the addition of 1 M TrisHCI, pH 9.0, and extracted three times with equal volumes of n-butanol. The

  13. Identification of a novel subgroup of Koala retrovirus from Koalas in Japanese zoos.

    PubMed

    Shojima, Takayuki; Yoshikawa, Rokusuke; Hoshino, Shigeki; Shimode, Sayumi; Nakagawa, So; Ohata, Takuji; Nakaoka, Rie; Miyazawa, Takayuki

    2013-09-01

    We identified a new subgroup of koala retrovirus (KoRV), named KoRV-J, which utilizes thiamine transport protein 1 as a receptor instead of the Pit-1 receptor used by KoRV (KoRV-A). By subgroup-specific PCR, KoRV-J and KoRV-A were detected in 67.5 and 100% of koalas originating from koalas from northern Australia, respectively. Altogether, our results indicate that the invasion of the koala population by KoRV-J may have occurred more recently than invasion by KoRV-A.

  14. Selected highlights on women and HIV from the 5th Conference on Retroviruses and Opportunistic Infections.

    PubMed

    Bartnof, H S

    1998-04-01

    Many sessions at the 5th Conference on Retroviruses and Opportunistic Infections dealt specifically with HIV infection and treatment in women. Highlights are presented from several sessions, including indinavir blood levels at various points in the menstrual cycle, abnormal kidney function associated with women taking indinavir, abnormal pap smears in women with high viral load, the relationship between viral load and the increased risk of death in women, and the impact of ddI crossing the placenta in pregnant women. Information is given on each presentation, including clinical trial results, side effects, and impacts on disease progression.

  15. Weak Palindromic Consensus Sequences Are a Common Feature Found at the Integration Target Sites of Many Retroviruses

    PubMed Central

    Wu, Xiaolin; Li, Yuan; Crise, Bruce; Burgess, Shawn M.; Munroe, David J.

    2005-01-01

    Integration into the host genome is one of the hallmarks of the retroviral life cycle and is catalyzed by virus-encoded integrases. While integrase has strict sequence requirements for the viral DNA ends, target site sequences have been shown to be very diverse. We carefully examined a large number of integration target site sequences from several retroviruses, including human immunodeficiency virus type 1, simian immunodeficiency virus, murine leukemia virus, and avian sarcoma-leukosis virus, and found that a statistical palindromic consensus, centered on the virus-specific duplicated target site sequence, was a common feature at integration target sites for these retroviruses. PMID:15795304

  16. HIV infection and HERV expression: a review

    PubMed Central

    2012-01-01

    The human genome contains multiple copies of retrovirus genomes known as endogenous retroviruses (ERVs) that have entered the germ-line at some point in evolution. Several of these proviruses have retained (partial) coding capacity, so that a number of viral proteins or even virus particles are expressed under various conditions. Human ERVs (HERVs) belong to the beta-, gamma-, or spuma- retrovirus groups. Endogenous delta- and lenti- viruses are notably absent in humans, although endogenous lentivirus genomes have been found in lower primates. Exogenous retroviruses that currently form a health threat to humans intriguingly belong to those absent groups. The best studied of the two infectious human retroviruses is the lentivirus human immunodeficiency virus (HIV) which has an overwhelming influence on its host by infecting cells of the immune system. One HIV-induced change is the induction of HERV transcription, often leading to induced HERV protein expression. This review will discuss the potential HIV-HERV interactions. Several studies have suggested that HERV proteins are unlikely to complement defective HIV virions, nor is HIV able to package HERV transcripts, probably due to low levels of sequence similarity. It is unclear whether the expression of HERVs has a negative, neutral, or positive influence on HIV-AIDS disease progression. A positive effect was recently reported by the specific expression of HERVs in chronically HIV-infected patients, which results in the presentation of HERV-derived peptides to CD8+ T-cells. These cytotoxic T-cells were not tolerant to HERV peptides, as would be expected for self-antigens, and consequently lysed the HIV-infected, HERV-presenting cells. This novel mechanism could control HIV replication and result in a low plasma viral load. The possibility of developing a vaccination strategy based on these HERV peptides will be discussed. PMID:22248111

  17. Intracellular trafficking of Gag and Env proteins and their interactions modulate pseudotyping of retroviruses.

    PubMed

    Sandrin, Virginie; Muriaux, Delphine; Darlix, Jean-Luc; Cosset, François-Loïc

    2004-07-01

    Glycoproteins derived from most retroviruses and from several families of enveloped viruses can form infectious pseudotypes with murine leukemia virus (MLV) and lentiviral core particles, like the MLV envelope glycoproteins (Env) that are incorporated on either virus type. However, coexpression of a given glycoprotein with heterologous core proteins does not always give rise to highly infectious viral particles, and restrictions on pseudotype formation have been reported. To understand the mechanisms that control the recruitment of viral surface glycoproteins on lentiviral and retroviral cores, we exploited the fact that the feline endogenous retrovirus RD114 glycoprotein does not efficiently pseudotype lentiviral cores derived from simian immunodeficiency virus, whereas it is readily incorporated onto MLV particles. Our results indicate that recruitment of glycoproteins by the MLV and lentiviral core proteins occurs in intracellular compartments and not at the cell surface. We found that Env and core protein colocalization in intracytoplasmic vesicles is required for pseudotype formation. By investigating MLV/RD114 Env chimeras, we show that signals in the cytoplasmic tail of either glycoprotein differentially influenced their intracellular localization; that of MLV allows endosomal localization and hence recruitment by both lentiviral and MLV cores. Furthermore, we found that upon membrane binding, MLV core proteins could relocalize Env glycoproteins in late endosomes and allow their incorporation on viral particles. Thus, intracellular colocalization, as well as interactions between Env and core proteins, may influence the recruitment of the glycoprotein onto viral particles and generate infectious pseudotyped viruses.

  18. 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.

  19. 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.

  20. A retrovirus isolated from cell lines derived from neurofibromas in bicolor damselfish (Pomacentrus partitus).

    PubMed

    Schmale, M C; Aman, M R; Gill, K A

    1996-06-01

    Damselfish neurofibromatosis (DNF) is a naturally occurring, neoplastic disease affecting bicolor damselfish (Pomacentrus partitus) living on coral reefs in southern Florida, USA. The disease consists of multiple neurofibromas, neurofibrosarcomas and chromatophoromas and has been proposed as an animal model for neurofibromatosis type 1 in humans. DNF is transmissible by injection of crude tumour homogenates, cell-free filtrates of homogenates or cells from tumour cell lines. An analysis of tumorigenic cell lines derived from fish with spontaneous or experimentally induced DNF revealed virus particles budding from cells and present in conditioned media. The 90-110 nm particles resembled type C retroviruses. This virus exhibited a buoyant density of 1.14-1.17 g/cm2 in sucrose, at least six virus proteins of 15 to 80 kDa and reverse transcriptase (RT) activity. RT activity was maximized with a poly(rC).oligo(dG) template.primer combination and Mn2+ at a concentration of 0.5-1.0 mM. The optimum temperature for RT was determined to be 20 degrees C, a finding consistent with the ambient temperatures encountered by this species. This retrovirus, tentatively named damselfish neurofibromatosis virus (DNFV) may be the aetiological agent of DNF. Whether DNFV or another, as yet unidentified, virus is the cause of DNF, this agent may be unique in virus oncogenesis; neoplastic transformation of the cell types involved in DNF, Schwann cells and chromatophores, has not been documented in any other transmissible tumour.

  1. 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

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

    PubMed Central

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

    2014-01-01

    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. PMID:25462343

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

    SciTech Connect

    Vargas, Amandine Thiery, Maxime Lafond, Julie Barbeau, Benoit

    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 were 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.

  4. A report from the Conference on Retroviruses and Opportunistic Infections (CROI) 2015 (February 23-26--Seattle, Washington, USA).

    PubMed

    Rabasseda, X

    2015-03-01

    The largest conference on HIV and AIDS, the annual Conference on Retroviruses and Opportunistic Infections (CROI) also encompasses opportunistic and other important pathogens, notably in 2015 hepatitis C virus and the Ebola virus. This year's conference involved 4 days of discussions on highly relevant basic, translational and clinical research and developments in the ongoing battle against these infections.

  5. Annual Conference on Human Retrovirus Testing (7th) held in Chicago, IL on March 3-5, 1992

    DTIC Science & Technology

    1993-04-05

    Microbiologist, California State Immunology, Indiana State Department Health Services Department of Health Patricia Garrett, Boston Biomedica Harold...V Biomedica . Inc. Counseling & Testing Program, Illinois Dr. Robert J Dwyer, Director, Diagnostic Department of Public Health Development. IAF Biochem...Keith Sayre, Retrovirus Product Manager. Steven Rala, Boston Biomedica Ortho Diagnostic Systems Michael Ramirez, Public Health Charles Schable. M.S

  6. The nucleotide sequence of koala (Phascolarctos cinereus) retrovirus: a novel type C endogenous virus related to Gibbon ape leukemia virus.

    PubMed

    Hanger, J J; Bromham, L D; McKee, J J; O'Brien, T M; Robinson, W F

    2000-05-01

    A novel retrovirus, morphologically consistent with mammalian C-type retroviruses, was detected by electron microscopy in mitogen-stimulated peripheral blood mononuclear cell cultures from 163 koalas and in lymphoma tissue from 3 koalas. PCR amplified provirus from the blood and tissues of 17 wild and captive koalas, and reverse transcriptase-PCR demonstrated viral mRNA, viral genomic RNA, and reverse transcriptase activity in koala serum and cell culture supernatants. Comparison of viral sequences derived from genomic DNA and mRNA showed identity indicative of a single retroviral species-here designated koala retrovirus (KoRV). Southern blot analysis of koala tissue genomic DNA using labelled KoRV probes demonstrated banding consistent with an endogenous retrovirus. Complete and apparently truncated proviruses were detected in DNA of both clinically normal koalas and those with hematopoietic disease. KoRV-related viruses were not detected in other marsupials, and phylogenetic analysis showed that KoRV paradoxically clusters with gibbon ape leukemia virus (GALV). The strong similarity between GALV and KoRV suggests that these viruses are closely related and that recent cross-host transmission has occurred. The complete proviral DNA sequence of KoRV is reported.

  7. Complete cure of established murine hepatocellular carcinoma is achievable by repeated injections of retroviruses carrying the herpes simplex virus thymidine kinase gene.

    PubMed

    Kuriyama, S; Masui, K; Kikukawa, M; Sakamoto, T; Nakatani, T; Nagao, S; Yamazaki, M; Yoshiji, H; Tsujinoue, H; Fukui, H; Yoshimatsu, T; Ikenaka, K

    1999-04-01

    Although xenotransplantation of retrovirus-producing cells into a tumor has been shown to be effective for the treatment of cancer, injections of recombinant retroviruses are much more feasible for clinical applications. We established a clone producing retroviruses carrying the herpes simplex virus thymidine kinase (HSVtk) gene with titers of up to 4 x 10(7) colony-forming units/ml, and examined the effectiveness of in vivo gene therapy against cancer. Syngeneic mice were inoculated subcutaneously with murine hepatocellular carcinoma (HCC) cells, BNL1ME A.7R.1, and the treatment was initiated after tumors were established. When mice were given an intratumoral injection of HSVtk-carrying retroviruses or their producing cells followed by ganciclovir (GCV) treatment, significantly prolonged survival periods were observed. When mice were treated with repeated intratumoral injections of HSVtk-carrying retrovirus-producing cells, significant antitumor responses and some cures were induced by GCV treatment. Furthermore, repeated intratumoral injections of HSVtk-carrying retroviruses and GCV treatment resulted in complete regression of established HCC tumors in all animals used in the experiment. Mice that completely eradicated tumors exhibited protective immunity against wild-type HCC tumors. These results suggest that repeated injections of HSVtk-carrying retroviruses followed by GCV treatment is a potent modality for the treatment of solid tumors.

  8. 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

  9. Natural killer cells recognize friend retrovirus-infected erythroid progenitor cells through NKG2D-RAE-1 interactions In Vivo.

    PubMed

    Ogawa, Tatsuya; Tsuji-Kawahara, Sachiyo; Yuasa, Takae; Kinoshita, Saori; Chikaishi, Tomomi; Takamura, Shiki; Matsumura, Haruo; Seya, Tsukasa; Saga, Toshihiko; Miyazawa, Masaaki

    2011-06-01

    Natural killer (NK) cells function as early effector cells in the innate immune defense against viral infections and also participate in the regulation of normal and malignant hematopoiesis. NK cell activities have been associated with early clearance of viremia in experimental simian immunodeficiency virus and clinical human immunodeficiency virus type 1 (HIV-1) infections. We have previously shown that NK cells function as major cytotoxic effector cells in vaccine-induced immune protection against Friend virus (FV)-induced leukemia, and NK cell depletion totally abrogates the above protective immunity. However, how NK cells recognize retrovirus-infected cells remains largely unclear. The present study demonstrates a correlation between the expression of the products of retinoic acid early transcript-1 (RAE-1) genes in target cells and their susceptibility to killing by NK cells isolated from FV-infected animals. This killing was abrogated by antibodies blocking the NKG2D receptor in vitro. Further, the expression of RAE-1 proteins on erythroblast surfaces increased early after FV inoculation, and administration of an RAE-1-blocking antibody resulted in increased spleen infectious centers and exaggerated pathology, indicating that FV-infected erythroid cells are recognized by NK cells mainly through the NKG2D-RAE-1 interactions in vivo. Enhanced retroviral replication due to host gene-targeting resulted in markedly increased RAE-1 expression in the absence of massive erythroid cell proliferation, indicating a direct role of retroviral replication in RAE-1 upregulation.

  10. A Novel Model of SCID-X1 Reconstitution Reveals Predisposition to Retrovirus-induced Lymphoma but No Evidence of γC Gene Oncogenicity.

    PubMed

    Scobie, Linda; Hector, Ralph D; Grant, Louise; Bell, Margaret; Nielsen, Anne A; Meikle, Sharon; Philbey, Adrain; Thrasher, Adrain J; Cameron, Ewan R; Blyth, Karen; Neil, James C

    2009-06-01

    The emergence of leukemia following gene transfer to restore common cytokine receptor γ chain (γC) function in X-linked severe combined immunodeficiency (SCID-X1) has raised important questions with respect to gene therapy safety. To explore the risk factors involved, we tested the oncogenic potential of human γC in new strains of transgenic mice expressing the gene under the control of the CD2 promoter and locus control region (LCR). These mice demonstrated mildly perturbed T-cell development, with an increased proportion of thymic CD8 cells, but showed no predisposition to tumor development even on highly tumor prone backgrounds or after γ-retrovirus infection. The human CD2-γC transgene rescued T and B-cell development in γC(-/-) mice but with an age-related delay, mimicking postnatal reconstitution in SCID-X1 gene therapy subjects. However, we noted that γC(-/-) mice are acutely susceptible to murine leukemia virus (MLV) leukemogenesis, and that this trait was not corrected by the γC transgene. We conclude that the SCID-X1 phenotype can be corrected safely by stable ectopic expression of γC and that the transgene is not significantly oncogenic when expressed in this context. However, an underlying predisposition conferred by the SCID-X1 background appears to collaborate with insertional mutagenesis to increase the risk of tumor development.

  11. A novel model of SCID-X1 reconstitution reveals predisposition to retrovirus-induced lymphoma but no evidence of gammaC gene oncogenicity.

    PubMed

    Scobie, Linda; Hector, Ralph D; Grant, Louise; Bell, Margaret; Nielsen, Anne A; Meikle, Sharon; Philbey, Adrian; Philbey, Adrain; Thrasher, Adrian J; Thrasher, Adrain J; Cameron, Ewan R; Blyth, Karen; Neil, James C

    2009-06-01

    The emergence of leukemia following gene transfer to restore common cytokine receptor gamma chain (gammaC) function in X-linked severe combined immunodeficiency (SCID-X1) has raised important questions with respect to gene therapy safety. To explore the risk factors involved, we tested the oncogenic potential of human gammaC in new strains of transgenic mice expressing the gene under the control of the CD2 promoter and locus control region (LCR). These mice demonstrated mildly perturbed T-cell development, with an increased proportion of thymic CD8 cells, but showed no predisposition to tumor development even on highly tumor prone backgrounds or after gamma-retrovirus infection. The human CD2-gammaC transgene rescued T and B-cell development in gammaC(-/-) mice but with an age-related delay, mimicking postnatal reconstitution in SCID-X1 gene therapy subjects. However, we noted that gammaC(-/-) mice are acutely susceptible to murine leukemia virus (MLV) leukemogenesis, and that this trait was not corrected by the gammaC transgene. We conclude that the SCID-X1 phenotype can be corrected safely by stable ectopic expression of gammaC and that the transgene is not significantly oncogenic when expressed in this context. However, an underlying predisposition conferred by the SCID-X1 background appears to collaborate with insertional mutagenesis to increase the risk of tumor development.

  12. Ectopic lymphokine gene expression in human peripheral blood lymphocytes in vivo

    SciTech Connect

    Chambers, C.A.; Kang, Joonsoo; Hozumi, Nobumichi Mount Sinai Hospital, Toronto, Ontario )

    1992-02-01

    An animal model to study the effects of ectopic expression of cytokines involved in cell growth and differentiation has been established. Retrovirus vectors containing the human interleukin 6 cDNA were used to produce high titer virus-producing lines. Human peripheral blood lymphocytes (hPBLs) were successfully infected with the retrovirus and engrafted into severe combined immunodeficient mice. The majority of the animals were engrafted with hPBLs, as determined by the presence of human glucose phosphate isomerase. Furthermore, six of seven mice engrafted with hPBLs infected with high titer virus and detectable hPBLs present in the spleen expressed the retroviral human interleukin 6 gene. Importantly, human interleukin 6 protein was expressed at physiologically significant levels in these mice. These results demonstrate that models for human disease and immunotherapy involving retrovirus-mediated gene transfer into human cells can be developed in mice.

  13. Contrastive prevalence of feline retrovirus infections between northern and southern Vietnam.

    PubMed

    Nakamura, K; Miyazawa, T; Ikeda, Y; Sato, E; Nishimura, Y; Nguyen, N T; Takahashi, E; Mochizuki, M; Mikami, T

    2000-08-01

    The prevalence of infections with three feline retroviruses; feline leukemia virus (FeLV), feline immunodeficiency virus (FIV) and feline foamy virus (FeFV), was examined in domestic cats (Felis catus) and leopard cats (Felis bengalensis) in southern Vietnam in 1998. We then compared this data with our previous study in northern Vietnam in 1997. None of the cats had FeLV antigens in both the northern and southern areas. In contrast, there is a great distinction in the seropositivity of FIV. Twenty-two percent of domestic cats had FIV antibodies whereas no FIV positive cats were detected in northern area. FIV may have entered southern Vietnam recently and spread rapidly. FeFV infections were found in both areas, suggesting that FeFV might be present in the cat populations in Vietnam from the earliest time.

  14. 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

  15. The sequence of carnation etched ring virus DNA: comparison with cauliflower mosaic virus and retroviruses

    PubMed Central

    Hull, R.; Sadler, J.; Longstaff, M.

    1986-01-01

    Carnation etched ring virus (CERV) DNA comprises 7932 bp. CERV primer binding sites and overall genome organization are similar to those of the related cauliflower mosaic virus (CaMV). The six open reading frames of CERV showed amino acid homology (50-80%) with CaMV ORFs I-VI; no homologues of CaMV ORFs VII or VIII were found. CERV ORFs 1-5 interface each other with the sequence ATGA. The comparison of CERV ORF5 with CaMV ORFV highlighted regions which show homologies to retrovirus gag/pol protease, RNase H and DNA polymerase domains; the possibility that the DNA polymerase domain comprises two subdomains, operating off different templates, is discussed. Both CERV and CaMV ORFs I have sequence homology to tobacco mosaic virus P30 and plastocyanin. PMID:16453731

  16. Identification of cellular factors required for the budding of koala retrovirus.

    PubMed

    Shimode, Sayumi; Nakaoka, Rie; Hoshino, Shigeki; Abe, Masumi; Shogen, Hiroko; Yasuda, Jiro; Miyazawa, Takayuki

    2013-07-01

    Koala retrovirus (KoRV) is a unique gammaretrovirus that is currently endogenizing into its host and considered to be associated with leukemia, lymphoma and immunosuppression in koalas (Phascolactos cinereus). In this study, it was demonstrated that WWP2 or WWP2-like E3 ubiquitin ligases possessing the WW domain closely related to WWP2 and Vps4A/B are involved in KoRV budding. These data suggest that KoRV Gag recruits the cellular endosomal sorting complex required for transport machinery through interaction of the PPPY L-domain with the WW domain(s) of WWP2 and that progeny virions are released from cells by utilizing the multivesicular body sorting pathway.

  17. [Pathological Diagnoses and Whole-genome Sequence Analyses of the Jaagsiekte Sheep Retrovirus in Xinjiang, China].

    PubMed

    Yang, Sufang; Liang, Tian; Zhao, Qingliang; Zhang, Dianqing; Si Junqiang; Zhang, Jing; Yang, Xia; Sheng, Jinliang

    2015-05-01

    To carry out pathologic diagnoses and whole-genome sequence analyses of the Jaagsiekte sheep retrovirus (JSRV) in Xinjiang, China, we first observed sheep suspected to have the JSRV. Then, the extracted virus suspension was observed by transmission electron microscopy (TEM). Total RNAs from lungs of JSRV-infected sheep were extracted and reverse-transcribed using a cDNA synthesis kit. Six pairs of primers were designed according to the exogenous reference virus strain (AF105220). Reverse transcription-polymerase chain reaction was carried out from JSRV-infected tissue, and the whole genome of the JSRV sequenced. Our results showed: flow of nasal fluid ("wheelbarrow test"); different sizes of adenoma lesions in the lungs; papillary hyperplasia of alveolar epithelial cells; alveolar cavity filled with macrophages; dissolute nuclei in central lesions. TEM revealed JSRV particles with a diameter of 88 nm to 125. 4 nm. The full-length of the viral genome sequence was 7456 bp. BLAST analyses showed nucleotide homology of 96% and 95% compared with that of the representative strain from the USA (AF105220) and UK (AF357971). Nucleotide homology was 89.8% and 89.9% compared with the endogenous Jaagsiekte sheep retrovirus, Inner Mongolia strain (DQ838493) and USA strain (EF680300). The specific pathogenic amino-acid sequence "YXXM" was found in the TM district, similar to the exogenous JSRV: this gene has been reported to be oncogenic. This is the first report of the complete genomic sequence of the exogenous JSRV from Xinjiang, and could lay the foundation for study of the biological characteristics and pathogenic mechanisms of the pulmonary adenomatosis virus in sheep.

  18. 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

  19. Increased transduction efficiency of primary hematopoietic cells by physical colocalization of retrovirus and target cells.

    PubMed

    Hutchings, M; Moriwaki, K; Dilloo, D; Hoffmann, T; Kimbrough, S; Johnsen, H E; Brenner, M K; Heslop, H E

    1998-06-01

    Efficient gene transfer into hematopoietic stem cells offers a number of potential therapeutic applications. However, the relatively low titer of retroviral supernatants and the requirement for cell division to ensure integration have meant that transduction efficiency has been low. We have modified a flowthrough approach to cell transduction and have been able consistently to increase gene transfer efficiency into human hematopoietic progenitor cells. We transduced CD34 cells with retroviral vectors encoding a truncated nerve growth factor receptor (NGFR) or neo. Retroviral supernatant was pulled through 0.2-micron polycarbonated membranes, followed by placement of cells on the filter. In the absence of cytokines, the transduction efficiency of CD34 cells with a NGFR vector was increased 3-11-fold over that obtained at an identical MOI in liquid culture to produce 11%-44% transduction. Furthermore, both Thy1+ and Thy1- subsets in a total CD34 population were transduced with similar efficiency, and transduction with a neo vector, as measured by G418 resistance in clonogenic assays, increased 1.5-5-fold. The mechanism by which gene transfer is improved may reflect colocalization of cells and retrovirus. Costaining of cells transduced on the filter with an NGFR retrovirus with both an NGFR antibody and a gp70 antibody that recognizes viral coat protein revealed high-level coexpression. The levels of in vitro gene transfer we obtain are equivalent to those observed when CD34 cells are cocultured in liquid culture with cytokines. However, culture with cytokines may commit CD34 cells to differentiation and has produced disappointingly low levels of subsequent in vivo gene transfer. Gene marking studies using distinguishable retroviral vectors will provide a means of learning whether the effects of flowthrough transduction genuinely enhance the efficiency of gene transfer to human marrow-repopulating cells.

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

    PubMed

    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

    2016-02-14

    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.

  1. Partial molecular cloning of the JHK retrovirus using gammaretrovirus consensus PCR primers

    PubMed Central

    Halligan, Brian D; Sun, Hai-Yuan; Kushnaryov, Vladimir M; Grossberg, Sidney E

    2013-01-01

    The JHK virus (JHKV) was previously described as a type C retrovirus that has some distinctive ultrastructural features and replicates constitutively in a human B-lymphoblastoid cell line, JHK-3. In order to facilitate the cloning of sequences from JHKV, a series of partially degenerate consensus retroviral PCR primers were created by a data-driven design approach based on an alignment of 14 diverse gammaretroviral genomes. These primers were used in the PCR amplification of purified JHK virion cDNA, and ana lysis of the resulting amplified sequence indicates that the JHKV is in the murine leukemia virus (MLV) family. The JHK sequence is nearly identical to the corresponding region of the Bxv-1 endogenous mouse retrovirus (GenBank accession AC115959) and distinct from XMRV. JHKV gag-specific amplification was demonstrated with nucleic acids from uncultivated, frozen, peripheral blood mononuclear cells (PBMCs) of the index patient, but not in PBMCs from nine healthy blood donors. Unlike earlier reports, in which MLV-like sequences were identified in human source material, which may have been due to murine contamination, budding retrovirions were demonstrated repeatedly by electron microscopy in uncultivated lymphocytes of the index patient that were morphologically identical in their development to the virions in the JHK-3 cells, and immunological evidence was obtained that the index patient produced IgG antibodies that bound to the budding viral particles in patient PBMCs and in the JHK-3 cells. These data indicate that the patient had been infected by JHKV, lending significance to the demonstration of JHKV amplicons in nucleic acids of the patient’s PBMCs. In future studies, the PCR primer sets described herein may expand the detection of an amplifiable subset of viruses related to MLV. PMID:24159361

  2. 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

  3. Early Steps of Jaagsiekte Sheep Retrovirus-Mediated Cell Transformation Involve the Interaction between Env and the RALBP1 Cellular Protein

    PubMed Central

    Monot, Margaux; Erny, Alexandra; Gineys, Barbara; Desloire, Sophie; Dolmazon, Christine; Aublin-Gex, Anne; Lotteau, Vincent; Archer, Fabienne

    2015-01-01

    ABSTRACT Ovine pulmonary adenocarcinoma is a naturally occurring lung cancer in sheep induced by the Jaagsiekte sheep retrovirus (JSRV). Its envelope glycoprotein (Env) carries oncogenic properties, and its expression is sufficient to induce in vitro cell transformation and in vivo lung adenocarcinoma. The identification of cellular partners of the JSRV envelope remains crucial for deciphering mechanisms leading to cell transformation. We initially identified RALBP1 (RalA binding protein 1; also known as RLIP76 or RIP), a cellular protein implicated in the ras pathway, as a partner of JSRV Env by yeast two-hybrid screening and confirmed formation of RALBP1/Env complexes in mammalian cells. Expression of the RALBP1 protein was repressed in tumoral lungs and in tumor-derived alveolar type II cells. Through its inhibition using specific small interfering RNA (siRNA), we showed that RALBP1 was involved in envelope-induced cell transformation and in modulation of the mTOR (mammalian target of rapamycin)/p70S6K pathway by the retroviral envelope. IMPORTANCE JSRV-induced lung adenocarcinoma is of importance for the sheep industry. While the envelope has been reported as the oncogenic determinant of the virus, the cellular proteins directly interacting with Env are still not known. Our report on the formation of RALBP/Env complexes and the role of this interaction in cell transformation opens up a new hypothesis for the dysregulation observed upon virus infection in sheep. PMID:26041289

  4. Human endogenous retrovirus W family envelope gene activates the small conductance Ca2+-activated K+ channel in human neuroblastoma cells through CREB.

    PubMed

    Li, S; Liu, Z C; Yin, S J; Chen, Y T; Yu, H L; Zeng, J; Zhang, Q; Zhu, F

    2013-09-05

    Numerous studies have shown that human endogenous retrovirus W family (HERV-W) envelope gene (env) is related to various diseases but the underlying mechanism has remained poorly understood. Our previous study showed that there was abnormal expression of HERV-W env in sera of patients with schizophrenia. In this paper, we reported that overexpression of the HERV-W env elevated the levels of small conductance Ca(2+)-activated K(+) channel protein 3 (SK3) in human neuroblastoma cells. Using a luciferase reporter system and RNA interference method, we found that functional cAMP response element site was required for the expression of SK3 triggered by HERV-W env. In addition, it was also found that the SK3 channel was activated by HERV-W env. Further study indicated that cAMP response element-binding protein (CREB) was required for the activation of the SK3 channel. Thus, a novel signaling mechanism of how HERV-W env influences neuronal activity and contributes to mental illnesses such as schizophrenia was proposed.

  5. The ecology of primate retroviruses – An assessment of 12 years of retroviral studies in the Taï national park area, Côte d'Ivoire

    PubMed Central

    Gogarten, Jan F.; Akoua-Koffi, Chantal; Calvignac-Spencer, Sebastien; Leendertz, Siv Aina J.; Weiss, Sabrina; Couacy-Hymann, Emmanuel; Koné, Inza; Peeters, Martine; Wittig, Roman M.; Boesch, Christophe; Hahn, Beatrice H.; Leendertz, Fabian H.

    2014-01-01

    The existence and genetic make-up of most primate retroviruses was revealed by studies of bushmeat and fecal samples from unhabituated primate communities. For these, detailed data on intra- and within-species contact rates are generally missing, which makes identification of factors influencing transmission a challenging task. Here we present an assessment of 12 years of research on primate retroviruses in the Taï National Park area, Côte d’Ivoire. We discuss insights gained into the prevalence, within- and cross-species transmission of primate retroviruses (including towards local human populations) and the importance of virus–host interactions in determining cross-species transmission risk. Finally we discuss how retroviruses ecology and evolution may change in a shifting environment and identify avenues for future research. PMID:25010280

  6. The ecology of primate retroviruses - an assessment of 12 years of retroviral studies in the Taï national park area, Côte d׳Ivoire.

    PubMed

    Gogarten, Jan F; Akoua-Koffi, Chantal; Calvignac-Spencer, Sebastien; Leendertz, Siv Aina J; Weiss, Sabrina; Couacy-Hymann, Emmanuel; Koné, Inza; Peeters, Martine; Wittig, Roman M; Boesch, Christophe; Hahn, Beatrice H; Leendertz, Fabian H

    2014-07-01

    The existence and genetic make-up of most primate retroviruses was revealed by studies of bushmeat and fecal samples from unhabituated primate communities. For these, detailed data on intra- and within-species contact rates are generally missing, which makes identification of factors influencing transmission a challenging task. Here we present an assessment of 12 years of research on primate retroviruses in the Taï National Park area, Côte d'Ivoire. We discuss insights gained into the prevalence, within- and cross-species transmission of primate retroviruses (including towards local human populations) and the importance of virus-host interactions in determining cross-species transmission risk. Finally we discuss how retroviruses ecology and evolution may change in a shifting environment and identify avenues for future research.

  7. Identification and classification of feline endogenous retroviruses in the cat genome using degenerate PCR and in silico data analysis.

    PubMed

    Song, Ning; Jo, Haiin; Choi, Minkyeung; Kim, Jin-Hoi; Seo, Han Geuk; Cha, Se-Yeoun; Seo, Kunho; Park, Chankyu

    2013-07-01

    The purpose of this study was to identify and classify endogenous retroviruses (ERVs) in the cat genome. Pooled DNA from five domestic cats was subjected to degenerate PCR with primers specific to the conserved retroviral pro/pol region. The 59 amplified retroviral sequences were used for in silico analysis of the cat genome (Felis_catus-6.2). We identified 219 ERV γ and β elements from cat genome contigs, which were classified into 42 ERV γ and 4 β families and further analysed. Among them, 99 γ and 5 β ERV elements contained the complete retroviral structure. Furthermore, we identified 757 spuma-like ERV elements based on the sequence homology to murine (Mu)ERV-L and human (H)ERV-L. To the best of our knowledge, this is the first detailed genome-scale analysis examining Felis catus endogenous retroviruses (FcERV) and providing advanced insights into their structural characteristics, localization in the genome, and diversity.

  8. 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.

  9. Retrovirus Restriction by TRIM5 Proteins Requires Recognition of Only a Small Fraction of Viral Capsid Subunits

    PubMed Central

    Shi, Jiong; Friedman, David B.

    2013-01-01

    The host restriction factors TRIM5α and TRIMCyp potently inhibit retrovirus infection by binding to the incoming retrovirus capsid. TRIM5 proteins are dimeric, and their association with the viral capsid appears to be enhanced by avidity effects owing to formation of higher-order oligomeric complexes. We examined the stoichiometric requirement for TRIM5 functional recognition by quantifying the efficiencies of restriction of HIV-1 and murine leukemia virus (MLV) particles containing various proportions of restriction-sensitive and -insensitive CA subunits. Both TRIMCyp and TRIM5α inhibited infection of retrovirus particles containing as little as 25% of the restriction-sensitive CA protein. Accordingly, we also observed efficient binding of TRIMCyp in vitro to capsid assemblies containing as little as one-fourth wild-type CA protein. Paradoxically, the ability of HIV-1 particles to abrogate TRIMCyp restriction in trans was more strongly dependent on the fraction of wild-type CA than was restriction of infection. Collectively, our results indicate that TRIM5 restriction factors bind to retroviral capsids in a highly cooperative manner and suggest that TRIM5 can engage a capsid lattice containing a minimum of three or fewer recognizable subunits per hexamer. Our study supports a model in which localized binding of TRIM5 to the viral capsid nucleates rapid polymerization of a TRIM5 lattice on the capsid surface. PMID:23785198

  10. Clinical Potential of the Acyclic Nucleoside Phosphonates Cidofovir, Adefovir, and Tenofovir in Treatment of DNA Virus and Retrovirus Infections

    PubMed Central

    De Clercq, Erik

    2003-01-01

    The acyclic nucleoside phosphonates HPMPC (cidofovir), PMEA (adefovir), and PMPA (tenofovir) have proved to be effective in vitro (cell culture systems) and in vivo (animal models and clinical studies) against a wide variety of DNA virus and retrovirus infections: cidofovir against herpesvirus (herpes simplex virus types 1 and 2 varicella-zoster virus, cytomegalovirus [CMV], Epstein-Barr virus, and human herpesviruses 6, 7, and 8), polyomavirus, papillomavirus, adenovirus, and poxvirus (variola virus, cowpox virus, vaccinia virus, molluscum contagiosum virus, and orf virus) infections; adefovir against herpesvirus, hepadnavirus (human hepatitis B virus), and retrovirus (human immunodeficiency virus types 1 [HIV-1] and 2 [HIV-2], simian immunodeficiency virus, and feline immunodeficiency virus) infections; and tenofovir against both hepadnavirus and retrovirus infections. Cidofovir (Vistide) has been officially approved for the treatment of CMV retinitis in AIDS patients, tenofovir disoproxil fumarate (Viread) has been approved for the treatment of HIV infections (i.e., AIDS), and adefovir dipivoxil (Hepsera) has been approved for the treatment of chronic hepatitis B. Nephrotoxicity is the dose-limiting side effect for cidofovir (Vistide) when used intravenously (5 mg/kg); no toxic side effects have been described for adefovir dipivoxil and tenofovir disoproxil fumarate, at the approved doses (Hepsera at 10 mg orally daily and Viread at 300 mg orally daily). PMID:14557287

  11. Infectious Entry Pathway Mediated by the Human Endogenous Retrovirus K Envelope Protein

    PubMed Central

    Robinson, Lindsey R.

    2016-01-01

    ABSTRACT Endogenous retroviruses (ERVs), the majority of which exist as degraded remnants of ancient viruses, comprise approximately 8% of the human genome. The youngest human ERVs (HERVs) belong to the HERV-K(HML-2) subgroup and were endogenized within the past 1 million years. The viral envelope protein (ENV) facilitates the earliest events of endogenization (cellular attachment and entry), and here, we characterize the requirements for HERV-K ENV to mediate infectious cell entry. Cell-cell fusion assays indicate that a minimum of two events are required for fusion, proteolytic processing by furin-like proteases and exposure to acidic pH. We generated an infectious autonomously replicating recombinant vesicular stomatitis virus (VSV) in which the glycoprotein was replaced by HERV-K ENV. HERV-K ENV imparts an endocytic entry pathway that requires dynamin-mediated membrane scission and endosomal acidification but is distinct from clathrin-dependent or macropinocytic uptake pathways. The lack of impediments to the replication of the VSV core in eukaryotic cells allowed us to broadly survey the HERV-K ENV-dictated tropism. Unlike extant betaretroviral envelopes, which impart a narrow species tropism, we found that HERV-K ENV mediates broad tropism encompassing cells from multiple mammalian and nonmammalian species. We conclude that HERV-K ENV dictates an evolutionarily conserved entry pathway and that the restriction of HERV-K to primate genomes reflects downstream stages of the viral replication cycle. IMPORTANCE Approximately 8% of the human genome is of retroviral origin. While many of those viral genomes have become inactivated, some copies of the most recently endogenized human retrovirus, HERV-K, can encode individual functional proteins. Here, we characterize the envelope protein (ENV) of the virus to define how it mediates infection of cells. We demonstrate that HERV-K ENV undergoes a proteolytic processing step and triggers membrane fusion in response to

  12. Modulation of ecotropic murine retroviruses by N-linked glycosylation of the cell surface receptor/amino acid transporter.

    PubMed Central

    Wang, H; Klamo, E; Kuhmann, S E; Kozak, S L; Kavanaugh, M P; Kabat, D

    1996-01-01

    The cell surface receptor for ecotropic host-range (infection limited to mice or rats) murine leukemia viruses (MuLVs) is the widely expressed system y+ transporter for cationic amino acids (CAT-1). Like other retroviruses, ecotropic MuLV infection eliminates virus-binding sites from cell surfaces and results in complete interference to superinfection. Surprisingly, infection causes only partial (ca 40 to 60%) loss of mouse CAT-1 transporter activity. The NIH/Swiss mouse CAT-1 (mCAT-1) contains 622 amino acids with 14 hydrophobic potential membrane-spanning sequences, and it is known that the third extracellular loop from the amino terminus is required for virus binding. Although loop 3 is hypervariable in different species and mouse strains, consistent with its proposed role in virus-host coevolution, loop 3 sequences of both susceptible and resistant species contain consensus sites for N-linked glycosylation. Both of the consensus sites in loop 3 of mCAT-1 are known to be glycosylated and to contain oligosaccharides with diverse sizes (J. W. Kim and J. M. Cunningham, J. Biol. Chem. 268:16316-16320, 1993). We confirmed by several lines of evidence that N-linked glycosylation occludes a potentially functional virus-binding site in the CAT-1 protein of hamsters, thus contributing to resistance of that species. To study the role of receptor glycosylation in animals susceptible to infection, we eliminated loop 3 glycosylation sites by mutagenesis of an mCAT-1 cDNA clone, and we expressed wild-type and mutant receptors in mink fibroblasts and Xenopus oocytes. These receptors had indistinguishable transport properties, as determined by kinetic and voltage-jump electrophysiological studies of arginine uptake in oocytes and by analyses Of L-[3H]arginine uptake in mink cells. Bindings of ecotropic envelope glycoprotein gp7O to the accessible receptor sites on surfaces of mink cells expressing wild-type or mutant mCAT-1 were not significantly different in kinetics or in

  13. Structural and biochemical studies of ALIX/AIP1 and its role in retrovirus budding.

    PubMed

    Fisher, Robert D; Chung, Hyo-Young; Zhai, Qianting; Robinson, Howard; Sundquist, Wesley I; Hill, Christopher P

    2007-03-09

    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 YPX(n)L 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. YPX(n)L late domains bind 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.

  14. Restriction of feline retroviruses: lessons from cat APOBEC3 cytidine deaminases and TRIM5alpha proteins.

    PubMed

    Münk, Carsten; Hechler, Torsten; Chareza, Sarah; Löchelt, Martin

    2010-03-15

    The interplay between viral and cellular factors determines the outcome of an initial contact between a given virus and its natural host or upon encounter of a novel host. Thus, the potential of inducing disease as well as crossing host species barriers are the consequences of the molecular interactions between the parasite and its susceptible, tolerant or resistant host. Cellular restriction factors, for instance APOBEC3 and TRIM5 proteins, targeting defined pathogens or groups of pathogens as well as viral genes counter-acting these cellular defense systems are of prime importance in this respect and may even represent novel targets for prevention and therapy of virus infections. Due to the importance of host-encoded antiviral restriction and viral counter-defense for pathogenicity and host tropism, the responsible molecular factors and mechanisms are currently under intense investigation. In this review we will introduce host restriction and retroviral counter-defense systems with a special emphasis on the cat and its naturally occurring exogenous retroviruses which is a valid model for human disease, a model that will contribute to increase our basic understanding and potential applications of these important aspects of host-virus interaction.

  15. Cells infected with Jaagsiekte sheep retrovirus are detected in the bone marrow of asymptomatic sheep.

    PubMed

    Borobia, Marta; Ortín, Aurora; Ferrer, Luis M; Ramos, Juán J; Lacasta, Delia; De Las Heras, Marcelo

    2014-07-01

    Ovine pulmonary adenocarcinoma (OPA) is a transmissible lung cancer caused by Jaggsiekte sheep retrovirus (JSRV). It is difficult to identify animals infected with JSRV but are clinically healthy. The virus does not induce a specific antibody response and, although proviral DNA sequences of JSRV can be found in mononuclear blood cells, the detection is inconsistent. The aim of this study was to investigate the presence of JSRV in the bone marrow of infected sheep and develop a more consistent screening method. Immunohistochemical examination of bone marrow samples from 8 asymptomatic JSRV-infected sheep revealed the presence of positively labelled cells. However, JSRV could not be detected by a highly sensitive polymerase chain reaction (PCR) in bone marrow aspirates periodically collected from these animals. Results suggest that JSRV-infected cells may be present in the bone marrow of symptomless animals, but the number is below the detectable level for PCR. Therefore, this technique does not seem to be helpful for preclinical diagnosis of OPA.

  16. Two Closely Related but Distinct Retroviruses Are Associated with Walleye Discrete Epidermal Hyperplasia

    PubMed Central

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

    1998-01-01

    Walleye discrete epidermal hyperplasia (WEH) is a hyperproliferative skin disease that is prevalent on adult walleye fish throughout North America. We have identified two retroviruses associated with WEH, designated here as walleye epidermal hyperplasia virus type 1 and type 2 (WEHV1 and WEHV2), that are closely related to one another (77% identity) and to walleye dermal sarcoma virus (64% identity) within the polymerase region. WEHV1 and/or WEHV2 viral DNA was readily detected by PCR in hyperplastic tissue samples, but only low levels of viral DNA were detected in uninvolved skin. Southern blot analysis showed one to three copies of integrated WEHV2 viral DNA in lesions but did not detect WEHV2 viral DNA in uninvolved skin from the same fish. Northern blots detected abundant levels of WEHV1 and/or WEHV2 virion RNA transcripts of approximately 13 kb in hyperplastic tissue, but virion RNA was not observed in uninvolved skin and muscle. These results suggest that WEHV1 and WEHV2 are the causative agents of discrete epidermal hyperplasia. PMID:9525688

  17. Human retroviruses and AIDS 1996. A compilation and analysis of nucleic acid and amino acid sequences

    SciTech Connect

    Myers, G.; Foley, B.; Korber, B.; Mellors, J.W.; Jeang, K.T.; Wain-Hobson, S.

    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. 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.

  18. Clonal fluctuation within the haematopoietic system of mice reconstituted with retrovirus-infected stem cells.

    PubMed Central

    Snodgrass, R; Keller, G

    1987-01-01

    The clonal make-up of the haematopoietic system of mice reconstituted with retrovirus-infected bone marrow cells was analysed at two different points in time following reconstitution. We have found that under these conditions, the haematopoietic system consists of clones that persist throughout the 5 month course of the experiment as well as those which undergo temporal changes. The various changes that we have observed included the appearance of a new clone(s) in all lineages, the loss of a clone from some lineages and the shift in the appearance of a clone from one lineage to another. In addition, we provide evidence which suggests that the clonal make-up of the thymus changes with time; early after reconstitution it consists of many clones, whereas at the later time-points it contains a limited number of predominant clones. These studies document the dramatic clonal changes which occur within the various lineages for a long time following reconstitution and highlight the difficulty in demonstrating lineage-specific stem cells. Images Fig. 2. Fig. 3. Fig. 4. PMID:2832146

  19. Structural and Biochemical Studies of ALIX/AlP1 and Its Role in Retrovirus Budding

    SciTech Connect

    Fisher,R.; Chung, H.; Zhai, Q.; Robinson, H.; Sundquist, W.; Hill, C.

    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 bind 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.

  20. Construction and characterization of an infectious molecular clone of Koala retrovirus.

    PubMed

    Shojima, Takayuki; Hoshino, Shigeki; Abe, Masumi; Yasuda, Jiro; Shogen, Hiroko; Kobayashi, Takeshi; Miyazawa, Takayuki

    2013-05-01

    Koala retrovirus (KoRV) is a gammaretrovirus that is currently endogenizing into koalas. Studies on KoRV infection have been hampered by the lack of a replication-competent molecular clone. In this study, we constructed an infectious molecular clone, termed plasmid pKoRV522, of a KoRV isolate (strain Aki) from a koala reared in a Japanese zoo. The virus KoRV522, derived from pKoRV522, grew efficiently in human embryonic kidney (HEK293T) cells, attaining 10(6) focus-forming units/ml. Several mutations in the Gag (L domain) and Env regions reported to be involved in reduction in viral infection/production in vitro are found in pKoRV522, yet KoRV522 replicated well, suggesting that any effects of these mutations are limited. Indeed, a reporter virus pseudotyped with pKoRV522 Env was found to infect human, feline, and mink cell lines efficiently. Analyses of KoRV L-domain mutants showed that an additional PPXY sequence, PPPY, in Gag plays a critical role in KoRV budding. Altogether, our results demonstrate the construction and characterization of the first infectious molecular clone of KoRV. The infectious clone reported here will be useful for elucidating the mechanism of endogenization of the virus in koalas and screening for antiretroviral drugs for KoRV-infected koalas.

  1. 12th international conference on human retrovirology: HTLV and related retroviruses

    PubMed Central

    Lairmore, Michael D; Fujii, Masahiro

    2005-01-01

    The 12th International Conference on Human Retrovirology: HTLV and Related Retroviruses, was held at the Half Moon Hotel in Montego Bay, Jamaica, from June 22nd to June 25th 2005. The scientific conference, sponsored by the International Retrovirology Association, is held biennially at rotating international venues around the world. The meeting brings together basic scientists, epidemiologists and clinical researchers to discuss findings to prevent HTLV infection or develop new therapies against HTLV-mediated diseases. The Association fosters the education and training of young scientists to bring new approaches to the complex problems of HTLV research, such as translational research to bring findings from the laboratory into clinical trials that benefit HTLV-infected patients. The breadth and quality of research presentations and workshops at the 12th International Conference indicate that these goals are being accomplished. As HTLV research enters its third decade a new generation of scientists face many challenges. However, HTLV scientists and clinicians displayed exciting new approaches and discoveries during plenary talks and poster sessions. The conference encouraged research in HTLV infections and disease, fostered collaborations, and stimulated new partnerships between clinicians and scientists to encourage clinical trials and novel therapeutic interventions. PMID:16202161

  2. Evolution of endogenous retrovirus-like elements of the woolly mammoth (Mammuthus primigenius) and its relatives.

    PubMed

    Greenwood, A D; Lee, F; Capelli, C; DeSalle, R; Tikhonov, A; Marx, P A; MacPhee, R D

    2001-05-01

    Endogenous retrovirus-like elements characterizable by a leucine tRNA primer (ERV-Ls) are reiterated genomic sequences known to be widespread in mammals, including humans. They may have arisen from an ancestral foamy virus-like element by successful germ line infection followed by copy number expansion. However, among mammals, only primates and rodents have thus far exhibited high copy number amplification and sequence diversification. Conventionally, empirical studies of proviral amplification and diversification have been limited to extant species, but taxa having good Quaternary fossil records could potentially be investigated using the techniques of "ancient" DNA research. To examine evolutionary parameters of ERV-Ls across both time and taxa, we characterized this proviral class in the extinct woolly mammoth (Mammuthus primigenius) and living elephants, as well as extant members of the larger clade to which they belong (Uranotheria, a group containing proboscideans, sirenians, hyraxes, and their extinct relatives). Ungulates and carnivores previously analyzed demonstrated low copy numbers of ERV-L sequences, and thus it was expected that uranotheres should as well. Here, we show that all uranothere taxa exhibit unexpectedly numerous and diverse ERV-L sequence complements, indicating active expansion within this group of lineages. Selection is the most parsimonious explanation for observed differences in ERV-L distribution and frequency, with relative success being reflected in the persistence of certain elements over a variety of sampled time depths (as can be observed by comparing sequences from fossil and extant elephantid samples).

  3. 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

  4. 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.

  5. The role of genes domesticated from LTR retrotransposons and retroviruses in mammals

    PubMed Central

    Kaneko-Ishino, Tomoko; Ishino, Fumitoshi

    2012-01-01

    The acquisition of multiple genes from long terminal repeat (LTR) retrotransposons occurred in mammals. Genes belonging to a sushi-ichi-related retrotransposon homologs (SIRH) family emerged around the time of the establishment of two viviparous mammalian groups, marsupials and eutherians. These genes encode proteins that are homologous to a retrotransposon Gag capsid protein and sometimes also have a Pol-like region. We previously demonstrated that PEG10 (SIRH1) and PEG11/RTL1 (SIRH2) play essential but different roles in placental development. PEG10 is conserved in both the marsupials and the eutherians, while PEG11/RTL1 is a eutherian-specific gene, suggesting that these two domesticated genes were deeply involved in the evolution of mammals via the establishment of the viviparous reproduction system. In this review, we introduce the roles of PEG10 and PEG11/RTL1 in mammalian development and evolution, and summarize the other genes domesticated from LTR retrotransposons and endogenous retroviruses (ERVs) in mammals. We also point out the importance of DNA methylation in inactivating and neutralizing the integrated retrotransposons and ERVs in the process of domestication. PMID:22866050

  6. The role of genes domesticated from LTR retrotransposons and retroviruses in mammals.

    PubMed

    Kaneko-Ishino, Tomoko; Ishino, Fumitoshi

    2012-01-01

    The acquisition of multiple genes from long terminal repeat (LTR) retrotransposons occurred in mammals. Genes belonging to a sushi-ichi-related retrotransposon homologs (SIRH) family emerged around the time of the establishment of two viviparous mammalian groups, marsupials and eutherians. These genes encode proteins that are homologous to a retrotransposon Gag capsid protein and sometimes also have a Pol-like region. We previously demonstrated that PEG10 (SIRH1) and PEG11/RTL1 (SIRH2) play essential but different roles in placental development. PEG10 is conserved in both the marsupials and the eutherians, while PEG11/RTL1 is a eutherian-specific gene, suggesting that these two domesticated genes were deeply involved in the evolution of mammals via the establishment of the viviparous reproduction system. In this review, we introduce the roles of PEG10 and PEG11/RTL1 in mammalian development and evolution, and summarize the other genes domesticated from LTR retrotransposons and endogenous retroviruses (ERVs) in mammals. We also point out the importance of DNA methylation in inactivating and neutralizing the integrated retrotransposons and ERVs in the process of domestication.

  7. Human Endogenous Retrovirus Protein Activates Innate Immunity and Promotes Experimental Allergic Encephalomyelitis in Mice

    PubMed Central

    Perron, Hervé; Dougier-Reynaud, Hei-Lanne; Lomparski, Christina; Popa, Iuliana; Firouzi, Reza; Bertrand, Jean-Baptiste; Marusic, Suzana; Portoukalian, Jacques; Jouvin-Marche, Evelyne; Villiers, Christian L.; Touraine, Jean-Louis; Marche, Patrice N.

    2013-01-01

    Multiple sclerosis (MS) is a complex multifactorial disease of the central nervous system (CNS) for which animal models have mainly addressed downstream immunopathology but not potential inducers of autoimmunity. In the absence of a pathogen known to cause neuroinflammation in MS, Mycobacterial lysate is commonly used in the form of complete Freund's adjuvant to induce autoimmunity to myelin proteins in Experimental Allergic Encephalomyelitis (EAE), an animal model for MS. The present study demonstrates that a protein from the human endogenous retrovirus HERV-W family (MSRV-Env) can be used instead of mycobacterial lysate to induce autoimmunity and EAE in mice injected with MOG, with typical anti-myelin response and CNS lesions normally seen in this model. MSRV-Env was shown to induce proinflammatory response in human macrophage cells through TLR4 activation pathway. The present results demonstrate a similar activation of murine dendritic cells and show the ability of MSRV-Env to trigger EAE in mice. In previous studies, MSRV-Env protein was reproducibly detected in MS brain lesions within microglia and perivascular macrophages. The present results are therefore likely to provide a model for MS, in which the upstream adjuvant triggering neuroinflammation is the one detected in MS active lesions. This model now allows pre-clinical studies with therapeutic agents targeting this endogenous retroviral protein in MS. PMID:24324591

  8. 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).

  9. Computer simulation of a cellular automata model for the immune response in a retrovirus system

    SciTech Connect

    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/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/).

  10. Human retroviruses and aids, 1992. A compilation and analysis of nucleic acid and amino acid sequences

    SciTech Connect

    Myers, G.; Korber, B.; Berzofsky, J.A.; Pavlakis, G.N.; Smith, R.F.

    1992-10-01

    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; (H) 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. 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. In addition to the general descriptions below of the parts of the compendium, the user should read the individual introductions for each part.

  11. Evidence for retrovirus infections in green turtles Chelonia mydas from the Hawaiian islands

    USGS Publications Warehouse

    Casey, R.N.; Quackenbush, S.L.; Work, T.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).

  12. Detection of avian retroviruses in vaccines by amplification on DF-1 cells with immunostaining and fluorescent product-enhanced reverse transcriptase endpoint methods.

    PubMed

    Birmingham, Cheryl L; Dupont, Dominique; Riou, Patrice; Armanet, Corinne; Edamura, Kerrie Nichol; Martinho, Briolange; Serres, Aurelie; Jacouton, Severine; Detrez, Valerie; McNeil, Bryan; Schreiber, Martha; Gaillac, David; Bonnevay, Thierry; Gisonni-Lex, Lucy; Mallet, Laurent

    2013-05-01

    In order to ensure the safety of vaccines produced on avian cells, rigorous testing for the absence of avian retroviruses must be performed. Current methods used to detect avian retroviruses often exhibit a high invalid-test/false-positive rate, rely on hard-to-secure reagents, and/or have readouts that are difficult to standardize. Herein, we describe the development and validation of two consistent and sensitive methods for the detection of avian retroviruses in vaccines: viral amplification on DF-1 cells followed by immunostaining for the detection of avian leukosis virus (ALV) and viral amplification on DF-1 cells followed by fluorescent product-enhanced reverse transcriptase (F-PERT) for the detection of all avian retroviruses. Both assays share an infectivity stage on DF-1 cells followed by a different endpoint readout depending on the retrovirus to be detected. Validation studies demonstrated a limit of detection of one 50% cell culture infectious dose (CCID(50))/ml for retrovirus in a 30-ml test inoculum volume for both methods, which was as sensitive as a classical method used in the vaccine industry, namely, viral amplification on primary chicken embryo fibroblasts followed by the complement fixation test for avian leukosis virus (COFAL). Furthermore, viral amplification on DF-1 cells followed by either immunostaining or F-PERT demonstrated a sensitivity that exceeds the regulatory requirements for detection of ALV strains. A head-to-head comparison of the two endpoint methods showed that viral amplification on DF-1 cells followed by F-PERT is a suitable method to be used as a stand-alone test to ensure that vaccine preparations are free from infectious avian retroviruses.

  13. 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

  14. Endogenous New World primate retrovirus: interspecies antigenic determinants shared with the major structural protein of type-D RNA viruses of Old World monkeys.

    PubMed Central

    Hino, S; Tronick, S R; Heberling, R L; Kalter, S S; Hellman, A; Aaronson, S A

    1977-01-01

    A reverse transcriptase-containing virus has recently been isolated from a squirrel monkey (Saimiri sciureus). Molecular hybridization studies demonstrate that the squirrel monkey retrovirus (SMRV) is endogenous to this New World primate, yet lacks detectable nucleotide sequence homology with cellular DNAs of representative Old World primates or with the genomes of previously isolated Old World primate retroviruses. The 35,000-dalton major structural protein (p35) of SMRV was purified and shown to possess antigenic determinants distinct from those of known retroviruses. While SMRV was found to lack antigenic determinants broadly shared among mammalian type-C viruses, immunologic crossreactivity was demonstrated between SMRV p35 and the major structural protein (p26) of Mason-Pfizer monkey virus, a prototype type-D retrovirus of Old World monkeys. These findings support the concept that SMRV and Mason-Pfizer monkey virus are evolutionarily related, and raise the possibility that a progenitor of type-D retroviruses became genetically associated with primates at a very early time in their evolution. PMID:74833

  15. Phylogeny-Directed Search for Murine Leukemia Virus-Like Retroviruses in Vertebrate Genomes and in Patients Suffering from Myalgic Encephalomyelitis/Chronic Fatigue Syndrome and Prostate Cancer

    PubMed Central

    Blomberg, Jonas; Sheikholvaezin, Ali; Elfaitouri, Amal; Blomberg, Fredrik; Sjösten, Anna; Mattson Ulfstedt, Johan; Pipkorn, Rüdiger; Källander, Clas; Öhrmalm, Christina; Sperber, Göran

    2011-01-01

    Gammaretrovirus-like sequences occur in most vertebrate genomes. Murine Leukemia Virus (MLV) like retroviruses (MLLVs) are a subset, which may be pathogenic and spread cross-species. Retroviruses highly similar to MLLVs (xenotropic murine retrovirus related virus (XMRV) and Human Mouse retrovirus-like RetroViruses (HMRVs)) reported from patients suffering from prostate cancer (PC) and myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) raise the possibility that also humans have been infected. Structurally intact, potentially infectious MLLVs occur in the genomes of some mammals, especially mouse. Mouse MLLVs contain three major groups. One, MERV G3, contained MLVs and XMRV/HMRV. Its presence in mouse DNA, and the abundance of xenotropic MLVs in biologicals, is a source of false positivity. Theoretically, XMRV/HMRV could be one of several MLLV transspecies infections. MLLV pathobiology and diversity indicate optimal strategies for investigating XMRV/HMRV in humans and raise ethical concerns. The alternatives that XMRV/HMRV may give a hard-to-detect “stealth” infection, or that XMRV/HMRV never reached humans, have to be considered. PMID:22315600

  16. Retroviruses and sexual size dimorphism in domestic cats (Felis catus L.).

    PubMed Central

    Pontier, D; Fromont, E; Courchamp, F; Artois, M; Yoccoz, N G

    1998-01-01

    Hochberg and co-workers have predicted that an increase in host adult mortality due to parasites is balanced by an earlier age at first reproduction. In polygynous species we hypothesize that such a pattern would lead to diverging selection pressure on body size between sexes and increased sexual size dimorphism. In polygynous mammals, male body size is considered to be an important factor for reproductive success. Thus, under the pressure of a virulent infection, males should be selected for rapid growth and/or higher body size to be able to compete successfully as soon as possible with opponents. In contrast, under the same selection pressure, females should be selected for lighter adult body size or rapid growth to reach sexual maturity earlier. We investigated this hypothesis in the domestic cat Felis catus. Orange cats have greater body size dimorphism than non-orange cats. Orange females are lighter than non-orange females, and orange males are heavier than non-orange males. Here, we report the extent to which orange and non-orange individuals differ in infection prevelance for two retroviruses, feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV). FIV is thought to be transmitted almost exclusively through aggressive contacts between individuals, whereas FeLV transmission occurs mainly through social contacts. The pattern of infection of both diseases is consistent with the higher aggressiveness of orange cats. In both sexes, orange cats are significantly more infected by FIV, and tend to be less infected by FeLV than other cats. The pattern of infection is also consistent with an earlier age at first reproduction in orange than in non-orange cats, at least for females. These results suggest that microparasitism may have played an important role in the evolution of sexual size dimorphism of domestic cats. PMID:9493404

  17. Integration and Fixation Preferences of Human and Mouse Endogenous Retroviruses Uncovered with Functional Data Analysis

    PubMed Central

    Pini, Alessia; Chiaromonte, Francesca; Makova, Kateryna D.

    2016-01-01

    Endogenous retroviruses (ERVs), the remnants of retroviral infections in the germ line, occupy ~8% and ~10% of the human and mouse genomes, respectively, and affect their structure, evolution, and function. Yet we still have a limited understanding of how the genomic landscape influences integration and fixation of ERVs. Here we conducted a genome-wide study of the most recently active ERVs in the human and mouse genome. We investigated 826 fixed and 1,065 in vitro HERV-Ks in human, and 1,624 fixed and 242 polymorphic ETns, as well as 3,964 fixed and 1,986 polymorphic IAPs, in mouse. We quantitated >40 human and mouse genomic features (e.g., non-B DNA structure, recombination rates, and histone modifications) in ±32 kb of these ERVs’ integration sites and in control regions, and analyzed them using Functional Data Analysis (FDA) methodology. In one of the first applications of FDA in genomics, we identified genomic scales and locations at which these features display their influence, and how they work in concert, to provide signals essential for integration and fixation of ERVs. The investigation of ERVs of different evolutionary ages (young in vitro and polymorphic ERVs, older fixed ERVs) allowed us to disentangle integration vs. fixation preferences. As a result of these analyses, we built a comprehensive model explaining the uneven distribution of ERVs along the genome. We found that ERVs integrate in late-replicating AT-rich regions with abundant microsatellites, mirror repeats, and repressive histone marks. Regions favoring fixation are depleted of genes and evolutionarily conserved elements, and have low recombination rates, reflecting the effects of purifying selection and ectopic recombination removing ERVs from the genome. In addition to providing these biological insights, our study demonstrates the power of exploiting multiple scales and localization with FDA. These powerful techniques are expected to be applicable to many other genomic investigations

  18. Express

    Integrated Risk Information System (IRIS)

    Express ; CASRN 101200 - 48 - 0 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Effect

  19. 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

  20. Expression of human adenosine deaminase in murine hematopoietic cells.

    PubMed Central

    Belmont, J W; MacGregor, G R; Wager-Smith, K; Fletcher, F A; Moore, K A; Hawkins, D; Villalon, D; Chang, S M; Caskey, C T

    1988-01-01

    Multiple replication-defective retrovirus vectors were tested for their ability to transfer and express human adenosine deaminase in vitro and in vivo in a mouse bone marrow transplantation model. High-titer virus production was obtained from vectors by using both a retrovirus long terminal repeat promoter and internal transcriptional units with human c-fos and herpes virus thymidine kinase promoters. After infection of primary murine bone marrow with one of these vectors, human adenosine deaminase was detected in 60 to 85% of spleen colony-forming units and in the blood of 14 of 14 syngeneic marrow transplant recipients. This system offers the opportunity to assess methods for increasing efficiency of gene transfer, for regulation of expression of foreign genes in hematopoietic progenitors, and for long-term measurement of the stability of expression in these cells. Images PMID:3072474

  1. 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

  2. The multiple sclerosis-associated retrovirus and its HERV-W endogenous family: a biological interface between virology, genetics, and immunology in human physiology and disease.

    PubMed

    Dolei, Antonina; Perron, Hervé

    2009-01-01

    This mini-review summarizes current knowledge of MSRV (multiple sclerosis-associated retrovirus), founder member of the type W family of human endogenous retroviruses (HERVs), its pathogenic potential and association with diseases. As retrotransposable elements, HERVs behave differently from stable genes, and cannot be studied with "Mendelian genetics" concepts only. They also display complex interactions with other HERV families, and with classical viruses. These concepts may contribute to unravelling the etiopathogenesis of complex diseases such as multiple sclerosis, schizophrenia, and other chronic multifactorial diseases.

  3. Soluble expression and enzymatic activity evaluation of protease from reticuloendotheliosis virus.

    PubMed

    Hu, Feng; Zhao, Yan; Qi, Xiaole; Cui, Hongyu; Gao, Yulong; Gao, Honglei; Liu, Changjun; Wang, Yongqiang; Zhang, Yanping; Li, Kai; Wang, Xiaomei; Wang, Yunfeng

    2015-10-01

    The protease (PR) encoded by most retroviruses is deeply involved in the lifecycle and infection process of retroviruses by possessing the specificity necessary to correctly cleave the viral polyproteins and host cell proteins. However, as an important representative of avian retroviruses, the enzymatic properties of PR from reticuloendotheliosis virus (REV) have not been clearly documented. The recombinant PR, its mutant fused with a His-tag, and its substrate p18-p30 fused with a GST-tag were expressed in the Escherichia coli system as soluble enzymes. The soluble PR and p18-p30 were purified using Ni-NTA His Bind Resin and Glutathione Sepharose 4B, respectively. The enzymatic activity of PR was analyzed using the substrate of p18-p30. The expressed prokaryotic protease has enzyme activity that is dependent on such conditions as temperature, pH, and ions, and its activity can be inhibited by caspase inhibitor and the divalent metal ions Ca(2+) and Ni(2+). In addition, the key role of the residue Thr (amino acids 28) for the enzymatic activity of PR was identified. Furthermore, the caspase inhibitor Z-VAD-FMK was confirmed to inhibit the PR enzymatic activity of REV. For the first time, the PR of REV was expressed in the soluble form, and the optimal enzymatic reaction system in vitro was developed and preliminarily used. This study provides essential tools and information for further understanding the infection mechanism of REV and for the development of antiviral drugs treating retroviruses.

  4. Comparison of serum sensitivities of pseudotype retroviruses produced from newly established packaging cell lines of human and feline origins.

    PubMed

    Watanabe, Rie; Miyazawa, Takayuki; Matsuura, Yoshiharu

    2004-01-01

    To apply retrovirus vectors for in vivo gene therapy in cats, it is necessary to develop vector systems that are not inactivated by cat serum. In this study, the retrovirus packaging cell lines 2SC-1 and AHCeB7 were newly established from human embryonic kidney (HEK) 293 and feline fibroblastic AH927 cells, respectively. Then the sensitivities of pseudotype viruses released from these cell lines to fresh sera from humans and cats were compared. Pseudotype viruses from the 2SC-1 cells were inactivated efficiently by cat serum but not by human serum. Pseudotype viruses from the AHCeB7 cells were also inactivated efficiently by human serum, however they were rather resistant to cat serum. When the xenoantigenicity of the cell lines was examined by flow cytometry, AH927 cells reacted with human serum, however, HEK293 cells did not react with cat serum. These results suggested that pseudotype viruses from 2SC-1 cells were inactivated by the fresh cat serum in an antibody-independent manner. Chelating experiments revealed that certain temperature-sensitive factor(s) other than complements might be involved in the inactivation. The usage of feline cells as packaging cells is suitable for in vivo gene therapy in cats.

  5. 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

  6. The HERV-K Human Endogenous Retrovirus Envelope Protein Antagonizes Tetherin Antiviral Activity

    PubMed Central

    Lemaître, Cécile; Harper, Francis; Pierron, Gérard

    2014-01-01

    ABSTRACT Endogenous retroviruses are the remnants of past retroviral infections that are scattered within mammalian genomes. In humans, most of these elements are old degenerate sequences that have lost their coding properties. The HERV-K(HML2) family is an exception: it recently amplified in the human genome and corresponds to the most active proviruses, with some intact open reading frames and the potential to encode viral particles. Here, using a reconstructed consensus element, we show that HERV-K(HML2) proviruses are able to inhibit Tetherin, a cellular restriction factor that is active against most enveloped viruses and acts by keeping the viral particles attached to the cell surface. More precisely, we identify the Envelope protein (Env) as the viral effector active against Tetherin. Through immunoprecipitation experiments, we show that the recognition of Tetherin is mediated by the surface subunit of Env. Similar to Ebola glycoprotein, HERV-K(HML2) Env does not mediate Tetherin degradation or cell surface removal; therefore, it uses a yet-undescribed mechanism to inactivate Tetherin. We also assessed all natural complete alleles of endogenous HERV-K(HML2) Env described to date for their ability to inhibit Tetherin and found that two of them (out of six) can block Tetherin restriction. However, due to their recent amplification, HERV-K(HML2) elements are extremely polymorphic in the human population, and it is likely that individuals will not all possess the same anti-Tetherin potential. Because of Tetherin's role as a restriction factor capable of inducing innate immune responses, this could have functional consequences for individual responses to infection. IMPORTANCE Tetherin, a cellular protein initially characterized for its role against HIV-1, has been proven to counteract numerous enveloped viruses. It blocks the release of viral particles from producer cells, keeping them tethered to the cell surface. Several viruses have developed strategies to

  7. Molecular characteristics of Human Endogenous Retrovirus type-W in schizophrenia and bipolar disorder.

    PubMed

    Perron, H; Hamdani, N; Faucard, R; Lajnef, M; Jamain, S; Daban-Huard, C; Sarrazin, S; LeGuen, E; Houenou, J; Delavest, M; Moins-Teisserenc, H; Moins-Teiserenc, H; Bengoufa, D; Yolken, R; Madeira, A; Garcia-Montojo, M; Gehin, N; Burgelin, I; Ollagnier, G; Bernard, C; Dumaine, A; Henrion, A; Gombert, A; Le Dudal, K; Charron, D; Krishnamoorthy, R; Tamouza, R; Leboyer, M

    2012-12-04

    Epidemiological and genome-wide association studies of severe psychiatric disorders such as schizophrenia (SZ) and bipolar disorder (BD), suggest complex interactions between multiple genetic elements and environmental factors. The involvement of genetic elements such as Human Endogenous Retroviruses type 'W' family (HERV-W) has consistently been associated with SZ. HERV-W envelope gene (env) is activated by environmental factors and encodes a protein displaying inflammation and neurotoxicity. The present study addressed the molecular characteristics of HERV-W env in SZ and BD. Hundred and thirty-six patients, 91 with BD, 45 with SZ and 73 healthy controls (HC) were included. HERV-W env transcription was found to be elevated in BD (P<10-4) and in SZ (P=0.012) as compared with HC, but with higher values in BD than in SZ group (P<0.01). The corresponding DNA copy number was paradoxically lower in the genome of patients with BD (P=0.0016) or SZ (P<0.0003) than in HC. Differences in nucleotide sequence of HERV-W env were found between patients with SZ and BD as compared with HC, as well as between SZ and BD. The molecular characteristics of HERV-W env also differ from what was observed in Multiple Sclerosis (MS) and may represent distinct features of the genome of patients with BD and SZ. The seroprevalence for Toxoplasma gondii yielded low but significant association with HERV-W transcriptional level in a subgroup of BD and SZ, suggesting a potential role in particular patients. A global hypothesis of mechanisms inducing such major psychoses is discussed, placing HERV-W at the crossroads between environmental, genetic and immunological factors. Thus, particular infections would act as activators of HERV-W elements in earliest life, resulting in the production of an HERV-W envelope protein, which then stimulates pro-inflammatory and neurotoxic cascades. This hypothesis needs to be further explored as it may yield major changes in our understanding and treatment of

  8. Comprehensive analysis of human endogenous retrovirus group HERV-W locus transcription in multiple sclerosis brain lesions by high-throughput amplicon sequencing.

    PubMed

    Schmitt, Katja; Richter, Christin; Backes, Christina; Meese, Eckart; Ruprecht, Klemens; Mayer, Jens

    2013-12-01

    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.

  9. The LTR enhancer of ERV-9 human endogenous retrovirus is active in oocytes and progenitor cells in transgenic zebrafish and humans

    PubMed Central

    Pi, Wenhu; Yang, Zhongan; Wang, Jian; Ruan, Ling; Yu, Xiuping; Ling, Jianhua; Krantz, Sanford; Isales, Carlos; Conway, Simon J.; Lin, Shuo; Tuan, Dorothy

    2004-01-01

    The solitary LTRs of ERV-9 human endogenous retrovirus are middle repetitive DNAs associated with 3,000–4,000 human gene loci including the β-globin gene locus where the ERV-9 LTR is juxtaposed to the locus control region (β-LCR) far upstream of the globin genes. The ERV-9 LTRs are conserved during primate evolution, but their function in the primate genomes is unknown. Here, we show that in transgenic zebrafish harboring the β-globin ERV-9 LTR coupled to the GFP gene, the LTR enhancer was active and initiated synthesis of GFP mRNA in oocytes but not in spermatozoa, and GFP expression in the embryos was maternally inherited. The LTR enhancer was active also in stem/progenitor cell regions of adult tissues of transgenic zebrafish. In human tissues, ERV-9 LTR enhancer was active also in oocytes and stem/progenitor cells but not in spermatozoa and a number of differentiated, adult somatic cells. Transcriptional analyses of the human β-globin gene locus showed that the β-globin ERV-9 LTR enhancer initiated RNA synthesis from the LTR in the direction of the downstream β locus control region and globin genes in ovary and erythroid progenitor cells. The findings suggest that, during oogenesis, ERV-9 LTR enhancers in the human genome could activate the cis-linked gene loci to synthesize maternal mRNAs required for early embryogenesis. Alternatively, the ERV-9 LTR enhancers, in initiating RNA syntheses into the downstream genomic DNAs, could transcriptionally potentiate and preset chromatin structure of the cis-linked gene loci in oocytes and adult stem/progenitor cells. PMID:14718667

  10. Epstein-Barr Virus Latent Membrane Protein LMP-2A Is Sufficient for Transactivation of the Human Endogenous Retrovirus HERV-K18 Superantigen

    PubMed Central

    Sutkowski, Natalie; Chen, Gang; Calderon, German; Huber, Brigitte T.

    2004-01-01

    Superantigens are microbial proteins that strongly stimulate T cells. We described previously that the Epstein-Barr virus (EBV) transactivates a superantigen encoded by the human endogenous retrovirus, HERV-K18. We now report that the transactivation is dependent upon the EBV latent cycle proteins. Moreover, LMP-2A is sufficient for induction of HERV-K18 superantigen activity. PMID:15220463

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  12. Comparative host protein interactions with HTLV-1 p30 and HTLV-2 p28: insights into difference in pathobiology of human retroviruses

    PubMed Central

    2012-01-01

    Background Human T lymphotropic virus type-1 (HTLV-1) and type 2 (HTLV-2) are closely related human retroviruses, but have unique disease associations. HTLV-1 is the causative agent of an aggressive T-cell leukemia known as adult T-cell leukemia (ATL), HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP), and other inflammatory diseases. HTLV-2 infection has not been clearly associated with any disease condition. Although both viruses can transform T cells in vitro, the HTLV-1 provirus is mainly detected in CD4+ T cells whereas HTLV-2 is mainly detected in CD8+ T cells of infected individuals. HTLV-1 and HTLV-2 encode accessory proteins p30 and p28, respectively, which share partial amino acid homology and are required for viral persistence in vivo. The goal of this study was to identify host proteins interacting with p30 and p28 in order to understand their role in pathogenesis. Results Affinity-tag purification coupled with mass spectrometric (MS) analyses revealed 42 and 22 potential interacting cellular partners of p30 and p28, respectively. Of these, only three cellular proteins, protein arginine methyltransferase 5 (PRMT5), hnRNP K and 60 S ribosomal protein L8 were detected in both p30 and p28 fractions. To validate the proteomic results, four interacting proteins were selected for further analyses using immunoblot assays. In full agreement with the MS analysis two cellular proteins REGγ and NEAF-interacting protein 30 (NIP30) selectively interacted with p30 and not with p28; heterogeneous nuclear ribonucleoprotein H1 (hnRNP H1) bound to p28 and not to p30; and PRMT5 interacted with both p30 and p28. Further studies demonstrated that reduced levels of PRMT5 resulted in decreased HTLV-2 viral gene expression whereas the viral gene expression of HTLV-1 was unchanged. Conclusion The comparisons of p30 and p28 host protein interaction proteome showed striking differences with some degree of overlap. PRMT5, one of the host proteins that

  13. Single cell analysis and selection of living retrovirus vector-corrected mucopolysaccharidosis VII cells using a fluorescence-activated cell sorting-based assay for mammalian beta-glucuronidase enzymatic activity.

    PubMed

    Lorincz, M C; Parente, M K; Roederer, M; Nolan, G P; Diwu, Z; Martin, D I; Herzenberg, L A; Wolfe, J H

    1999-01-08

    Mutations in the acid beta-glucuronidase gene lead to systemic accumulation of undegraded glycosaminoglycans in lysosomes and ultimately to clinical manifestations of mucopolysaccharidosis VII (Sly disease). Gene transfer by retrovirus vectors into murine mucopolysaccharidosis VII hematopoietic stem cells or fibroblasts ameliorates glycosaminoglycan accumulation in some affected tissues. The efficacy of gene therapy for mucopolysaccharidosis VII depends on the levels of beta-glucuronidase secreted by gene-corrected cells; therefore, enrichment of transduced cells expressing high levels of enzyme prior to transplantation is desirable. We describe the development of a fluorescence-activated cell sorter-based assay for the quantitative analysis of beta-glucuronidase activity in viable cells. Murine mucopolysaccharidosis VII cells transduced with a beta-glucuronidase retroviral vector can be isolated by cell sorting on the basis of beta-glucuronidase activity and cultured for further use. In vitro analysis revealed that sorted cells have elevated levels of beta-glucuronidase activity and secrete higher levels of cross-correcting enzyme than the population from which they were sorted. Transduced fibroblasts stably expressing beta-glucuronidase after subcutaneous passage in the mucopolysaccharidosis VII mouse can be isolated by cell sorting and expanded ex vivo. A relatively high percentage of these cells maintain stable expression after secondary transplantation, yielding significantly higher levels of enzymatic activity than that generated in the primary transplant.

  14. Surveys of endogenous retroviruses (ERVs) in the freshwater crocodile (Crocodylus johnstoni) suggest that ERVs in Crocodylus spp. vary between species.

    PubMed

    Chong, Amanda Y; Kjeldsen, Shannon R; Gongora, Jaime

    2015-04-01

    Endogenous retroviruses (ERVs) are one of many families of transposable elements present in vertebrate genomes. We have examined the ERV complement of the freshwater crocodile (Crocodylus johnstoni) in order to investigate the diversity of ERVs present and possibility of ERV or retroviral activity in a diseased individual of this species. Amplification and sequencing of the highly conserved retroviral pro-pol domains revealed high levels of sequence variation in these ERVs. Phylogenetic analyses of these ERVs and those previously identified in other crocodilian species suggest that although many crocodilians share the same ERV lineages, the relative numbers of retroelement insertions from each of these lineages may vary greatly between species. The data generated in this study provide evidence for the presence of a unique and varied complement of ERVs in crocodilians. This study has also demonstrated the presence of species-specific evolution in ancient retroviral infections.

  15. Cytolytic T lymphocytes specific for tumors and infected cells from mice with a retrovirus-induced immunodeficiency syndrome.

    PubMed Central

    Erbe, J G; Green, K A; Crassi, K M; Morse, H C; Green, W R

    1992-01-01

    LP-BM5 retrovirus complex-infected C57BL/6 mice develop immunodeficiency, somewhat analogous to AIDS, termed murine AIDS (MAIDS). After secondary stimulation with syngeneic B-cell lymphomas from LP-BM5-infected mice, C57BL/6 mice produced vigorous CD8+ cytotoxic T lymphocytes specific for MAIDS-associated tumors. An anti-LP-BM5 specificity was suggested because spleen and lymph node cells from LP-BM5-infected mice served as target cells in competition assays, and cells from LP-BM5, but not ecotropic, virus-infected mice functioned as secondary in vitro stimulators to generate cytotoxic T lymphocytes to MAIDS tumors. PMID:1560546

  16. Molecular cloning of the unintegrated squirrel monkey retrovirus genome: organization and distribution of related sequences in primate DNAs.

    PubMed Central

    Chiu, I M; Andersen, P R; Aaronson, S A; Tronick, S R

    1983-01-01

    The closed circular form of the endogenous squirrel monkey type D retrovirus (SMRV) was molecularly cloned in a bacteriophage vector. The restriction map of the biologically active clone was determined and found to be identical to that of the parental SMRV linear DNA except for the deletion of one long terminal repeat. Restriction enzyme analysis and Southern blotting indicated that the SMRV long terminal repeat was approximately 300 base pairs long. The SMRV restriction map was oriented to the viral RNA by using a gene-specific probe from baboon endogenous virus. Restriction enzyme digests of a variety of vertebrate DNAs were analyzed for DNA sequence homology with SMRV by using the cloned SMRV genome as a probe. Consistent with earlier studies, multiple copies of SMRV were detected in squirrel monkey DNA. Related fragments were also detected in the DNAs from other primate species, including humans. Images PMID:6312076

  17. The 14th International Conference on Human Retrovirology: HTLV and related retroviruses (July 1–4, 2009; Salvador, Brazil)

    PubMed Central

    Willems, Luc

    2009-01-01

    The "14th International Conference on Human Retrovirology: HTLV and Related Retroviruses" was held in Salvador, Bahia, from July 1st to July 4th 2009. The aim of this biennial meeting is to promote discussion and share new findings between researchers and clinicians for the benefit of patients infected by human T-lymphotropic virus (HTLV). HTLV infects approximately 15–20 million individuals worldwide and causes a broad spectrum of diseases including neurodegeneration and leukemia. The scientific program included a breadth of HTLV research topics: epidemiology, host immune response, basic mechanisms of protein function, virology, pathogenesis, clinical aspects and treatment. Exciting new findings were presented in these different fields, and the new advances have led to novel clinical trials. Here, highlights from this conference are summarized. PMID:19686596

  18. 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.

  19. Changes in lymphocyte and macrophage subsets due to morphine and ethanol treatment during a retrovirus infection causing murine AIDS

    SciTech Connect

    Watson, R.R.; Prabhala, R.H.; Darban, H.R.; Yahya, M.D.; Smith, T.L.

    1988-01-01

    The number of lymphocytes of various subsets were not significantly changed by the ethanol exposure except those showing activation markers which were reduced. The percentage of peripheral blood cells showing markers for macrophage functions and their activation were significantly reduced after binge use of ethanol. Ethanol retarded suppression of cells by retroviral infection. However by 25 weeks of infection there was a 8.6% survival in the ethanol fed mice infected with retrovirus which was much less than virally infected controls. Morphine treatment also increased the percentage of cells with markers for macrophages and activated macrophages in virally infected mice, while suppressing them in uninfected mice. The second and third morphine injection series suppressed lymphocyte T-helper and T-suppressor cells, but not total T cells. However, suppression by morphine was significantly less during retroviral disease than suppression caused by the virus only. At 25 weeks of infection 44.8% of morphine treated, infected mice survived.

  20. NICER elements: a family of nerve growth factor-inducible cAMP-extinguishable retrovirus-like elements.

    PubMed Central

    Cho, K O; Minsk, B; Wagner, J A

    1990-01-01

    We have shown previously that the transcription of the gene designated d5 is induced by nerve growth factor (NGF) in rat adrenal pheochromocytoma PC-12 cells and that this NGF induction is repressed by cAMP. In this paper we demonstrate that d5 is a member of a gene family that contains several hundred members, which is closely related to retroviruses and retrotransposons, as demonstrated by the following observations: (i) the original d5 cDNA hybridized to numerous restriction fragments in genomic DNA; (ii) d5 cDNA hybridized to genomic clones with various intensities, and genomic clones can be isolated with a frequency suggesting that this family includes several hundred members; and (iii) there were minor sequence variations in four independently isolated cDNA clones that were homologous to d5 cDNA. Primer extension studies show that initiation of the 5.7-kilobase d5 mRNA(s) occurs at a unique site relative to a synthetic primer. The 5' end of the cDNA sequence was homologous to Rasheed rat sarcoma virus; and a genomic clone contained several elements that are typical of a long terminal repeat (LTR), including a CCAAT box, a TATA box, a primer binding site, a poly(A) addition signal, and a poly(A) addition site. Furthermore, there is a LTR at the 3' end of at least one of the genes in this family, and there appeared to be a four-base duplication at the probable site of integration into host DNA. Since several members of this family retain responses to NGF and cAMP, we conclude that the regulatory elements present in the LTR have been conserved in many members of this family. We have named this family of genes the NICER elements because they are a family of NGF-inducible cAMP-extinguishable retrovirus-like elements. Images PMID:2160077

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

    SciTech Connect

    Barabás, Orsolya; Németh, Veronika; Vértessy, Beáta G.

    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 vicinity 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.

  2. CArG, CCAAT, and CCAAT-like protein binding sites in avian retrovirus long terminal repeat enhancers.

    PubMed Central

    Zachow, K R; Conklin, K F

    1992-01-01

    A strong enhancer element is located within the long terminal repeats (LTRs) of exogenous, oncogenic avian retroviruses, such as Rous sarcoma virus (RSV) and the avian leukosis viruses. The LTRs of a second class of avian retroviruses, the endogenous viruses (evs), lack detectable enhancer function, a property that correlates with major sequence differences between the LTRs of these two virus groups. Despite this lack of independent enhancer activity, we previously identified sequences in ev LTRs that were able to functionally replace essential enhancer domains from the RSV enhancer with which they share limited sequence similarity. To identify candidate enhancer domains in ev LTRs that are functionally equivalent to those in RSV LTRs, we analyzed and compared ev and RSV LTR-specific DNA-protein interactions. Using this approach, we identified two candidate enhancer domains and one deficiency in ev LTRs. One of the proposed ev enhancer domains was identified as a CArG box, a motif also found upstream of several muscle-specific genes, and as the core sequence of the c-fos serum response element. The RSV LTR contains two CArG motifs, one at a previously identified site and one identified in this report at the same relative location as the ev CArG motif. A second factor binding site that interacts with a heat-stable protein was also identified in ev LTRs and, contrary to previous suggestions, appears to be different from previously described exogenous virus enhancer binding proteins. Finally, a deficiency in factor binding was found within the one inverted CCAAT box in ev LTRs, affirming the importance of sequences that flank CCAAT motifs in factor binding and providing a candidate defect in the ev enhancer. Images PMID:1312613

  3. Strong purifying selection in endogenous retroviruses in the saltwater crocodile (Crocodylus porosus) in the Northern Territory of Australia

    PubMed Central

    2012-01-01

    Background Endogenous retroviruses (ERVs) are remnants of exogenous retroviruses that have integrated into the nuclear DNA of a germ-line cell. Here we present the results of a survey into the ERV complement of Crocodylus porosus, the saltwater crocodile, representing 45 individuals from 17 sampling locations in the Northern Territory of Australia. These retroelements were compared with published ERVs from other species of Crocodylia (Crocodilians; alligators, caimans, gharials and crocodiles) as well as representatives from other vertebrates. This study represents one of the first in-depth studies of ERVs within a single reptilian species shedding light on the diversity of ERVs and proliferation mechanisms in crocodilians. Results Analyses of the retroviral pro-pol gene region have corroborated the presence of two major clades of ERVs in C. porosus and revealed 18 potentially functional fragments out of the 227 recovered that encode intact pro-pol ORFs. Interestingly, we have identified some patterns of diversification among those ERVs as well as a novel sequence that suggests the presence of an additional retroviral genus in C. porosus. In addition, considerable diversity but low genetic divergence within one of the C. porosus ERV lineages was identified. Conclusions We propose that the ERV complement of C. porosus has come about through a combination of recent infections and replication of ancestral ERVs. Strong purifying selection acting on these clades suggests that this activity is recent or still occurring in the genome of this species. The discovery of potentially functional elements is an interesting development that warrants further investigation. PMID:23217152

  4. Retroviral Restriction Factors and Infectious Risk in Xenotransplantation

    PubMed Central

    Meije, Yolanda; Tönjes, Ralf R.; Fishman, Jay A.

    2010-01-01

    The clinical application of xenotransplantation poses immunologic, ethical, and microbiologic challenges. Significant progress has been made in the investigation of each of these areas. Among concerns regarding infectious risks for human xenograft recipients is the identification in swine of infectious agents including porcine endogenous retroviruses (PERV) that are capable of replication in some human cell lines. PERV replication has, however, been difficult to demonstrate in primate-derived cell lines and in preclinical studies of non-human primates receiving porcine xenografts. Endogenous “retroviral restriction factors” are intracellular proteins and components of the innate immune system that act at various steps in retroviral replication. Recent studies suggest that some of these factors may have applications in the management of endogenous retroviruses in xenotransplantation. The risks of PERV infection and the potential role of retroviral restriction factors in xenotransplantation are reviewed in detail. PMID:20642677

  5. Specific detection of RT activity in culture supernantants of retrovirus-producing cells, using synthetic DNA as competitor in polymerase enhanced reverse transcriptase assay.

    PubMed

    Voisset, C; Tönjes, R R; Breyton, P; Mandrand, B; Paranhos-Baccalà, G

    2001-05-01

    The polymerase enhanced reverse transcriptase (PERT) assay is a highly sensitive assay for the detection of reverse transcriptase (RT) activity in culture supernatants of retrovirus-producing cells. However, some cellular DNA-dependent DNA polymerases exhibit RT-like activities in this assay. A synthetic DNA competitor which suppresses the RT-like activities of cellular DNA-dependent DNA polymerases was used in a modified PERT assay technique for specific detection of RT activity in culture supernatants of retrovirus-producing cells. We determined the optimum condition of the assay and evaluated its specificity. This improved PERT assay is easy to perform and is able to detect minute amounts of purified RT, as well as RT in crude cell lysates and concentrated culture supernatants.

  6. Isolation and characterization of NIH 3T3 cells expressing polyomavirus small T antigen

    SciTech Connect

    Noda, T.; Satake, M.; Robins, T.; Ito, Y.

    1986-10-01

    The polyomavirus small T-antigen gene, together with the polyomavirus promoter, was inserted into retrovirus vector pGV16 which contains the Moloney sarcoma virus long terminal repeat and neomycin resistance gene driven by the simian virus 40 promoter. This expression vector, pGVST, was packaged into retrovirus particles by transfection of PSI2 cells which harbor packaging-defective murine retrovirus genome. NIH 3T3 cells were infected by this replication-defective retrovirus containing pGVST. Of the 15 G418-resistant cell clones, 8 express small T antigen at various levels as revealed by immunoprecipitation. A cellular protein with an apparent molecular weight of about 32,000 coprecipitates with small T antigen. Immunofluorescent staining shows that small T antigen is mainly present in the nuclei. Morphologically, cells expressing small T antigen are indistinguishable from parental NIH 3T3 cells and have a microfilament pattern similar to that in parental NIH 3T3 cells. Cells expressing small T antigen form a flat monolayer but continue to grow beyond the saturation density observed for parental NIH 3T3 cells and eventually come off the culture plate as a result of overconfluency. There is some correlation between the level of expression of small T antigen and the growth rate of the cells. Small T-antigen-expressing cells form small colonies in soft agar. However, the proportion of cells which form these small colonies is rather small. A clone of these cells tested did not form tumors in nude mice within 3 months after inoculation of 10/sup 6/ cells per animal. Thus, present studies establish that the small T antigen of polyomavirus is a second nucleus-localized transforming gene product of the virus (the first one being large T antigen) and by itself has a function which is to stimulate the growth of NIH 3T3 cells beyond their saturation density in monolayer culture.

  7. Implications of the evolution pattern of human T-cell leukemia retroviruses on their pathogenic virulence (Review).

    PubMed

    Azran, Inbal; Schavinsky-Khrapunsky, Yana; Priel, Esther; Huleihel, Mahmoud; Aboud, Mordechai

    2004-11-01

    Simian retroviruses pose a serious threat to public health, as two human pathogenic retroviruses, HIV and HTLV, have been already proved to originate from such non-human viruses. Therefore, studying their natural prevalence among wild non-human primates is important for planning strategies to prevent the emergence of additional human retroviral pathogens. This article is focused on tracing the origin and evolution of the human T-cell leukemia viruses HTLV-I and HTLV-II in comparison to that of the simian lymphotropic viruses STLV-I, STLV-II and STLV-L, which are phylo-genically classified into a common group called primate T-lymphotropic viruses (PTLV). Thus, HTLV-I and STLV-I are referred to as PTLV-I and HTLV-II and STLV-II as PTLV-II, whereas STLV-L, which is highly divergent from both HTLV types, comprises a third subgroup called PTLV-L. The phylogeny of PTLV indicates that both, HTLV-I and HTLV-II emerged from a simian origin, but their subsequent evolution continued in different patterns. HTLV-I includes 6 subtypes which evolved from STLV-I through several times of different geographic interspecies transmission between simian and human hosts. These repeated invasions to new primate species are likely to give rise to viral strains with increasing pathogenic potential. On the other hand, HTLV-II includes 4 subtypes which appear to originate from a common human ancestor virus that emerged from only one simian to human transmission, whereas the subsequent evolution of HTLV-II and STLV-II strains continued separately only within the Homo sapiens and Pan paniscus species respectively, without repeated interspecies jumps. Such evolution pattern likely involves less genetic changes and selection of viral strains with low pathogenic virulence that could co-exist with their hosts for long time. These different evolution patterns can explain the much wider implication of HTLV-I with human clinical disorders than HTLV-II. Of note, however, more recently HTLV-II started

  8. New way of regulating alternative splicing in retroviruses: the promoter makes a difference.

    PubMed

    Bohne, Jens; Schambach, Axel; Zychlinski, Daniela

    2007-04-01

    Alternative splicing has been recognized as a major mechanism for creating proteomic diversity from a limited number of genes. However, not all determinants regulating this process have been characterized. Using subviral human immunodeficiency virus (HIV) env constructs we observed an enhanced splicing of the RNA when expression was under control of the cytomegalovirus (CMV) promoter instead of the HIV long terminal repeat (LTR). We extended these observations to LTR- or CMV-driven murine leukemia proviruses, suggesting that retroviral LTRs are adapted to inefficient alternative splicing at most sites in order to maintain balanced gene expression.

  9. Purification and N-terminal amino acid sequence comparisons of structural proteins from retrovirus-D/Washington and Mason-Pfizer monkey virus.

    PubMed Central

    Henderson, L E; Sowder, R; Smythers, G; Benveniste, R E; Oroszlan, S

    1985-01-01

    A new D-type retrovirus originally designated SAIDS-D/Washington and here referred to as retrovirus-D/Washington (R-D/W) was recently isolated at the University of Washington Primate Center, Seattle, Wash., from a rhesus monkey with an acquired immunodeficiency syndrome and retroperitoneal fibromatosis. To better establish the relationship of this new D-type virus to the prototype D-type virus, Mason-Pfizer monkey virus (MPMV), we have purified and compared six structural proteins from each virus. The proteins purified from each D-type retrovirus include p4, p10, p12, p14, p27, and a phosphoprotein designated pp18 for MPMV and pp20 for R-D/W. Amino acid analysis and N-terminal amino acid sequence analysis show that the p4, p12, p14, and p27 proteins of R-D/W are distinct from the homologous proteins of MPMV but that these proteins from the two different viruses share a high degree of amino acid sequence homology. The p10 proteins from the two viruses have similar amino acid compositions, and both are blocked to N-terminal Edman degradation. The phosphoproteins from the two viruses each contain phosphoserine but are different from each other in amino acid composition, molecular weight, and N-terminal amino acid sequence. The data thus show that each of the R-D/W proteins examined is distinguishable from its MPMV homolog and that a major difference between these two D-type retroviruses is found in the viral phosphoproteins. The N-terminal amino acid sequences of D-type retroviral proteins were used to search for sequence homologies between D-type and other retroviral amino acid sequences. An unexpected amino acid sequence homology was found between R-D/W pp20 (a gag protein) and a 28-residue segment of the env precursor polyprotein of Rous sarcoma virus. The N-terminal amino acid sequences of the D-type major gag protein (p27) and the nucleic acid-binding protein (p14) show only limited amino acid sequence homology to functionally homologous proteins of C

  10. Retrovirus-induced osteopetrosis in mice. Effects of viral infection on osteogenic differentiation in skeletoblast cell cultures.

    PubMed Central

    Schmidt, J.; Casser-Bette, M.; Murray, A. B.; Luz, A.; Erfle, V.

    1987-01-01

    Newborn female strain NMRI mice were injected with a mouse retrovirus (OA MuLV) known to induce osteopetrosis. Primary skeletoblast cell cultures were established from humeri and calvaria of 3-day-old, 7-day-old, and 28-day-old animals. Infectious ecotropic MuLV was found in all humerus cultures from infected animals and in 7-day and 28-day calvaria cell cultures. Levels of alkaline phosphatase activity were markedly higher in cultures of calvaria and humeri from infected mice than in those from controls. In vitro infection of undifferentiated periosteal cells was followed by a decrease in cell growth and an increase in alkaline phosphatase activity. In contrast, differentiated osteoblast-like cells were barely susceptible to OA MuLV infection, and the virus did not influence their cell growth or differentiation. Electron-microscopic studies of skeletal tissue from infected old osteopetrotic mice showed virus particles associated with and budding from osteocytes and accumulated in devitalized osteocyte lacunae. The results indicate that progenitor cells of the osteoblastic lineage represent the target cells for OA MuLV in bone tissue, that virus infection induces an increase in osteoblastic activity, and that infected cells produce virus until full development of the disease. Images Figure 1 Figure 2 Figure 5 PMID:2827489

  11. Investigation of Human Cancers for Retrovirus by Low-Stringency Target Enrichment and High-Throughput Sequencing

    PubMed Central

    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

    2015-01-01

    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. PMID:26285800

  12. The role of molecular mimicry and other factors in the association of Human Endogenous Retroviruses and autoimmunity.

    PubMed

    Trela, Malgorzata; Nelson, Paul N; Rylance, Paul B

    2016-01-01

    Human Endogenous Retroviruses (HERVs) have been implicated in autoimmune and other diseases. Molecular mimicry has been postulated as a potential mechanism of autoimmunity. Exogenous viruses have also been reported to be associated with the same diseases, as have genetic and environmental factors. If molecular mimicry were to be shown to be an initiating mechanism of some autoimmune diseases, then therapeutic options of blocking antibodies and peptides might be of benefit in halting diseases at the outset. Bioinformatic and molecular modelling techniques have been employed to investigate molecular mimicry and the evidence for the association of HERVs and autoimmunity is reviewed. The most convincing evidence for molecular mimicry is in rheumatoid arthritis, where HERV K-10 shares amino acid sequences with IgG1Fc, a target for rheumatoid factor. Systemic lupus erythematosus is an example of a condition associated with several autoantibodies, and several endogenous and exogenous viruses have been reported to be associated with the disease. The lack of a clear link between one virus and this condition, and the spectrum of clinical manifestations, suggests that genetic, environmental and the inflammatory response to a virus or viruses might also be major factors in the pathogenesis of lupus and other autoimmune conditions. Where there are strong associations between a virus and an autoimmune condition, such as in hepatitis C and cryoglobulinaemia, the use of bioinformatics and molecular modelling can also be utilized to help to understand the role of molecular mimicry in how HERVs might trigger disease.

  13. 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.

  14. ‘There and back again’: revisiting the pathophysiological roles of human endogenous retroviruses in the post-genomic era

    PubMed Central

    Magiorkinis, Gkikas; Belshaw, Robert; Katzourakis, Aris

    2013-01-01

    Almost 8% of the human genome comprises endogenous retroviruses (ERVs). While they have been shown to cause specific pathologies in animals, such as cancer, their association with disease in humans remains controversial. The limited evidence is partly due to the physical and bioethical restrictions surrounding the study of transposons in humans, coupled with the major experimental and bioinformatics challenges surrounding the association of ERVs with disease in general. Two biotechnological landmarks of the past decade provide us with unprecedented research artillery: (i) the ultra-fine sequencing of the human genome and (ii) the emergence of high-throughput sequencing technologies. Here, we critically assemble research about potential pathologies of ERVs in humans. We argue that the time is right to revisit the long-standing questions of human ERV pathogenesis within a robust and carefully structured framework that makes full use of genomic sequence data. We also pose two thought-provoking research questions on potential pathophysiological roles of ERVs with respect to immune escape and regulation. PMID:23938753

  15. Interaction between endoplasmic reticulum stress and caspase 8 activation in retrovirus MoMuLV-ts1-infected astrocytes.

    PubMed

    Liu, Na; Scofield, Virginia L; Qiang, Wenan; Yan, Mingshan; Kuang, Xianghong; Wong, Paul K Y

    2006-05-10

    The murine retrovirus, MoMuLV-ts1, induces progressive paralysis and immune deficiency in FVB/N mice. We have reported previously that ts1 infection causes apoptosis in astrocytes via endoplasmic reticulum (ER) and mitochondrial stress (Liu, N., Kuang, X., Kim, H.T., Stoica, G., Qiang, W., Scofield, V.L., Wong, P.K.Y. Wong. 2004. Possible involvement of both endoplasmic reticulum- and mitochondria-dependent pathways in MoMuLV-ts1-induced apoptosis in astrocytes. J. NeuroVirol. 10, 189-198). In the present study, we show that caspase 8 activation in these cells is mediated through ER stress-associated elevation of death receptor DR5 and the C/EBP homologous protein (GADD153/CHOP), an ER stress-initiated transcription factor, rather than through TNFalpha and TNF-R1 interactions on the cell surface. Treatment with Z-IETD-FMK, a specific inhibitor of caspase 8 enzymatic activity, reduced ER stress by two mechanisms: by inhibiting caspase 8 activation, and by preventing cleavage of the ER-associated membrane protein BAP31 into BAP20, which exacerbates the ER stress response. These findings suggest that caspase 8- and ER stress-associated apoptotic pathways are linked in ts1-infected astrocytes.

  16. Selective cell targeting and lineage tracing of human induced pluripotent stem cells using recombinant avian retroviruses.

    PubMed

    Hildebrand, Laura; Seemann, Petra; Kurtz, Andreas; Hecht, Jochen; Contzen, Jörg; Gossen, Manfred; Stachelscheid, Harald

    2015-12-01

    Human induced pluripotent stem cells (hiPSC) differentiate into multiple cell types. Selective cell targeting is often needed for analyzing gene function by overexpressing proteins in a distinct population of hiPSC-derived cell types and for monitoring cell fate in response to stimuli. However, to date, this has not been possible, as commonly used viruses enter the hiPSC via ubiquitously expressed receptors. Here, we report for the first time the application of a heterologous avian receptor, the tumor virus receptor A (TVA), to selectively transduce TVA(+) cells in a mixed cell population. Expression of the TVA surface receptor via genetic engineering renders cells susceptible for infection by avian leucosis virus (ALV). We generated hiPSC lines with this stably integrated, ectopic TVA receptor gene that expressed the receptor while retaining pluripotency. The undifferentiated hiPSC(TVA+) as well as their differentiating progeny could be infected by recombinant ALV (so-called RCAS virus) with high efficiency. Due to incomplete receptor blocking, even sequential infection of differentiating or undifferentiated TVA(+) cells was possible. In conclusion, the TVA/RCAS system provides an efficient and gentle gene transfer system for hiPSC and extends our possibilities for selective cell targeting and lineage tracing studies.

  17. 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

  18. Retrovirus-mediated transduction of a cytosine deaminase gene preserves the stemness of mesenchymal stem cells.

    PubMed

    Park, Jin Sung; Chang, Da-Young; Kim, Ji-Hoi; Jung, Jin Hwa; Park, JoonSeong; Kim, Se-Hyuk; Lee, Young-Don; Kim, Sung-Soo; Suh-Kim, Haeyoung

    2013-02-22

    Human mesenchymal stem cells (MSCs) have emerged as attractive cellular vehicles to deliver therapeutic genes for ex-vivo therapy of diverse diseases; this is, in part, because they have the capability to migrate into tumor or lesion sites. Previously, we showed that MSCs could be utilized to deliver a bacterial cytosine deaminase (CD) suicide gene to brain tumors. Here we assessed whether transduction with a retroviral vector encoding CD gene altered the stem cell property of MSCs. MSCs were transduced at passage 1 and cultivated up to passage 11. We found that proliferation and differentiation potentials, chromosomal stability and surface antigenicity of MSCs were not altered by retroviral transduction. The results indicate that retroviral vectors can be safely utilized for delivery of suicide genes to MSCs for ex-vivo therapy. We also found that a single retroviral transduction was sufficient for sustainable expression up to passage 10. The persistent expression of the transduced gene indicates that transduced MSCs provide a tractable and manageable approach for potential use in allogeneic transplantation.

  19. Defective virus is associated with induction of murine retrovirus-induced immunodeficiency syndrome.

    PubMed Central

    Chattopadhyay, S K; Morse, H C; Makino, M; Ruscetti, S K; Hartley, J W

    1989-01-01

    C57BL/6 mice infected with a mixture of murine leukemia viruses (MuLV) develop a syndrome characterized by lymphoproliferation and profound immunodeficiency. Analyses of this viral mixture (LP-BM5 MuLV) showed that it includes replication-competent ecotropic and mink cell focus-inducing MuLV and defective viruses with genome sizes of 3.8-6.5 kilobases. The ecotropic and mink cell focus-inducing MuLV biologically cloned from the mixture did not induce disease, whereas viral preparations containing the ecotropic MuLV and 4.8-kilobase defective virus were active. Cells producing the 4.8-kilobase defective virus expressed an unusual gag-encoded polyprotein of Mr 60,000. Images PMID:2542949

  20. Retrovirus-induced oxidative stress with neuroimmunodegeneration is suppressed by antioxidant treatment with a refined monosodium alpha-luminol (Galavit).

    PubMed

    Jiang, Yuhong; Scofield, Virginia L; Yan, Mingshan; Qiang, Wenan; Liu, Na; Reid, Amy J; Lynn, William S; Wong, Paul K Y

    2006-05-01

    Oxidative stress is involved in many human neuroimmunodegenerative diseases, including human immunodeficiency virus disease/AIDS. The retrovirus ts1, a mutant of Moloney murine leukemia virus, causes oxidative stress and progressive neuro- and immunopathology in mice infected soon after birth. These pathological changes include spongiform neurodegeneration, astrogliosis, thymic atrophy, and T-cell depletion. Astrocytes and thymocytes are directly infected and killed by ts1. Neurons are not infected, but they also die, most likely as an indirect result of local glial infection. Cytopathic effects of ts1 infection in cultured astrocytes are associated with accumulation of the viral envelope precursor protein gPr80env in the endoplasmic reticulum (ER), which triggers ER stress and oxidative stress. We have reported (i) that activation of the Nrf2 transcription factor and upregulation of antioxidative defenses occurs in astrocytes infected with ts1 in vitro and (ii) that some ts1-infected astrocytes survive infection by mobilization of these pathways. Here, we show that treatment with a refined monosodium alpha-luminol (Galavit; GVT) suppresses oxidative stress and Nrf2 activation in cultured ts1-infected astrocytes. GVT treatment also inhibits the development of spongiform encephalopathy and gliosis in the central nervous system (CNS) in ts1-infected mice, preserves normal cytoarchitecture in the thymus, and delays paralysis, thymic atrophy, wasting, and death. GVT treatment of infected mice reduces ts1-induced oxidative stress, cell death, and pathogenesis in both the CNS and thymus of treated animals. These studies suggest that oxidative stress mediates ts1-induced neurodegeneration and T-cell loss.

  1. Lung Adenocarcinoma Originates from Retrovirus Infection of Proliferating Type 2 Pneumocytes during Pulmonary Post-Natal Development or Tissue Repair

    PubMed Central

    Murgia, Claudio; Caporale, Marco; Ceesay, Ousman; Di Francesco, Gabriella; Ferri, Nicola; Varasano, Vincenzo; de las Heras, Marcelo; Palmarini, Massimo

    2011-01-01

    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. PMID:21483485

  2. Disparities in outcomes for African American and Latino subjects in the Flexible Initial Retrovirus Suppressive Therapies (FIRST) trial.

    PubMed

    Giordano, Thomas P; Bartsch, Glenn; Zhang, Yafeng; Tedaldi, Ellen; Absalon, Judith; Mannheimer, Sharon; Thomas, Avis; MacArthur, Rodger D

    2010-05-01

    To benefit maximally from antiretroviral therapy, patients with HIV infection must enter care before their disease is advanced and adhere to care. We sought to determine if and where on this continuum of care racial/ethnic disparities were evident. Data from the Flexible Initial Retrovirus Suppressive Therapies (FIRST) trial, which evaluated three strategies for initial HIV therapy, were compared for White, African American, and Latino subjects. Outcomes included progression of disease and death, HIV viral suppression, and change in CD4(+) cell count. Multivariate Cox proportional hazard models adjusted for known predictors of survival. There were 1357 subjects, including 368 non-Latino white, 751 non-Latino African American, and 238 Latino subjects. At baseline, the two latter groups were more likely to have had AIDS and had lower CD4(+) cell counts than white subjects. In follow-up, African American subjects had lower self-reported adherence to therapy, lower CD4(+) cell count increases, and lower odds of viral suppression. African American and Latino subjects had unadjusted hazard ratios of progression of disease or death of 1.57 (1.17, 2.10; p = 0.0025) and 1.57 (1.09, 2.26; p = 0.02), respectively. Adjusting for baseline differences and differences in adherence, CD4(+) cell count change, and viral suppression accounted for the disparities in outcomes. Opportunities to reduce disparities in outcomes for African American and Latino patients exist along the continuum of HIV care. Efforts to promote access to HIV testing and care and to improve adherence have the potential to reduce racial/ethnic disparities in outcomes of patients with HIV infection.

  3. Transport of cationic amino acids by the mouse ecotropic retrovirus receptor.

    PubMed

    Kim, J W; Closs, E I; Albritton, L M; Cunningham, J M

    1991-08-22

    Susceptibility of rodent cells to infection by ecotropic murine leukaemia viruses (MuLV) is determined by binding of the virus envelope to a membrane receptor that has multiple membrane-spanning domains. Cells infected by ecotropic MuLV synthesize envelope protein, gp70, which binds to this receptor, thereby preventing additional infections. The consequences of envelope-MuLV receptor binding for the infected host cell have not been directly determined, partly because the cellular function of the MuLV receptor protein is unknown. Here we report a coincidence in the positions of the first eight putative membrane-spanning domains found in the virus receptor and in two related proteins, the arginine and histidine permeases of Saccharomyces cerevisiae (Fig. 1), but not in any other proteins identified by computer-based sequence comparison of the GenBank data base. Xenopus oocytes injected with receptor-encoding messenger RNA show increased uptake of L-arginine, L-lysine and L-ornithine. The transport properties and the expression pattern of the virus receptor behave in ways previously attributed to y+, the principal transporter of cationic L-amino acids in mammalian cells.

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

    SciTech Connect

    Myers, G.; Korber, B.; Wain-Hobson, S.; Smith, R.F.; Pavlakis, G.N.

    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.

  5. Review of "the Twelfth West Coast Retrovirus Meeting" and "the Twenty-third Annual Symposium on Nonhuman Primate Models for AIDS".

    PubMed

    Cairns, J Scott

    2006-01-11

    Two recent meetings held on the west coast of the USA highlighted current work being done in the field of retrovirology and AIDS. The meetings, "The Twelfth West Coast Retrovirus Meeting" (Palm Springs CA; October 6-8, 2005), and the "Twenty-third Annual Symposium on Nonhuman Primate Models for AIDS" (Portland OR; September 21-24) covered a broad range of topics. The highlights covered here are not meant to be inclusive but reflect presentations of interest in the identification and development of new HIV therapies and the role played by animal models in their development.

  6. Human Endogenous Retrovirus Family HERV-K(HML-5): Status, Evolution, and Reconstruction of an Ancient Betaretrovirus in the Human Genome†

    PubMed Central

    Lavie, Laurence; Medstrand, Patrik; Schempp, Werner; Meese, Eckart; Mayer, Jens

    2004-01-01

    The human genome harbors numerous distinct families of so-called human endogenous retroviruses (HERV) which are remnants of exogenous retroviruses that entered the germ line millions of years ago. We describe here the hitherto little-characterized betaretrovirus HERV-K(HML-5) family (named HERVK22 in Repbase) in greater detail. Out of 139 proviruses, only a few loci represent full-length proviruses, and many lack gag protease and/or env gene regions. We generated a consensus sequence from multiple alignment of 62 HML-5 loci that displays open reading frames for the four major retroviral proteins. Four HML-5 long terminal repeat (LTR) subfamilies were identified that are associated with monophyletic proviral bodies, implying different evolution of HML-5 LTRs and genes. Sequence analysis indicated that the proviruses formed approximately 55 million years ago. Accordingly, HML-5 proviral sequences were detected in Old World and New World primates but not in prosimians. No recent activity is associated with this HERV family. We also conclude that the HML-5 consensus sequence primer binding site is identical to methionine tRNA. Therefore, the family should be designated HERV-M. Our study provides important insights into the structure and evolution of the oldest betaretrovirus in the primate genome known to date. PMID:15280487

  7. Thrombocytopenia in homosexual patients. Prognosis, response to therapy, and prevalence of antibody to the retrovirus associated with the acquired immunodeficiency syndrome.

    PubMed

    Walsh, C; Krigel, R; Lennette, E; Karpatkin, S

    1985-10-01

    Thirty-three homosexual patients with thrombocytopenia (mean [+/- SE] platelet count, 50 000 +/- 7000/mm3; range, 7 to 135 000/mm3) have been followed for a mean period of 20 +/- 2 months. Six patients have developed the acquired immunodeficiency syndrome 1 to 37 months after the diagnosis of thrombocytopenia. Six patients spontaneously reverted to normal platelet counts 5 to 27 months (median, 10 months) after the diagnosis of thrombocytopenia, in the absence of splenectomy and while not receiving corticosteroids. Sixteen of seventeen patients had a moderate to excellent response while on corticosteroid treatment. Ten of ten patients had an excellent response to splenectomy which has persisted. Fifteen patients did not require treatment for their thrombocytopenia. Thirteen of fourteen patients had antibody against the retrovirus associated with the acquired immunodeficiency syndrome, as did 4 of 12 homosexual controls without thrombocytopenia. Thrombocytopenia in homosexuals is part of the complex related to the acquired immunodeficiency syndrome.

  8. Single residues in the surface subunits of oncogenic sheep retrovirus envelopes distinguish receptor-mediated triggering for fusion at low pH and infection

    SciTech Connect

    Cote, Marceline; Zheng, Yi-Min; Albritton, Lorraine M.; Liu, Shan-Lu

    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 amino 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.

  9. Koala retrovirus (KoRV) genotyping analyses reveal a low prevalence of KoRV-A in Victorian koalas and an association with clinical disease.

    PubMed

    Legione, Alistair Raymond; Patterson, Jade L S; Whiteley, Pam; Firestone, Simon M; Curnick, Megan; Bodley, Kate; Lynch, Michael; Gilkerson, James R; Sansom, Fiona M; Devlin, Joanne M

    2016-12-28

    Koala retrovirus (KoRV) is currently undergoing endogenisation 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 using PCRs to detect pol and env genes. The prevalence of KoRV in these Victorian koalas was 24.7% (160/648) (95% confidence interval [CI]: 21.3, 28.1%). KoRV-A was detected in 141/160 cases but KoRV-B, a genotype associated with neoplasia in captive koalas, was not detected. Detection may have been precluded by genomic differences between KoRV in Victoria and type strains. 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% CI 1.21, 2.98). Our findings have important implications for the conservation of this iconic species, particularly in regards to translocation potential.

  10. Recombinant retrovirus-derived virus-like particle-based vaccines induce hepatitis C virus-specific cellular and neutralizing immune responses in mice.

    PubMed

    Huret, Christophe; Desjardins, Delphine; Miyalou, Mathilde; Levacher, Béatrice; Amadoudji Zin, Martin; Bonduelle, Olivia; Combadière, Béhazine; Dalba, Charlotte; Klatzmann, David; Bellier, Bertrand

    2013-03-01

    While the immunological correlates of hepatitis C virus (HCV)-specific immunity are not well understood, it is now admitted that an effective vaccine against HCV will need to induce both cellular and humoral immune responses and address viral heterogeneity to prevent immune escape. We developed a vaccine platform specifically aimed at inducing such responses against HCV antigens displayed by recombinant retrovirus-based virus-like particles (VLPs) made of Gag of murine leukemia virus. Both ex vivo produced VLPs and plasmid DNA encoding VLPs can be used as vaccines. Here, we report that immunizations with plasmid DNA forming VLPs pseudotyped with HCV E1 and E2 envelope glycoproteins (HCV-specific plasmo-retroVLPs) induce strong T-cell-mediated immune responses that can be optimized by using proper DNA delivery methods and/or genetic adjuvants. Additionally, multigenotype or multi-specific T-cell responses were observed after immunization with plasmids that encode VLPs pseudotyped with E1E2 derived from numerous viral genotypes and/or displaying NS3 antigen in capsid proteins. While homologous prime-boost immunizations with HCV-specific plasmo-retroVLPs or ex vivo produced VLPs induce a low level of specific antibody responses, optimal combination of plasmo-retroVLPs and VLPs was identified for inducing HCV-specific T-cell and B-cell responses as well as neutralizing antibodies. Altogether, these results have important meanings for the development of anti-HCV preventive vaccines and exemplify the flexibility and potential of our retrovirus-based platform in inducing broad cellular and humoral immune responses.

  11. Serial bone marrow transplantation reveals in vivo expression of the pCLPG retroviral vector

    PubMed Central

    2010-01-01

    Background Gene therapy in the hematopoietic system remains promising, though certain aspects of vector design, such as transcriptional control elements, continue to be studied. Our group has developed a retroviral vector where transgene expression is controlled by p53 with the intention of harnessing the dynamic and inducible nature of this tumor suppressor and transcription factor. We present here a test of in vivo expression provided by the p53-responsive vector, pCLPG. For this, we used a model of serial transplantation of transduced bone marrow cells. Results We observed, by flow cytometry, that the eGFP transgene was expressed at higher levels when the pCLPG vector was used as compared to the parental pCL retrovirus, where expression is directed by the native MoMLV LTR. Expression from the pCLPG vector was longer lasting, but did decay along with each sequential transplant. The detection of eGFP-positive cells containing either vector was successful only in the bone marrow compartment and was not observed in peripheral blood, spleen or thymus. Conclusions These findings indicate that the p53-responsive pCLPG retrovirus did offer expression in vivo and at a level that surpassed the non-modified, parental pCL vector. Our results indicate that the pCLPG platform may provide some advantages when applied in the hematopoietic system. PMID:20096105

  12. Epstein-Barr virus nuclear antigen 2 specifically induces expression of the B-cell activation antigen CD23

    SciTech Connect

    Wang, F.; Gregory, C.D.; Rowe, M.; Rickinson, A.B.; Wang, D.; Birkenbach, M.; Kikutani, H.; Kishimoto, T.; Kieff, E.

    1987-05-01

    Epstein-Barr virus (EBV) infection of EBV-negative Burkitt lymphoma (BL) cells includes some changes similar to those seen in normal B lymphocytes that have been growth transformed by EBV. The role of individual EBV genes in this process was evaluated by introducing each of the viral genes that are normally expressed in EBV growth-transformed and latently infected lymphoblasts into an EBV-negative BL cell line, using recombinant retrovirus-mediated transfer. Clones of cells were derived that stably express the EBV nuclear antigen 1 (EBNA-1), EBNA-2, EBNA-3, EBNA-leader protein, or EBV latent membrane protein (LMP). These were compared with control clones infected with the retrovirus vector. All 10 clones converted to EBNA-2 expression differed from control clones or clones expressing other EBV proteins by growth in tight clumps and by markedly increased expression of one particular surface marker of B-cell activation, CD23. Other activation antigens were unaffected by EBNA-2 expression, as were markers already expressed on the parent BL cell line. The results indicate that EBNA-2 is a specific direct or indirect trans-activator of CD23. This establishes a link between an EBV gene and cell gene expression. Since CD23 has been implicated in the transduction of B-cell growth signals, its specific induction by EBNA-2 could be important in EBV induction of B-lymphocyte transformation.

  13. Retrovirus-like vectors for Saccharomyces cerevisiae: integration of foreign genes controlled by efficient promoters into yeast chromosomal DNA.

    PubMed

    Jacobs, E; Dewerchin, M; Boeke, J D

    1988-07-30

    Using modified Saccharomyces cerevisiae Ty1 elements located on a 2 mu plasmid, reverse-transcriptase-mediated transposition into yeast chromosomes of expression cassettes containing a foreign gene can be induced. These expression cassettes consist of the yeast ARG3 and CUP1 promoter sequences fused to the Escherichia coli galK structural gene. Expression cassettes as large as 2 kb can be inserted into Ty elements and transposed efficiently to various sites in the yeast genome. A third yeast promoter (from the yeast CAR1 gene) seems to be unsuitable for use in the expression cassette. This may be because it does not allow the transcription run-through necessary for Ty1 transposition. Ways of improving this vector system are discussed, as are its advantages over episomal vector systems.

  14. The drug monosodium luminol (GVT) preserves crypt-villus epithelial organization and allows survival of intestinal T cells in mice infected with the ts1 retrovirus.

    PubMed

    Scofield, Virginia L; Yan, Mingshan; Kuang, Xianghong; Kim, Soo-Jin; Wong, Paul K Y

    2009-02-21

    Of the cytopathic retroviruses that affect mammals, including HIV-1, many selectively infect CD4+ T cells and cause immunosuppressive syndromes. These diseases destroy both the thymus and the small and large intestines, after infecting and killing T-lineage cells in both tissues. A mutant of the murine leukemia retrovirus MoMuLV-TB, called ts1, causes this syndrome in susceptible strains of mice. In FVB/N strain mice that are infected at birth, thymic atrophy, CD4+ T cell loss, intestinal collapse, body wasting, and death occur by approximately 30-40 days postinfection (dpi). Apoptosis of ts1-infected T-lineage cells, in the thymus, peripheral lymphoid system and intestines is caused by accumulation of the ts1 mutant viral envelope preprotein gPr80(env), which is inefficiently cleaved into the mature viral proteins gp70 and PrP15E. We show here that ts1 infection in the small intestine is followed by loss of intestinal epithelial cell (IEC) thyroid-stimulating hormone (TSH) and cell cycling gradients (along the crypt-villus axes), accumulation of gPr80(env) in intestinal cells, apoptosis of developing T cells in the lamina propria (LP), and intestinal collapse by approximately 30 dpi. In infected mice treated with the antioxidant drug monosodium luminol (GVT), however, normal intestinal epithelial cell gradients are still in place at 30 dpi, and IECs covering both the crypts and villi contain large amounts of the antioxidant transcription factor Nrf2. In addition, no apoptotic cells are present, and accumulated gpr80(env) is absent from the tissue at this time. We conclude that GVT treatment can make ts1 a noncytopathic virus for intestinal lymphoid cells, as it does for thymocytes [25]. As in the thymus, GVT may protect the intestine by reducing oxidant stress in infected intestinal T cells, perhaps by prevention of gPr80(env) accumulation via Nrf2 upregulation in the IECs. These results identify GVT as a potential therapy for intestinal diseases or inflammatory

  15. High Expression of Endogenous Retroviral Envelope Gene in the Equine Fetal Part of the Placenta

    PubMed Central

    Stefanetti, Valentina; Marenzoni, Maria Luisa; Passamonti, Fabrizio; Cappelli, Katia; Garcia-Etxebarria, Koldo; Coletti, Mauro; Capomaccio, Stefano

    2016-01-01

    Endogenous retroviruses (ERVs) are proviral phases of exogenous retroviruses that have co-evolved with vertebrate genomes for millions of years. Previous studies have identified the envelope (env) protein genes of retroviral origin preferentially expressed in the placenta which suggests a role in placentation based on their membrane fusogenic capacity and therefore they have been named syncytins. Until now, all the characterized syncytins have been associated with three invasive placentation types: the endotheliochorial (Carnivora), the synepitheliochorial (Ruminantia), and the hemochorial placentation (human, mouse) where they play a role in the syncytiotrophoblast formation. The purpose of the present study was to evaluate whether EqERV env RNA is expressed in horse tissues as well and investigate if the horse, possessing an epitheliochorial placenta, has “captured” a common retroviral env gene with syncytin-like properties in placental tissues. Interestingly, although in the equine placenta there is no syncytiotrophoblast layer at the maternal-fetal interface, our results showed that EqERV env RNA is highly expressed at that level, as expected for a candidate syncytin-like gene but with reduced abundance in the other somatic tissues (nearly 30-fold lower) thus suggesting a possible role in the placental tissue. Although the horse is one of the few domestic animals with a sequenced genome, few studies have been conducted about the EqERV and their expression in placental tissue has never been investigated. PMID:27176223

  16. 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.

  17. Characterization of retrovirus-based reporter viruses pseudotyped with the precursor membrane and envelope glycoproteins of four serotypes of dengue viruses

    SciTech Connect

    Hu, H.-P.; Hsieh, S.-C.; King, C.-C.; Wang, W.-K.

    2007-11-25

    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 inhibited 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.

  18. A human endogenous retrovirus K dUTPase triggers a TH1, TH17 cytokine response: does it have a role in psoriasis?

    PubMed

    Ariza, Maria-Eugenia; Williams, Marshall V

    2011-12-01

    Psoriasis is a chronic inflammatory immune disease of the skin characterized by a complex interplay between multiple risk genes and their interactions with environmental factors. Recent haplotype analyses have suggested that deoxyuridine triphosphate nucleotidohydrolase (dUTPase) encoded by a human endogenous retrovirus K (HERV-K) may be a candidate gene for the psoriasis susceptibility 1 locus. However, no functional studies have been conducted to determine the role of HERV-K dUTPase in psoriasis. For this purpose, we constructed an HERV-K dUTPase wild-type sequence, as well as specific mutations reflecting the genotype characteristic of high- and low-risk haplotypes, purified the recombinant proteins, and evaluated whether they could modulate innate and/or adaptive immune responses. In this study, we demonstrate that wild-type and mutant HERV-K dUTPase proteins induce the activation of NF-κB through Toll-like receptor 2, independent of enzymatic activity. Proteome array studies revealed that treatment of human primary cells with wild-type and mutant HERV-K dUTPase proteins triggered the secretion of T(H)1 and T(H)17 cytokines involved in the formation of psoriatic plaques, including IL-12p40, IL-23, IL-17, tumor necrosis factor-α, IL-8, and CCL20, in dendritic/Langerhans-like cells and to a lesser extent in keratinocytes. These data support HERV-K dUTPase as a potential contributor to psoriasis pathophysiology.

  19. Transfer of Fv-1 locus-specific resistance to murine N-tropic and B-tropic retroviruses by cytoplasmic RNA.

    PubMed Central

    Yang, W K; Tennant, R W; Rascati, R J; Otten, J A; Schluter, B; Kiggans, J O; Myer, F E; Brown, A

    1978-01-01

    A standardized bioassay for transfer of Fv-1 gene-specific resistance to N-tropic and B-tropic murine retroviruses was developed using X plaque reduction in SC-1 (Fv-1-) cells inoculated with virus. Testing of subcellular fractions of restrictive cells showed that the resistance transfer activity was present in the cytoplasmic (microsomal and cytosol) fractions. The activity of the cytoplasmic extract was destroyed by treatment with ribonuclease, but not with deoxyribonuclease or proteases. RNA prepared by phenol-chloroform extraction of mouse tissues, including embryos and livers of weanling mice, transferred Fv-1 locus-specific resistance into DEAE-dextran-treated SC-1 cells. The activity of isolated RNA preparations against virus of the appropriate host-range type has been demonstrated to correspond to the Fv-1 genotypes of the cell sources. The specific transfer of resistance with cellular RNA was effective within a 5- to 6-h period from 2 h before to 4 to 5 after virus infection. Sucrose gradient centrifugation of the RNA showed that the activity sedimented as a broad peak, with an apparent maximum in the 22S region. Affinity chromatography of whole-cell RNA on polyuridylic acid-Sepharose tended to separate more activity into the polyadenylic acid RNA fraction than the non-polyadenylic acid RNA fraction. Except for the reciprocal inhibitory activity for the two host-range virus types, the RNAs of Fv-1n and Fv-1b specificities showed similar properties in all aspects studied. PMID:211261

  20. Interferon Regulatory Factor 4 Contributes to Transformation of v-Rel-Expressing Fibroblasts

    PubMed Central

    Hrdličková, Radmila; Nehyba, Jiří; Bose, Henry R.

    2001-01-01

    The avian homologue of the interferon regulatory factor 4 (IRF-4) and a novel splice variant lacking exon 6, IRF-4ΔE6, were isolated and characterized. Chicken IRF-4 is expressed in lymphoid organs, less in small intestine, and lungs. IRF-4ΔE6 mRNA, though less abundant than full-length IRF-4, was detected in lymphoid tissues, with the highest levels observed in thymic cells. IRF-4 is highly expressed in v-Rel-transformed lymphocytes, and the expression of IRF-4 is increased in v-Rel- and c-Rel-transformed fibroblasts relative to control cells. The expression of IRF-4 from retrovirus vectors morphologically transformed primary fibroblasts, increased their saturation density, proliferation, and life span, and promoted their growth in soft agar. IRF-4 and v-Rel cooperated synergistically to transform fibroblasts. The expression of IRF-4 antisense RNA eliminated formation of soft agar colonies by v-Rel and reduced the proliferation of v-Rel-transformed cells. v-Rel-transformed fibroblasts produced interferon 1 (IFN1), which inhibits fibroblast proliferation. Infection of fibroblasts with retroviruses expressing v-Rel resulted in an increase in the mRNA levels of IFN1, the IFN receptor, STAT1, JAK1, and 2′,5′-oligo(A) synthetase. The exogenous expression of IRF-4 in v-Rel-transformed fibroblasts decreased the production of IFN1 and suppressed the expression of several genes in the IFN transduction pathway. These results suggest that induction of IRF-4 expression by v-Rel likely facilitates transformation of fibroblasts by decreasing the induction of this antiproliferative pathway. PMID:11533227

  1. Filariae-Retrovirus Co-infection in Mice is Associated with Suppressed Virus-Specific IgG Immune Response and Higher Viral Loads

    PubMed Central

    Dietze, Kirsten Katrin; Dittmer, Ulf; Koudaimi, Daniel Karim; Schimmer, Simone; Reitz, Martina

    2016-01-01

    Worldwide more than 2 billion people are infected with helminths, predominantly in developing countries. Co-infections with viruses such as human immunodeficiency virus (HIV) are common due to the geographical overlap of these pathogens. Helminth and viral infections induce antagonistic cytokine responses in their hosts. Helminths shift the immune system to a type 2-dominated immune response, while viral infections skew the cytokine response towards a type 1 immune response. Moreover, chronic helminth infections are often associated with a generalized suppression of the immune system leading to prolonged parasite survival, and also to a reduced defence against unrelated pathogens. To test whether helminths affect the outcome of a viral infection we set up a filarial/retrovirus co-infection model in C57BL/6 mice. Although Friend virus (FV) infection altered the L. sigmodontis-specific immunoglobulin response towards a type I associated IgG2 isotype in co-infected mice, control of L. sigmodontis infection was not affected by a FV-superinfection. However, reciprocal control of FV infection was clearly impaired by concurrent L. sigmodontis infection. Spleen weight as an indicator of pathology and viral loads in spleen, lymph nodes (LN) and bone marrow (BM) were increased in L. sigmodontis/FV-co-infected mice compared to only FV-infected mice. Numbers of FV-specific CD8+ T cells as well as cytokine production by CD4+ and CD8+ cells were alike in co-infected and FV-infected mice. Increased viral loads in co-infected mice were associated with reduced titres of neutralising FV-specific IgG2b and IgG2c antibodies. In summary our findings suggest that helminth infection interfered with the control of retroviral infection by dampening the virus-specific neutralising antibody response. PMID:27923052

  2. Adaptation of Chimeric Retroviruses In Vitro and In Vivo: Isolation of Avian Retroviral Vectors with Extended Host Range

    PubMed Central

    Barsov, Eugene V.; Payne, William S.; Hughes, Stephen H.

    2001-01-01

    We have designed and characterized two new replication-competent avian sarcoma/leukosis virus-based retroviral vectors with amphotropic and ecotropic host ranges. The amphotropic vector RCASBP-M2C(797-8), was obtained by passaging the chimeric retroviral vector RCASBP-M2C(4070A) (6) in chicken embryos. The ecotropic vector, RCASBP(Eco), was created by replacing the env-coding region in the retroviral vector RCASBP(A) with the env region from an ecotropic murine leukemia virus. It replicates efficiently in avian DFJ8 cells that express murine ecotropic receptor. For both vectors, permanent cell lines that produce viral stocks with titers of about 5 × 106 CFU/ml on mammalian cells can be easily established by passaging transfected avian cells. Some chimeric viruses, for example, RCASBP(Eco), replicate efficiently without modifications. For those chimeric viruses that do require modification, adaptation by passage in vitro or in vivo is a general strategy. This strategy has been used to prepare vectors with altered host range and could potentially be used to develop vectors that would be useful for targeted gene delivery. PMID:11333876

  3. Adaptation of chimeric retroviruses in vitro and in vivo: isolation of avian retroviral vectors with extended host range.

    PubMed

    Barsov, E V; Payne, W S; Hughes, S H

    2001-06-01

    We have designed and characterized two new replication-competent avian sarcoma/leukosis virus-based retroviral vectors with amphotropic and ecotropic host ranges. The amphotropic vector RCASBP-M2C(797-8), was obtained by passaging the chimeric retroviral vector RCASBP-M2C(4070A) (6) in chicken embryos. The ecotropic vector, RCASBP(Eco), was created by replacing the env-coding region in the retroviral vector RCASBP(A) with the env region from an ecotropic murine leukemia virus. It replicates efficiently in avian DFJ8 cells that express murine ecotropic receptor. For both vectors, permanent cell lines that produce viral stocks with titers of about 5 x 10(6) CFU/ml on mammalian cells can be easily established by passaging transfected avian cells. Some chimeric viruses, for example, RCASBP(Eco), replicate efficiently without modifications. For those chimeric viruses that do require modification, adaptation by passage in vitro or in vivo is a general strategy. This strategy has been used to prepare vectors with altered host range and could potentially be used to develop vectors that would be useful for targeted gene delivery.

  4. Human Endogenous Retrovirus K(HML-2) Gag- and Env-Specific T-Cell Responses Are Infrequently Detected in HIV-1-Infected Subjects Using Standard Peptide Matrix-Based Screening

    PubMed Central

    John, Vivek M.; Hunter, Diana V.; Martin, Eric; Mujib, Shariq; Mihajlovic, Vesna; Burgers, Peter C.; Luider, Theo M.; Gyenes, Gabor; Sheppard, Neil C.; SenGupta, Devi; Tandon, Ravi; Yue, Feng-Yun; Benko, Erika; Kovacs, Colin; Nixon, Douglas F.; Ostrowski, Mario A.

    2012-01-01

    T-cell responses to human endogenous retrovirus (HERV) K(HML-2) Gag and Env were mapped in HIV-1-infected subjects using 15mer peptides. Small peptide pools and high concentrations were used to maximize sensitivity. In the 23 subjects studied, only three bona fide HERV-K(HML-2)-specific responses were detected. At these high peptide concentrations, we detected false-positive responses, three of which were mapped to an HIV-1 Gag peptide contaminant. Thus, HERV-K(HML-2) Gag- and Env-specific T-cell responses are infrequently detected by 15mer peptide mapping. PMID:22205657

  5. Genome-wide Profiling Reveals Remarkable Parallels Between Insertion Site Selection Properties of the MLV Retrovirus and the piggyBac Transposon in Primary Human CD4+ T Cells

    PubMed Central

    Gogol-Döring, Andreas; Ammar, Ismahen; Gupta, Saumyashree; Bunse, Mario; Miskey, Csaba; Chen, Wei; Uckert, Wolfgang; Schulz, Thomas F; Izsvák, Zsuzsanna; Ivics, Zoltán

    2016-01-01

    The inherent risks associated with vector insertion in gene therapy need to be carefully assessed. We analyzed the genome-wide distributions of Sleeping Beauty (SB) and piggyBac (PB) transposon insertions as well as MLV retrovirus and HIV lentivirus insertions in human CD4+ T cells with respect to a panel of 40 chromatin states. The distribution of SB transposon insertions displayed the least deviation from random, while the PB transposon and the MLV retrovirus showed unexpected parallels across all chromatin states. Both MLV and PB insertions are enriched at transcriptional start sites (TSSs) and co-localize with BRD4-associated sites. We demonstrate physical interaction between the PB transposase and bromodomain and extraterminal domain proteins (including BRD4), suggesting convergent evolution of a tethering mechanism that directs integrating genetic elements into TSSs. We detect unequal biases across the four systems with respect to targeting genes whose deregulation has been previously linked to serious adverse events in gene therapy clinical trials. The SB transposon has the highest theoretical chance of targeting a safe harbor locus in the human genome. The data underscore the significance of vector choice to reduce the mutagenic load on cells in clinical applications. PMID:26755332

  6. Cleavage of tRNA within the mature tRNA sequence by the catalytic RNA of RNase P: implication for the formation of the primer tRNA fragment for reverse transcription in copia retrovirus-like particles.

    PubMed Central

    Kikuchi, Y; Sasaki, N; Ando-Yamagami, Y

    1990-01-01

    The retrovirus-like particles of Drosophila are intermediates of retrotransposition of the transposable element copia. In these particles, a 39-nucleotide-long fragment from the 5' region of Drosophila initiator methionine tRNA (tRNA(iMet) is used as the primer for copia minus-strand reverse transcription. To function as primer for this reverse transcription, the Drosophila tRNA(iMet) must be cleaved in vivo at the site between nucleotides 39 and 40. When a synthetic Drosophila tRNA(iMet) precursor was incubated with M1RNA, the catalytic RNA of Escherichia coli RNase P, other cleavages within the mature tRNA sequence were detected in addition to the efficient removal of the 5' leader sequence of this tRNA precursor. One of these cleavage sites is between nucleotides 39 and 40 of Drosophila tRNA(iMet). Based on this result, we propose a model for formation of the primer tRNA fragment for reverse transcription in copia retrovirus-like particles. Images PMID:1700426

  7. Expression of human factor IX in rat capillary endothelial cells: Toward somatic gene therapy for hemophilia B

    SciTech Connect

    Shounan Yao; Wilson, J.M.; Nabel, E.G.; Kurachi, Sumiko; Hachiya, H.L.; Kurachi, Kotoku )

    1991-09-15

    In aiming to develop a gene therapy approach for hemophilia B, the authors expressed and characterized human factor IX in rat capillary endothelial cells (CECs). Moloney murine leukemia virus-derived retrovirus vectors that contain human factor IX cDNA linked to heterologous promoters and the neomycin-resistant gene were constructed and employed to prepare recombinant retroviruses. Rat CECs and NIH 3T3 cells infected with these viruses were selected with the neomycin analogue, G418 sulfate, and tested for expression of factor IX. A construct with the factor IX cDNA under direct control by long terminal repeat gave the highest level of expression as quantitated by immunoassays as well as clotting activity assays. A single RNA transcript of 4.4 kilobases predicted by the construct and a recombinant factor IX were found. The recombinant human factor IX produced showed full clotting activity, demonstrating that CECs have an efficient mechanism for posttranslational modifications, including {gamma}-carboxylation, essential for its biological activity. These results, in addition to other properties of the endothelium, including large number of cells, accessibility, and direct contact with the circulating blood, suggest that CECs can serve as an efficient drug delivery vehicle producing factor IX in a somatic gene therapy for hemophilia B.

  8. Evolutionary conservation of a molecular machinery for export and expression of mRNAs with retained introns

    PubMed Central

    Wang, Baomin; Rekosh, David

    2015-01-01

    Intron retention is one of the least studied forms of alternative splicing. Through the use of retrovirus and other model systems, it was established many years ago that mRNAs with retained introns are subject to restriction both at the level of nucleocytoplasmic export and cytoplasmic expression. It was also demonstrated that specific cis-acting elements in the mRNA could serve to bypass these restrictions. Here we show that one of these elements, the constitutive transport element (CTE), first identified in the retrovirus MPMV and subsequently in the human NXF1 gene, is a highly conserved element. Using GERP analysis, CTEs with strong primary sequence homology, predicted to display identical secondary structure, were identified in NXF genes from >30 mammalian species. CTEs were also identified in the predicted NXF1 genes of zebrafish and coelacanths. The CTE from the zebrafish NXF1 was shown to function efficiently to achieve expression of mRNA with a retained intron in human cells in conjunction with zebrafish Nxf1 and cofactor Nxt proteins. This demonstrates that all essential functional components for expression of mRNA with retained introns have been conserved from fish to man. PMID:25605961

  9. 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

  10. Recent patents on alphavirus protein expression and vector production.

    PubMed

    Aranda, Alejandro; Ruiz-Guillen, Marta; Quetglas, Jose I; Bezunartea, Jaione; Casales, Erkuden; Smerdou, Cristian

    2011-12-01

    Alphaviruses contain a single-strand RNA genome that can be modified to express heterologous genes at high levels. Alphavirus vectors can be packaged within viral particles (VPs) or used as DNA/RNA layered systems. The broad tropism and high expression levels of alphavirus vectors have made them very attractive for applications like recombinant protein expression, vaccination or gene therapy. Expression mediated by alphavirus vectors is generally transient due to induction of apoptosis. However, during the last years several non-cytopathic mutations have been identified within the replicase sequence of different alphaviruses, allowing prolonged protein expression in culture cells. Some of these mutants, which have been patented, have allowed the generation of stable cell lines able to express recombinant proteins for extended periods of time in a constitutive or inducible manner. Production of alphavirus VPs usually requires cotransfection of cells with vector and helper RNAs providing viral structural proteins in trans. During this process full-length wild type (wt) genomes can be generated through recombination between different RNAs. Several new strategies to reduce wt virus generation during packaging, optimize VP production, increase packaging capacity, and provide VPs with specific targeting have been recently patented. Finally, hybrid vectors between alphavirus and other types of viruses have led to a number of patents with applications in vaccination, cancer therapy or retrovirus production.

  11. Cytotoxic immune response after retroviral-mediated hepatic gene transfer in rat does not preclude expression from adeno-associated virus 1 transduced muscles.

    PubMed

    Aubert, Dominique; Pichard, Virginie; Durand, Sophie; Moullier, Philippe; Ferry, Nicolas

    2003-03-20

    Intravenous delivery of nls-lacZ retroviral vectors to the regenerating liver triggers a cytotoxic immune response directed against transduced hepatocytes. We sought to determine whether prior immunization with retroviral vectors impacted on adeno-associated virus (AAV)-mediated muscular expression of the same transgene. The first group of rats first received nls-lacZ retroviral vectors intravenously after a partial hepatectomy. Thirty days later they received AAV vectors intramuscularly in both legs. In the second group, animals received the same vectors in the opposite sequence (i.e., AAV first and retroviruses 20 days later). In the first group, immune response occurred after retrovirus delivery with appearance of anti-beta-galactosidase antibodies and elimination of transduced hepatocytes. However, the immune response did not prevent sustained (9-month) beta-galactosidase expression in AAV-injected muscles. In the second group, AAV injections did not induce immune response and resulted in beta-galactosidase expression in myofibers. In this group, subsequent delivery of retroviral vectors triggered appearance of immune response and elimination of transduced hepatocytes. However, the immune response did not modify beta-galactosidase expression in AAV-transduced myofibers for up to 9 months. These results demonstrate a differential susceptibility between retrovirally transduced liver and AAV-transduced muscles to immune response against the transgene product.

  12. SARS coronavirus, but not human coronavirus NL63, utilizes cathepsin L to infect ACE2-expressing cells.

    PubMed

    Huang, I-Chueh; Bosch, Berend Jan; Li, Fang; Li, Wenhui; Lee, Kyoung Hoa; Ghiran, Sorina; Vasilieva, Natalya; Dermody, Terence S; Harrison, Stephen C; Dormitzer, Philip R; Farzan, Michael; Rottier, Peter J M; Choe, Hyeryun

    2006-02-10

    Viruses require specific cellular receptors to infect their target cells. Angiotensin-converting enzyme 2 (ACE2) is a cellular receptor for two divergent coronaviruses, SARS coronavirus (SARS-CoV) and human coronavirus NL63 (HCoV-NL63). In addition to hostcell receptors, lysosomal cysteine proteases are required for productive infection by some viruses. Here we show that SARS-CoV, but not HCoV-NL63, utilizes the enzymatic activity of the cysteine protease cathepsin L to infect ACE2-expressing cells. Inhibitors of cathepsin L blocked infection by SARS-CoV and by a retrovirus pseudotyped with the SARS-CoV spike (S) protein but not infection by HCoV-NL63 or a retrovirus pseudotyped with the HCoV-NL63 S protein. Expression of exogenous cathepsin L substantially enhanced infection mediated by the SARS-CoV S protein and by filovirus GP proteins but not by the HCoV-NL63 S protein or the vesicular stomatitis virus G protein. Finally, an inhibitor of endosomal acidification had substantially less effect on infection mediated by the HCoV-NL63 S protein than on that mediated by the SARS-CoV S protein. Our data indicate that two coronaviruses that utilize a common receptor nonetheless enter cells through distinct mechanisms.

  13. Expression of Dual-Specificity Phosphatase 5 Pseudogene 1 (DUSP5P1) in Tumor Cells

    PubMed Central

    Staege, Martin S.; Müller, Katja; Kewitz, Stefanie; Volkmer, Ines; Mauz-Körholz, Christine; Bernig, Toralf; Körholz, Dieter

    2014-01-01

    Sequencing of individual clones from a newly established cDNA library from the chemoresistant Hodgkin's lymphoma cell line L-1236 led to the isolation of a cDNA clone corresponding to a short sequence from chromosome 1. Reverse transcriptase-polymerase chain reaction indicated high expression of this sequence in Hodgkin's lymphoma derived cell lines but not in normal blood cells. Further characterization of this sequence and the surrounding genomic DNA revealed that this sequence is part of a human endogenous retrovirus locus. The sequence of this endogenous retrovirus is interrupted by a pseudogene of the dual specificity phosphatase 5 (DUSP5). Reverse transcriptase-polymerase chain reaction revealed high expression of this pseudogene (DUSP5P1) in HL cell lines but not in normal blood cells or Epstein-Barr virus-immortalized B cells. Cells from other tumor types (Burkitt's lymphoma, leukemia, neuroblastoma, Ewing sarcoma) also showed a higher DUSP5P1/DUSP5 ratio than normal cells. Furthermore, we observed that higher expression of DUSP5 in relation to DUSP5P1 correlated with the expression of the pro-apoptotic factor B cell leukemia/lymphoma 2-like 11 (BCL2L11) in peripheral blood cells and HL cells. Knock-down of DUSP5 in HL cells resulted in down-regulation of BCL2L11. Thus, the DUSP5/DUSP5P1 system could be responsible for regulation of BCL2L11 leading to inhibition of apoptosis in these tumor cells. PMID:24651368

  14. HTLV-1 p30II: selective repressor of gene expression.

    PubMed

    Green, Patrick L

    2004-11-24

    Human T-lymphotropic virus type-1 (HTLV-1) is a complex retrovirus that causes adult T-cell leukemia/lymphoma (ATL) and is implicated in a variety of lymphocyte-mediated disorders. HTLV-1 pX ORF II encodes two proteins, p13II and p30II whose roles are beginning to be defined in the virus life cycle. Previous studies indicate the importance of these viral proteins in the ability of the virus to maintain viral loads and persist in an animal model of HTLV-1 infection. Intriguing new studies indicate that p30II is a multifunctional regulator that differentially modulates CREB and Tax-responsive element-mediated transcription through its interaction with CREB-binding protein (CBP)/p300 and specifically binds and represses tax/rex mRNA nuclear export. A new study characterized the role of p30II in regulation of cellular gene expression using comprehensive human gene arrays. Interestingly, p30II is an overall repressor of cellular gene expression, while selectively favoring the expression of regulatory gene pathways important to T lymphocytes. These new findings suggest that HTLV-1, which is associated with lymphoproliferative diseases, uses p30II to selectively repress cellular and viral gene expression to favor the survival of cellular targets ultimately resulting in leukemogenesis.

  15. EP300-ZNF384 fusion gene product up-regulates GATA3 gene expression and induces hematopoietic stem cell gene expression signature in B-cell precursor acute lymphoblastic leukemia cells.

    PubMed

    Yaguchi, Akinori; Ishibashi, Takeshi; Terada, Kazuki; Ueno-Yokohata, Hitomi; Saito, Yuya; Fujimura, Junya; Shimizu, Toshiaki; Ohki, Kentaro; Manabe, Atsushi; Kiyokawa, Nobutaka

    2017-04-04

    ZNF384-related fusion genes are associated with a distinct subgroup of B-cell precursor acute lymphoblastic leukemias in childhood, with a frequency of approximately 3-4%. We previously identified a novel EP300-ZNF384 fusion gene. Patients with the ZNF384-related fusion gene exhibit a hematopoietic stem cell (HSC) gene expression signature and characteristic immunophenotype with negative or low expression of CD10 and aberrant expression of myeloid antigens, such as CD33 and CD13. However, the molecular basis of this pathogenesis remains completely unknown. In the present study, we examined the biological effects of EP300-ZNF384 expression induced by retrovirus-mediated gene transduction in an REH B-cell precursor acute lymphoblastic leukemia cell line, and observed the acquisition of the HSC gene expression signature and an up-regulation of GATA3 gene expression, as assessed by microarray analysis. In contrast, the gene expression profile induced by wild-type ZNF384 in REH cells was significantly different from that by EP300-ZNF384 expression. Together with the results of reporter assays, which revealed the enhancement of GATA3-promoter activity by EP300-ZNF384 expression, these findings suggest that EP300-ZNF384 mediates GATA3 gene expression and may be involved in the acquisition of the HSC gene expression signature and characteristic immunophenotype in B-cell precursor acute lymphoblastic leukemia cells.

  16. Negative regulation in correct tissue-specific expression of mouse mammary tumor virus in transgenic mice.

    PubMed Central

    Ross, S R; Hsu, C L; Choi, Y; Mok, E; Dudley, J P

    1990-01-01

    Mouse mammary tumor virus (MMTV) is an endogenous murine retrovirus that is expressed in the epithelial cells of the mammary and salivary glands, lungs, kidneys, and seminal vesicles and in the lymphoid cells of the spleen and thymus. Several studies have shown that the long terminal repeat (LTR) of this virus can direct the expression of reporter genes to the same tissues in transgenic mice. To determine whether multiple regulatory elements within the LTR are involved in this tissue-specific expression, we have established lines of transgenic mice containing transgenes that have deletions in the MMTV LTR. Deletions of all LTR sequences upstream of -364 or of LTR sequences from -165 to -665 both result in the expression of linked reporter genes such as the simian virus 40 early region or the bacterial enzyme chloramphenicol acetyltransferase in novel sites, such as the heart, brain, and skeletal muscle; expression of endogenous MMTV and transgenes containing the full-length LTR is not detected in these organs. Negative regulation appears to involve more than one region, since deletion of sequences between either -201 and -471 or -201 and -344, as well as sequences upstream of -364, results in inappropriate expression in heart, brain, and skeletal muscle. Therefore, a negative regulatory element(s) in the MMTV LTR can suppress transcription from the viral promoter in several different organs. This represents the first example of generalized negative regulatory elements that act in many different tissues in transgenic mice to prevent inappropriate expression of a gene. Images PMID:1700274

  17. Differential expression of perforin in cytotoxic lymphocyte in HIV/AIDS patients of China.

    PubMed

    Qi, Wang; Yongjun, Jiang; Yanan, Wang; Zining, Zhang; Xiaoxu, Han; Jing, Liu; Hong, Shang

    2006-07-01

    Cytotoxic lymphocytes are critical in the control of HIV replication, it has been shown that perforin is the key effector of killing machinery for CTLs and NK cells, so we investigated the circulating levels of perforin in CD8+ T cells and NK cells by flow cytometry intracellular stain in Chinese HIV infected individuals, its association with disease progression was analyzed. Our results showed that NK cells express perforin more efficiently than CD8+ T cells, CD8+ T cells expressed perforin higher than that of healthy controls, but NK cells expressed lower perforin than that of healthy controls, both were not correlated with disease progression. but significantly associated with their numbers, anti-retrovirus therapy had no evident effects on peforin expression in CD8+ T cells, but enhanced perfrin expression in NK cells, perforin expression in CD8+ T cells and CD16+ NK cells correlate with CD4+ T cell counts significantly in HAART-treated group. Therefore, different mechanisms may be involved in regulating peripheral perforin expression in different cell types.

  18. 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.

  19. Basic calcium phosphate crystal-induced Egr-1 expression stimulates mitogenesis in human fibroblasts

    SciTech Connect

    Zeng, Xiao R.; Sun Yubo; Wenger, Leonor; Cheung, Herman S. . E-mail: hcheung@med.miami.edu

    2005-05-13

    Previously, we have reported that basic calcium phosphate (BCP) crystals stimulate mitogenesis and synthesis of matrix metalloproteinases in cultured human foreskin and synovial fibroblasts. However, the detailed mechanisms involved are still unclear. In the present study, using RT-PCR and Egr-1 promoter analysis we showed that BCP crystals could stimulate early growth response gene Egr-1 transcription through a PKC{alpha}-dependent p44/p42 MAPK pathway. Using a retrovirus gene expression system (Clontech) to overexpress Egr-1 in human fibroblast BJ-1 cells resulted in promotion of mitogenesis measured either by MTT cell proliferation analysis or by direct cell counting. The results demonstrate that Egr-1 may play a key role in mediating BCP crystal-induced synovial fibroblast mitogenesis.

  20. Identification of developmental competence-related genes in mature porcine oocytes.

    PubMed

    Yuan, Ye; Ida, Jennifer M; Paczkowski, Melissa; Krisher, Rebecca L

    2011-08-01

    Oocyte competence is a key factor limiting female fertility, yet the underlying molecular mechanisms that contribute to oocyte competence remain unclear. The objective of this study was to elucidate specific genes whose function contributes to oocyte competence. We observed that 6 of 20 target genes examined were differentially expressed between adult (more competent) and prepubertal (less competent) porcine in vitro matured (IVM) oocytes. These genes were the cholesterol synthesis-related gene HMG-CoA reductase (HMGCR), fatty acid oxidation genes acyl-CoA synthetase long-chain family member 3 (ACSL3) and long-chain acyl-CoA dehydrogenase (ACADL), glycolytic genes fructose 1,6 bisphosphate aldolase (ALDOA) and lactate dehydrogenase C (LDHC), and tumor necrosis factor-α (TNF). These 6 genes, as well as 3 other genes [porcine endogenous retrovirus (PERV), transcribed loci 10 (TL10), serine/arginine-rich splicing factor 1 (SRSF1)], were further analyzed by comparing transcript abundance in IVM and in vivo matured (VVM) prepubertal and adult porcine oocytes. Among these 9 target genes, 5 were differentially expressed between IVM and VVM prepubertal oocytes, while 8 genes were differentially expressed between IVM and VVM adult oocytes. No genes were differentially expressed between VVM prepubertal and adult oocytes. A functional study of TNF demonstrated that depletion of endogenous TNF decreased oocyte competence and TNFAIP6 expression in cumulus cells, while TNF in IVM medium regulated TNFAIP6 expression in cumulus cells. Differential expression of the genes identified in this study suggests that these genes may be functionally relevant to oocyte competence.

  1. Elevated HERV-K mRNA expression in PBMC is associated with a prostate cancer diagnosis particularly in older men and smokers.

    PubMed

    Wallace, Tiffany A; Downey, Ronan F; Seufert, Caleb J; Schetter, Aaron; Dorsey, Tiffany H; Johnson, Carol A; Goldman, Radoslav; Loffredo, Christopher A; Yan, Peisha; Sullivan, Francis J; Giles, Francis J; Wang-Johanning, Feng; Ambs, Stefan; Glynn, Sharon A

    2014-09-01

    Aberrant expression of subgroup k human endogenous retroviruses (HERV-K) has been observed in prostate cancer. This subgroup is unique because it encodes sequences in the human genome containing open reading frames for near intact retroviruses. We hypothesized that HERV-K reactivation could serve as a non-invasive early disease detection marker for prostate cancer. We evaluated HERV-K gag messenger RNA (mRNA) expression in blood samples of African-American and European-American men using a case-control design via quantitative real-time PCR. Additionally, we examined HERV-K envelope protein expression in prostate tumors by immunohistochemistry. HERV-K envelope protein was commonly upregulated in prostate tumors, but more so in tumors of African-American than European-American patients (61% versus 40%, P < 0.01). Examining HERV-K gag expression in peripheral blood mononuclear cells (PBMC) from 294 cases and 135 healthy men, we found that the abundance of HERV-K gag message was significantly higher in cases than controls and was associated with increased plasma interferon-γ. Men with gag expression in the highest quartile had >12-fold increased odds {odds ratio = 12.87 [95% confidence interval 6.3-26.25]} of being diagnosed with prostate cancer than those in the lowest quartile. Moreover, our results showed that HERV-K expression may perform better as a disease biomarker in older than younger men (whereas the sensitivity of prostate-specific antigen (PSA) testing decreases with age) and in men with a smoking history compared with never smokers. Combining non-invasive HERV-K testing with PSA testing may improve the efficacy of prostate cancer detection specifically among older men and smokers who tend to develop a more aggressive disease.

  2. Increased proliferation and chemosensitivity of human mesenchymal stromal cells expressing fusion yeast cytosine deaminase.

    PubMed

    Kucerova, Lucia; Poturnajova, Martina; Tyciakova, Silvia; Matuskova, Miroslava

    2012-03-01

    Mesenchymal stromal cells (MSCs) are considered to be suitable vehicles for cellular therapy in various conditions. The expression of reporter and/or effector protein(s) enabled both the identification of MSCs within the organism and the exploitation in targeted tumor therapies. The aim of this study was to evaluate cellular changes induced by retrovirus-mediated transgene expression in MSCs in vitro. Human Adipose Tissue-derived MSCs (AT-MSCs) were transduced to express (i) the enhanced green fluorescent protein (EGFP) reporter transgene, (ii) the fusion yeast cytosine deaminase::uracil phosphoribosyltransferase (CDy::UPRT) enzyme along with the expression of dominant positive selection gene NeoR or (iii) the selection marker NeoR alone (MOCK). CDy::UPRT expression resulted in increased proliferation of CDy::UPRT-MSCs versus naïve AT-MSCs, MOCK-MSCs or EGFP-MSCs. Furthermore, CDy::UPRT-MSCs were significantly more sensitive to 5-fluorouracil (5FU), cisplatin, cyclophosphamide and cytosine arabinoside as determined by increased Caspase 3/7 activation and/or decreased relative proliferation. CDy::UPRT-MSCs in direct cocultures with breast cancer cells MDA-MB-231 increased tumor cell killing induced by low concentrations of 5FU. Our data demonstrated the changes in proliferation and chemoresistance in engineered MSCs expressing transgene with enzymatic function and suggested the possibilities for further augmentation of targeted MSC-mediated antitumor therapy.

  3. Stable knockdown of CREB, HIF-1 and HIF-2 by replication-competent retroviruses abrogates the responses to hypoxia in hepatocellular carcinoma

    PubMed Central

    Shneor, D; Folberg, R; Pe'er, J; Honigman, A; Frenkel, S

    2017-01-01

    The fast proliferation of tumor cells develops faster than the vasculature, resulting, in most malignant tumors, in generation of hypoxic regions. Hypoxia renders solid tumors resistant to radiation and chemotherapeutics while providing opportunities for tumor-selective therapies targeting tumor hypoxia. Here we exploit two properties of tumors: propagation of tumor cells and ongoing generation of hypoxic regions to construct a system that preferentially leads to the death of tumor cells and thus hinders tumor growth. We constructed murine leukemia virus replication-competent (RCR) viruses that infect only propagating cells. These viruses express small hairpin RNAs (shRNAs) targeting cyclic AMP-response-element binding protein (CREB), hypoxia-inducible factors 1 (HIF)-1 or HIF-2 individually or all three together (X3). These viruses efficiently infected in vitro human hepatocellular carcinoma (HepG2 and FLC4) cells and established persistence of the virus and knocked down the expression of the regulators of the hypoxia-responding genes. Knockdown of either HIF-1 or CREB or both in hypoxia reduced the expression of hypoxia-response elements- and CRE-mediated gene expression, diminished cell proliferation and increased caspase-3 activity. We did not detect any significant effect of the efficiently knocked down HIF-2 on any of the functions tested in vitro. Moreover, severe combined immunodeficiency mice implanted subcutaneously with HepG2 stably infected with recombinant RCRs showed reduction of tumor growth and vascular endothelial growth factor expression, and no hypoxia-guided neovascularization. Combined treatment (RCRs+doxorubicin) improved efficacy in the context of in vitro hypoxia and in vivo (with either vACE-CREB or vACE-X3). This synergistic effect may lead to an improved efficacy and safety profile of the treatment that may result in fewer side effects. PMID:27934882

  4. Generation of cloned transgenic cats expressing red fluorescence protein.

    PubMed

    Yin, Xi Jun; Lee, Hyo Sang; Yu, Xian Feng; Choi, Eugene; Koo, Bon Chul; Kwon, Mo Sun; Lee, Young S; Cho, Su Jin; Jin, Guang Zhen; Kim, Lyoung Hyo; Shin, Hyoung Doo; Kim, Teoan; Kim, Nam Hyung; Kong, Il Keun

    2008-03-01

    A method for engineering and producing genetically modified cats is important for generating biomedical models of human diseases. Here we describe the use of somatic cell nuclear transfer to produce cloned transgenic cats that systemically express red fluorescent protein. Immature oocytes were collected from superovulating cat ovaries. Donor fibroblasts were obtained from an ear skin biopsy of a white male Turkish Angora cat, cultured for one to two passages, and subjected to transduction with a retrovirus vector designed to transfer and express the red fluorescent protein (RFP) gene. A total of 176 RFP cloned embryos were transferred into 11 surrogate mothers (mean = 16 +/- 7.5 per recipient). Three surrogate mothers were successfully impregnated (27.3%) and delivered two liveborn and one stillborn kitten at 65 to 66 days of gestation. Analysis of nine feline-specific microsatellite loci confirmed that the cloned cats were genetically identical to the donor cat. Presence of the RFP gene in the transgenic cat genome was confirmed by PCR and Southern blot analyses. Whole-body red fluorescence was detected 60 days after birth in the liveborn transgenic (TG) cat but not in the surrogate mother cat. Red fluorescence was detected in tissue samples, including hair, muscle, brain, heart, liver, kidney, spleen, bronchus, lung, stomach, intestine, tongue, and even excrement of the stillborn TG cat. These results suggest that this nuclear transfer procedure using genetically modified somatic cells could be useful for the efficient production of transgenic cats.

  5. Glutathione Depletion Is Linked with Th2 Polarization in Mice with a Retrovirus-Induced Immunodeficiency Syndrome, Murine AIDS: Role of Proglutathione Molecules as Immunotherapeutics

    PubMed Central

    Brundu, Serena; Palma, Linda; Picceri, Giusi Giada; Ligi, Daniela; Orlandi, Chiara; Galluzzi, Luca; Chiarantini, Laura; Casabianca, Anna; Schiavano, Giuditta Fiorella; Santi, Martina; Mannello, Ferdinando; Smietana, Michaël; Magnani, Mauro

    2016-01-01

    ABSTRACT Injection of the LP-BM5 murine leukemia virus into mice causes murine AIDS, a disease characterized by many dysfunctions of immunocompetent cells. To establish whether the disease is characterized by glutathione imbalance, reduced glutathione (GSH) and cysteine were quantified in different organs. A marked redox imbalance, consisting of GSH and/or cysteine depletion, was found in the lymphoid organs, such as the spleen and lymph nodes. Moreover, a significant decrease in cysteine and GSH levels in the pancreas and brain, respectively, was measured at 5 weeks postinfection. The Th2 immune response was predominant at all times investigated, as revealed by the expression of Th1/Th2 cytokines. Furthermore, investigation of the activation status of peritoneal macrophages showed that the expression of genetic markers of alternative activation, namely, Fizz1, Ym1, and Arginase1, was induced. Conversely, expression of inducible nitric oxide synthase, a marker of classical activation of macrophages, was detected only when Th1 cytokines were expressed at high levels. In vitro studies revealed that during the very early phases of infection, GSH depletion and the downregulation of interleukin-12 (IL-12) p40 mRNA were correlated with the dose of LP-BM5 used to infect the macrophages. Treatment of LP-BM5-infected mice with N-(N-acetyl-l-cysteinyl)-S-acetylcysteamine (I-152), an N-acetyl-cysteine supplier, restored GSH/cysteine levels in the organs, reduced the expression of alternatively activated macrophage markers, and increased the level of gamma interferon production, while it decreased the levels of Th2 cytokines, such as IL-4 and IL-5. Our findings thus establish a link between GSH deficiency and Th1/Th2 disequilibrium in LP-BM5 infection and indicate that I-152 can be used to restore the GSH level and a balanced Th1/Th2 response in infected mice. IMPORTANCE The first report of an association between Th2 polarization and alteration of the redox state in LP-BM5

  6. Domain structure of the Moloney murine leukemia virus reverse transcriptase: mutational analysis and separate expression of the DNA polymerase and RNase H activities.

    PubMed Central

    Tanese, N; Goff, S P

    1988-01-01

    The reverse transcriptase of Moloney murine leukemia virus, like that of all retroviruses, exhibits a DNA polymerase activity capable of synthesis on RNA or DNA templates and an RNase H activity with specificity for RNA in the form of an RNA.DNA hybrid. We have generated a library of linker insertion mutants of the Moloney murine leukemia virus enzyme expressed in bacteria and assayed these mutants for both enzymatic activities. Those mutations affecting the DNA polymerase activity were clustered in the 5'-proximal two-thirds of the gene, and those affecting RNase H were in the remaining 3' one-third. Based on these maps, plasmids were made that expressed each one of the domains separately; assays of the proteins encoded by these plasmids showed that each domain exhibited only the expected activity. Images PMID:2450347

  7. 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

  8. 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…

  9. New Phase of Growth for Xenogeneic-Based Bioartificial Organs.

    PubMed

    Pitkin, Zorina

    2016-09-21

    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.

  10. 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

  11. Symbiotic Expressions

    NASA Astrophysics Data System (ADS)

    Bernecky, Robert; Herhut, Stephan; Scholz, Sven-Bodo

    We introduce symbiotic expressions, a method for algebraic simplification within a compiler, in lieu of an SMT solver, such as Yices or the Omega Calculator. Symbiotic expressions are compiler-generated expressions, temporarily injected into a program's abstract syntax tree (AST). The compiler's normal optimizations interpret and simplify those expressions, making their results available for the compiler to use as a basis for decisions about further optimization of the source program. The expressions are symbiotic, in the sense that both parties benefit: an optimization benefits, by using the compiler itself to simplify expressions that have been attached, lamprey-like, to the AST by the optimization; the program being compiled benefits, from improved run-time in both serial and parallel environments.

  12. Fine tuning of the temporal expression of HTLV-1 and HTLV-2

    PubMed Central

    Cavallari, Ilaria; Rende, Francesca; Bender, Cecilia; Romanelli, Maria G.; D'Agostino, Donna M.; Ciminale, Vincenzo

    2013-01-01

    Human T-cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2) are delta retroviruses that share a common overall genetic organization, splicing pattern, and ability to infect and immortalize T-cells in vitro. However, HTLV-1 and HTLV-2 exhibit a clearly distinct pathogenic potential in infected patients. To find clues to the possible viral determinants of the biology of these viruses, recent studies investigated the timing of expression and the intracellular compartmentalization of viral transcripts in ex-vivo samples from infected patients. Results of these studies revealed a common overall pattern of expression of HTLV-1 and -2 with a two-phase kinetics of expression and a nuclear accumulation of minus-strand transcripts. Studies in cells transfected with HTLV-1 molecular clones demonstrated the strict Rex-dependency of this “two-phase” kinetics. These studies also highlighted interesting differences in the relative abundance of transcripts encoding the Tax and Rex regulatory proteins, and that of the accessory proteins controlling Rex expression and function, thus suggesting a potential basis for the different pathobiology of the two viruses. PMID:24032027

  13. Inhibition of simian immunodeficiency virus by foamy virus vectors expressing siRNAs

    SciTech Connect

    Park, Jeonghae; Nadeau, Peter; Zucali, James R.; Johnson, Calvin M.; Mergia, Ayalew . E-mail: mergiaa@mail.vetmed.ufl.edu

    2005-12-20

    Viral vectors available for gene therapy are either inefficient or suffer from safety concerns for human applications. Foamy viruses are non-pathogenic retroviruses that offer several unique opportunities for gene transfer in various cell types from different species. In this report, we describe the use of simian foamy virus type 1 (SFV-1) vector to examine the efficacy of therapeutic genes. Hairpin short-interfering RNA (siRNA) that targets the simian immunodeficiency virus (SIV) rev/env was placed under the control of the PolIII U6 snRNA promoter for expression and screened for silencing target genes using cognate target-reporter fusions. We have identified an effective siRNA (designated R2) which reduces the rev and env gene expression by 89% and 95%, respectively. Using the simian foamy virus type 1 (SFV-1) based vector, we delivered the PolIII expressed R2 siRNA into cultured cells and challenged with SIV. The results show that the R2 siRNA is a potent inhibitor of SIV replication as determined by p27 expression and reverse transcriptase assays. Vectors based on a non-pathogenic SFV-1 vector may provide a safe and efficient alternative to currently available vectors, and the SIV model will help devise protocols for effective anti-HIV gene therapy.

  14. Cytotoxic immune response blunts long-term transgene expression after efficient retroviral-mediated hepatic gene transfer in rat.

    PubMed

    Aubert, Dominique; Ménoret, Séverine; Chiari, Estelle; Pichard, Virginie; Durand, Sophie; Tesson, Laurent; Moullier, Philippe; Anegon, Ignacio; Ferry, Nicolas

    2002-04-01

    Vectors derived from oncoretroviruses can transduce a small proportion of hepatocytes when injected in the regenerating liver. Transgene expression may be sustained for months without immune response. In striking contrast, we observed a rapid extinction when the intravenous injection of a high input of nuclear beta-galactosidase (beta-gal) expression vector, one day after partial hepatectomy, led to a significant proportion of transduced cells in the liver. Extinction was associated with liver inflammation on tissue sections and appearance of antibodies against the transgene product, while vector genomes became undetectable in liver tissue by PCR. These observations suggested the elimination of transduced cells by an immune response. Transgenic rats tolerant for cytoplasmic beta-gal, or normal rats depleted in CD8 T lymphocytes, steadily expressed the beta-gal vector. In the spleen of normal rats, we detected cytotoxic cells directed against cells expressing beta-gal after the injection of the beta-gal vector. In jaundiced Gunn rats deficient in bilirubin glucuronosyl transferase (BGT1) and treated with a human BGT1 cDNA expression vector, we observed the same kinetics of extinction as well as the appearance of anti-BGT1 antibodies. This study demonstrates that retrovirus-mediated gene transfer may induce cytotoxic T lymphocytes specifically directed against transgene-expressing cells.

  15. Differential expression of HERV-K (HML-2) proviruses in cells and virions of the teratocarcinoma cell line Tera-1.

    PubMed

    Bhardwaj, Neeru; Montesion, Meagan; Roy, Farrah; Coffin, John M

    2015-03-04

    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.

  16. Targeted entry via somatostatin receptors using a novel modified retrovirus glycoprotein that delivers genes at levels comparable to those of wild-type viral glycoproteins.

    PubMed

    Li, Fang; Ryu, Byoung Y; Krueger, Robin L; Heldt, Scott A; Albritton, Lorraine M

    2012-01-01

    Here we report a novel viral glycoprotein created by replacing a natural receptor-binding sequence of the ecotropic Moloney murine leukemia virus envelope glycoprotein with the peptide ligand somatostatin. This new chimeric glycoprotein, which has been named the Sst receptor binding site (Sst-RBS), gives targeted transduction based on three criteria: (i) a gain of the use of a new entry receptor not used by any known virus; (ii) targeted entry at levels comparable to gene delivery by wild-type ecotropic Moloney murine leukemia virus and vesicular stomatitis virus (VSV) G glycoproteins; and (iii) a loss of the use of the natural ecotropic virus receptor. Retroviral vectors coated with Sst-RBS gained the ability to bind and transduce human 293 cells expressing somatostatin receptors. Their infection was specific to target somatostatin receptors, since a synthetic somatostatin peptide inhibited infection in a dose-dependent manner and the ability to transduce mouse cells bearing the natural ecotropic receptor was effectively lost. Importantly, vectors coated with the Sst-RBS glycoprotein gave targeted entry of up to 1 × 10(6) transducing U/ml, a level comparable to that seen with infection of vectors coated with the parental wild-type ecotropic Moloney murine leukemia virus glycoprotein through the ecotropic receptor and approaching that of infection of VSV G-coated vectors through the VSV receptor. To our knowledge, this is the first example of a glycoprotein that gives targeted entry of retroviral vectors at levels comparable to the natural capacity of viral envelope glycoproteins.

  17. Nemo-like kinase (NLK) expression in osteoblastic cells and suppression of osteoblastic differentiation

    SciTech Connect

    Nifuji, Akira; Ideno, Hisashi; Ohyama, Yoshio; Takanabe, Rieko; Araki, Ryoko; Abe, Masumi; Noda, Masaki; Shibuya, Hiroshi

    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 NLK 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.

  18. Nemo-like kinase (NLK) expression in osteoblastic cells and suppression of osteoblastic differentiation.

    PubMed

    Nifuji, Akira; Ideno, Hisashi; Ohyama, Yoshio; Takanabe, Rieko; Araki, Ryoko; Abe, Masumi; Noda, Masaki; Shibuya, Hiroshi

    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 NLK 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.

  19. Nonsense surveillance regulates expression of diverse classes of mammalian transcripts and mutes genomic noise.

    PubMed

    Mendell, Joshua T; Sharifi, Neda A; Meyers, Jennifer L; Martinez-Murillo, Francisco; Dietz, Harry C

    2004-10-01

    Premature termination codons induce rapid transcript degradation in eukaryotic cells through nonsense-mediated mRNA decay (NMD). This pathway can modulate phenotypes arising from nonsense or frameshift mutations, but little is known about the physiologic role of NMD in higher eukaryotes. To address this issue, we examined expression profiles in mammalian cells depleted of Rent1 (also called hUpf1), a factor essential for NMD. Upregulated transcripts included those with upstream open reading frames in the 5' untranslated region, alternative splicing that introduces nonsense codons or frameshifts, introns in the 3' untranslated region or selenocysteine codons. Transcripts derived from ancient transposons and endogenous retroviruses were also upregulated. These RNAs are unified by the presence of a spliced intron at least 50 nucleotides downstream of a termination codon, a context sufficient to initiate NMD. Consistent with direct regulation by NMD, representative upregulated transcripts decayed more slowly in cells deficient in NMD. In addition, inhibition of NMD induced by amino acid starvation upregulated transcripts that promote amino acid homeostasis. These results document that nonsense surveillance is a crucial post-transcriptional regulatory event that influences the expression of broad classes of physiologic transcripts, has been functionally incorporated into essential homeostatic mechanisms and suppresses expression of evolutionary remnants.

  20. Retroviral-mediated gene transfer and expression of human phenylalanine hydroxylase in primary mouse hepatocytes.

    PubMed Central

    Peng, H; Armentano, D; MacKenzie-Graham, L; Shen, R F; Darlington, G; Ledley, F D; Woo, S L

    1988-01-01

    Genetic therapy for phenylketonuria (severe phenylalanine hydroxylase deficiency) may require introduction of a normal phenylalanine hydroxylase gene into hepatic cells of patients. We report development of a recombinant retrovirus based on the N2 vector for gene transfer and expression of human phenylalanine hydroxylase cDNA in primary mouse hepatocytes. This construct contains an internal promoter of the human alpha 1-antitrypsin gene driving transcription of the phenylalanine hydroxylase cDNA. Primary mouse hepatocytes were isolated from newborn mice, infected with the recombinant virus, and selected for expression of the neomycin-resistance gene. Hepatocytes transformed with the recombinant virus contained high levels of human phenylalanine hydroxylase mRNA transcripts originating form the retroviral and internal promoters. These results demonstrate that the transcriptional regulatory elements of the alpha 1-antitrypsin gene retain their tissue-specific function in the recombinant provirus and establish a method for efficient transfer and high-level expression of human phenylalanine hydroxylase in primary hepatocytes. Images PMID:3186716

  1. 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.

  2. Altered Immune Cytokine Expression Associated with KoRV B Infection and Season in Captive Koalas

    PubMed Central

    Higgins, Damien P.

    2016-01-01

    Koala (Phascolarctos cinereus) populations are increasingly vulnerable and one of the main threats is chlamydial infection. Koala retrovirus (KoRV) has been proposed as an underlying cause of the koala’s susceptibility to infection with Chlamydia and high rates of lymphoid neoplasia; however, the regionally ubiquitous, endogenous nature of this virus suggests that KoRV A infection is not sufficient for immune suppression to occur. A recently discovered exogenous variant of KoRV, KoRV B, has several structural elements that cause increased pathogenicity in related retroviruses and was associated with lymphoid neoplasia in one study. The present study assesses whether KoRV B infection is associated with alterations in immune function. Cytokine gene expression by mitogen stimulated lymphocytes of KoRV B positive (n = 5–6) and negative (n = 6–7) captive koalas was evaluated by qPCR four times (April 2014-February 2015) to control for seasonal variation. Key immune genes in the Th1 pathway (IFNγ, TNFα), Th2 pathway (IL 10, IL4, IL6) and Th17 pathway (IL17A), along with CD4:CD8 ratio, were assessed. KoRV B positive koalas showed significantly increased up-regulation of IL17A and IL10 in three out of four sampling periods and IFNγ, IL6, IL4 and TNFα in two out of four. IL17A is an immune marker for chlamydial pathogenesis in the koala; increased expression of IL17A in KoRV B positive koalas, and concurrent immune dysregulation, may explain the differences in susceptibility to chlamydial infection and severity of disease seen between individuals and populations. There was also marked seasonal variation in up-regulation for most of the cytokines and the CD4:CD8 ratio. The up-regulation in both Th1 and Th2 cytokines mirrors changes associated with immune dysregulation in humans and felids as a result of retroviral infections. This is the first report of altered immune expression in koalas infected by an exogenous variant of KoRV and also the first report of

  3. Microarray-based Identification of Individual HERV Loci Expression: Application to Biomarker Discovery in Prostate Cancer

    PubMed Central

    Pérot, Philippe; Cheynet, Valérie; Decaussin-Petrucci, Myriam; Oriol, Guy; Mugnier, Nathalie; Rodriguez-Lafrasse, Claire; Ruffion, Alain; Mallet, François

    2013-01-01

    The prostate-specific antigen (PSA) is the main diagnostic biomarker for prostate cancer in clinical use, but it lacks specificity and sensitivity, particularly in low dosage values1​​. ‘How to use PSA' remains a current issue, either for diagnosis as a gray zone corresponding to a concentration in serum of 2.5-10 ng/ml which does not allow a clear differentiation to be made between cancer and noncancer2 or for patient follow-up as analysis of post-operative PSA kinetic parameters can pose considerable challenges for their practical application3,4. Alternatively, noncoding RNAs (ncRNAs) are emerging as key molecules in human cancer, with the potential to serve as novel markers of disease, e.g. PCA3 in prostate cancer5,6 and to reveal uncharacterized aspects of tumor biology. Moreover, data from the ENCODE project published in 2012 showed that different RNA types cover about 62% of the genome. It also appears that the amount of transcriptional regulatory motifs is at least 4.5x higher than the one corresponding to protein-coding exons. Thus, long terminal repeats (LTRs) of human endogenous retroviruses (HERVs) constitute a wide range of putative/candidate transcriptional regulatory sequences, as it is their primary function in infectious retroviruses. HERVs, which are spread throughout the human genome, originate from ancestral and independent infections within the germ line, followed by copy-paste propagation processes and leading to multicopy families occupying 8% of the human genome (note that exons span 2% of our genome). Some HERV loci still express proteins that have been associated with several pathologies including cancer7-10. We have designed a high-density microarray, in Affymetrix format, aiming to optimally characterize individual HERV loci expression, in order to better understand whether they can be active, if they drive ncRNA transcription or modulate coding gene expression. This tool has been applied in the prostate cancer field (Figure 1

  4. First update of the International Xenotransplantation Association consensus statement on conditions for undertaking clinical trials of porcine islet products in type 1 diabetes--Chapter 5: recipient monitoring and response plan for preventing disease transmission.

    PubMed

    Denner, Joachim; Tönjes, Ralf R; Takeuchi, Yasu; Fishman, Jay; Scobie, Linda

    2016-01-01

    Xenotransplantation of porcine cells, tissues, and organs may be associated with the transmission of porcine microorganisms to the human recipient. A previous, 2009, version of this consensus statement focused on strategies to prevent transmission of porcine endogenous retroviruses (PERVs). This version addresses potential transmission of all porcine microorganisms including monitoring of the recipient and provides suggested approaches to the monitoring and prevention of disease transmission. Prior analyses assumed that most microorganisms other than the endogenous retroviruses could be eliminated from donor animals under appropriate conditions which have been called "designated pathogen-free" (DPF) source animal production. PERVs integrated as proviruses in the genome of all pigs cannot be eliminated in that manner and represent a unique risk. Certain microorganisms are by nature difficult to eliminate even under DPF conditions; any such clinically relevant microorganisms should be included in pig screening programs. With the use of porcine islets in clinical trials, special consideration has to be given to the presence of microorganisms in the isolated islet tissue to be used and also to the potential use of encapsulation. It is proposed that microorganisms absent in the donor animals by sensitive microbiological examination do not need to be monitored in the transplant recipient; this will reduce costs and screening requirements. Valid detection assays for donor and manufacturing-derived microorganisms must be established. Special consideration is needed to preempt potential unknown pathogens which may pose a risk to the recipient. This statement summarizes the main achievements in the field since 2009 and focus on issues and solutions with microorganisms other than PERV.

  5. The putative zinc finger of a caulimovirus is essential for infectivity but does not influence gene expression.

    PubMed

    Scholthof, H B; Wu, F C; Kiernan, J M; Shepherd, R J

    1993-04-01

    Plant pararetroviruses, such as caulimoviruses, and animal retroviruses have in common the presence of a highly conserved arrangement of cysteines and a histidine in the precursor of the capsid protein. The composition of these amino acids resembles a zinc finger element, a structure that is common to a class of eukaryotic proteins that regulate gene expression. The role of the putative zinc finger in the life-cycle of caulimoviruses was investigated by introducing specific mutations in the coat protein coding region of a cloned and infectious form of figwort mosaic virus, a caulimovirus. This mutated viral genome, which no longer encoded the conserved cysteine and histidine residues, was not infectious in plants. Transient expression assays in protoplasts showed that expression of a reporter gene inserted at different places in the genome was not detectably influenced by the coat protein or its putative zinc finger. It appears that the zinc finger-like element of caulimoviruses is not involved in the regulation of gene expression. These observations support a model which predicts a function of the zinc finger in specific recognition and packaging of viral RNA into virions prior to reverse transcription.

  6. World Reference Center for Arboviruses and Retroviruses

    DTIC Science & Technology

    1989-05-01

    Diagnosis, ELISA, Complement-fixation 06 11 Dengue , Japanese encephalitis virus, Monoclonal Antibodies, 06 01 HIV, Yellow fever vaccine, Crimean-Congo...encephalitis in Georgia.......................... 7 Dengue -l virus identified from blood of a patient with serum yellow fever IgM reactivity in Angola...AnlO7237 VEE complex virus from sentinel mouse, Sao Paulo, Brazil ......................... 10 Identification of Kagoshima virus from Culicoides of

  7. Polarized expression of the membrane ASP protein derived from HIV-1 antisense transcription in T cells

    PubMed Central

    2011-01-01

    Background Retroviral gene expression generally depends on a full-length transcript that initiates in the 5' LTR, which is either left unspliced or alternatively spliced. We and others have demonstrated the existence of antisense transcription initiating in the 3' LTR in human lymphotropic retroviruses, including HTLV-1, HTLV-2, and HIV-1. Such transcripts have been postulated to encode antisense proteins important for the establishment of viral infections. The antisense strand of the HIV-1 proviral DNA contains an ORF termed asp, coding for a highly hydrophobic protein. However, although anti-ASP antibodies have been described to be present in HIV-1-infected patients, its in vivo expression requires further support. The objective of this present study was to clearly demonstrate that ASP is effectively expressed in infected T cells and to provide a better characterization of its subcellular localization. Results We first investigated the subcellular localization of ASP by transfecting Jurkat T cells with vectors expressing ASP tagged with the Flag epitope to its N-terminus. Using immunofluorescence microscopy, we found that ASP localized to the plasma membrane in transfected Jurkat T cells, but with different staining patterns. In addition to an entire distribution to the plasma membrane, ASP showed an asymmetric localization and could also be detected in membrane connections between two cells. We then infected Jurkat T cells with NL4.3 virus coding for ASP tagged with the Flag epitope at its C-terminal end. By this approach, we were capable of showing that ASP is effectively expressed from the HIV-1 3' LTR in infected T cells, with an asymmetric localization of the viral protein at the plasma membrane. Conclusion These results demonstrate for the first time that ASP can be detected when expressed from full-length HIV-1 proviral DNA and that its localization is consistent with Jurkat T cells overexpressing ASP. PMID:21929758

  8. HIV-1 Tat regulates the expression of the dcw operon and stimulates the proliferation of bacteria.

    PubMed

    Wei, Jinsong; Zhang, Yumin; Knapp, Pamela E; Zhao, Tianyong

    2016-01-01

    Infections of pathogenic bacteria are very common in acquired immunodeficiency syndrome (AIDS) patients. However, the biological effects of HIV-1 Tat on bacteria are incompletely understood. In this study, HIV-1 Tat was expressed in Escherichia coli and Pseudomonas aeruginosa (PA01) to investigate its biological effects on bacteria. Bacterial cells expressing either HIV-1 Tat1-86 (Tat1-86) or HIV-1 Tat1-72 (Tat1-72) grow significantly faster than those with either only an empty vector or an unrelated control (GFP or Rluc). Supplementation of purified HIV-1 Tat1-86 or Tat1-101 protein into bacterial culture medium stimulated the growth of both E. coli and PA01. The expression profile of certain cell division-associated genes, such as those in the division cell wall (dcw) operon (ftsA, ftsQ, ftsW and ftsZ), yafO and zipA, was altered in HIV-1 Tat1-86 expressing E. coli BL21(DE3). Furthermore, the expression of firefly luciferase (Fluc) reporter gene, when engineered for control by the dcw promoter and terminator, was enhanced by HIV-1 Tat in E. coli, confirming that HIV-1 Tat transcriptionally regulates the expression of the dcw operon. The finding that HIV-1 Tat stimulates bacterial growth whether it is produced intracellularly or applied extracellularly may have relevance for HIV patients who are highly susceptible to opportunistic bacterial infections. Contents category: Viruses -Retroviruses. The GenBank accession number for the sequence of HIV-1 Tat1-86 is AF324439.1.

  9. Thyroid and glucocorticoid hormones induce expression of lactase-phlorizin hydrolase gene in CDX-2/HNF-1α co-transfected IEC-6 cells.

    PubMed

    Suzuki, Takuji; Mochizuki, Kazuki; Goda, Toshinao

    2014-01-01

    Thyroid and glucocorticoid hormones and several transcriptional factors such as caudal type homeobox (CDX)-2 and hepatocyte nuclear factor (HNF)-1α are important for the differentiation of small intestinal absorptive cells and the consequent expression of genes related to the digestion/absorption of carbohydrates. In this study, we investigated whether thyroid and glucocorticoid hormones enhanced the expression of lactase-phlorizin hydrolase (LPH) gene, an intestine-specific gene that encodes an enzyme for lactose digestion, in small intestinal stem-like IEC-6 cells co-transfected with CDX-2 and HNF-1α using a retrovirus system. Changes in expression of intestine-specific genes caused by treatment with thyroid and/or glucocorticoid hormones were monitored in empty vector-transfected cells and in CDX-2/HNF-1α co-transfected cells by qRT-PCR. Stable co-transfection with CDX-2 and HNF-1α evoked the expression of the LPH gene in IEC-6 cells. Furthermore, treatment with a thyroid hormone, triiodothyronine, and a glucocorticoid receptor agonist, dexamethasone, significantly enhanced expression of the LPH, CDX-2 and HNF-1α genes in CDX-2/HNF-1α co-transfected IEC-6 cells. These results suggest that thyroid and glucocorticoid hormones synergistically enhance expression of the LPH gene in CDX-2/HNF-1α co-transfected IEC-6 cells.

  10. Immunodiagnosis of episomal Banana streak MY virus using polyclonal antibodies to an expressed putative coat protein.

    PubMed

    Sharma, Susheel Kumar; Kumar, P Vignesh; Baranwal, Virendra Kumar

    2014-10-01

    A cryptic Badnavirus species complex, known as banana streak viruses (BSV) poses a serious threat to banana production and genetic improvement worldwide. Due to the presence of integrated BSV sequences in the banana genome, routine detection is largely based on serological and nucleo-serological diagnostic methods which require high titre specific polyclonal antiserum. Viral structural proteins like coat protein (CP) are the best target for in vitro expression, to be used as antigen for antiserum production. However, in badnaviruses precise CP sequences are not known. In this study, two putative CP coding regions (p48 and p37) of Banana streak MY virus (BSMYV) were identified in silico by comparison with caulimoviruses, retroviruses and Rice tungro bacilliform virus. The putative CP coding region (p37) was in vitro expressed in pMAL system and affinity purified. The purified fusion protein was used as antigen for raising polyclonal antiserum in rabbit. The specificity of antiserum was confirmed in Western blots, immunosorbent electron microscopy (ISEM) and antigen coated plate-enzyme linked immunosorbent assay (ACP-ELISA). The antiserum (1:2000) was successfully used in ACP-ELISA for specific detection of BSMYV infection in field and tissue culture raised banana plants. The antiserum was also utilized in immuno-capture PCR (IC-PCR) based indexing of episomal BSMYV infection. This is the first report of in silico identification of putative CP region of BSMYV, production of polyclonal antiserum against recombinant p37 and its successful use in immunodetection.

  11. Loss of CD4 membrane expression and CD4 mRNA during acute human immunodeficiency virus replication

    PubMed Central

    1988-01-01

    Using mAbs and genomic probe to the CD4 molecule, the HIV receptor, we demonstrated that HIV replication induces the disappearance of its functional receptor from the cell surface by two distinct mechanisms. First, after being expressed onto the cell surface, HIV envelope gp110 will complex CD4, efficiently masking the CD4 epitope used by the virus to bind its receptor. This phenomenon occurs on the surface of each infected cell and is not due to the release of soluble gp110; infection with recombinant HIV/vaccinia viruses expressing a mutated HIV env gene designed to prevent gp110 release from the cell surface induces a similar gp/CD4 complexes formation. Second, virus replication induces a dramatic and rapid loss of CD4 mRNA transcripts, preventing new CD4 molecules from being synthesized. These two mechanisms of receptor modulation could have been developed to avoid reinfection of cells replicating the virus as well as to produce more infectious particles. These results suggest that the classical virus interference documented for other retroviruses might not only be due to receptor/envelope interaction, but might also depend on receptor gene expression. PMID:3264318

  12. 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.

  13. Receptor for Activated Protein Kinase C: Requirement for Efficient MicroRNA Function and Reduced Expression in Hepatocellular Carcinoma

    PubMed Central

    Otsuka, Motoyuki; Takata, Akemi; Yoshikawa, Takeshi; Kojima, Kentaro; Kishikawa, Takahiro; Shibata, Chikako; Takekawa, Mutsuhiro; Yoshida, Haruhiko; Omata, Masao; Koike, Kazuhiko

    2011-01-01

    MicroRNAs (miRNAs) are important regulators of gene expression that control physiological and pathological processes. A global reduction in miRNA abundance and function is a general trait of human cancers, playing a causal role in the transformed phenotype. Here, we sought to newly identify genes involved in the regulation of miRNA function by performing a genetic screen using reporter constructs that measure miRNA function and retrovirus-based random gene disruption. Of the six genes identified, RACK1, which encodes “receptor for activated protein kinase C” (RACK1), was confirmed to be necessary for full miRNA function. RACK1 binds to KH-type splicing regulatory protein (KSRP), a member of the Dicer complex, and is required for the recruitment of mature miRNAs to the RNA-induced silencing complex (RISC). In addition, RACK1 expression was frequently found to be reduced in hepatocellular carcinoma. These findings suggest the involvement of RACK1 in miRNA function and indicate that reduced miRNA function, due to decreased expression of RACK1, may have pathologically relevant roles in liver cancers. PMID:21935400

  14. Nickel-induced heritable alterations in retroviral transforming gene expression.

    PubMed Central

    Biggart, N W; Gallick, G E; Murphy, E C

    1987-01-01

    Determination of the mutagenic effects of carcinogenic nickel compounds has been difficult because, like many metals, nickel is poorly or nonmutagenic in procaryotic mutagenicity assays. We attempted to characterize nickel-induced genetic lesions by assessing the effect of nickel chloride on the conditionally defective expression of the v-mos transforming gene in normal rat kidney cells infected with the Murine sarcoma virus mutant ts110 (MuSVts110) retrovirus. MuSVts110 contains an out-of-frame gag gene-mos gene junction that prevents the expression of the v-mos gene at the nonpermissive temperature (39 degrees C). In MuSVts110-infected cells (6m2 cells) grown at 33 degrees C, however, this defect can be suppressed by a splicing event that restores the mos reading frame, allowing the expression of a gag-mos fusion protein which induces the transformed phenotype. The capacity to splice the viral transcript at 33 degrees C, but not at 39 degrees C, is an intrinsic property of the viral RNA. This property allowed us to target the MuSVts110 genome using a positive selection scheme whereby nickel was used to induce genetic changes which resulted in expression of the transformed phenotype at 39 degrees C. We treated 6m2 cells with NiCl2 and isolated foci consisting of cells which had reverted to the transformed phenotype at 39 degrees C. We found that brief nickel treatment increased the reversion frequency of 6m2 cells grown at 39 degrees C sevenfold over the spontaneous reversion frequency. The nickel-induced revertants displayed the following heritable characteristics: They stably maintained the transformed phenotype at 39 degrees C; unlike the MuSVts110 RNA in 6m2 cells, the nickel-induced revertant viral RNA could be spliced efficiently at 39 degrees C; as a consequence of the enhanced accumulation of spliced viral RNA, the nickel-induced revertants produced substantial amounts of the transforming v-mos protein P85gag-mos at 39 degrees C; the nickel

  15. Expression of immunoglobulin genes in the avian embryo bone marrow revealed by retroviral transformation.

    PubMed

    Benatar, T; Iacampo, S; Tkalec, L; Ratcliffe, M J

    1991-10-01

    Analysis of the early stages of avian B lymphocyte differentiation has been hampered by the low frequency of extra-bursal B lineage cells in sites of hematopoiesis. Consequently, little is known about B lineage precursors prior to their migration into the bursa of Fabricius. Colonization of the bursa typically occurs between about days 8 and 14 of embryonic (e) development, although cells which can colonize the bursa, functionally defined as pre-bursal stem cells, can be demonstrated in embryo bone marrow up until about the time of hatch. As a novel approach to analyzing early stages of avian B lymphocyte development, we show here that transformed B lineage cells can be derived from chick embryo bone marrow after infection in vitro with the replication-defective retrovirus REV-T produced in the context of the non-cytopathic CSV helper virus. Thus, exposure of day 14e-15e chick embryo bone marrow cells to REV-T (CSV) results in the generation of transformed, polyclonal lines of cells. From these lines, cells expressing cell surface immunoglobulin were readily isolated by flow cytometric cell sorting and single cell cloning. Analysis of the phenotype of REV-T(CSV)-transformed clones with a panel of monoclonal antibody reagents demonstrated that transformation by v-rel likely leads to marked changes in cell surface antigen expression. Nonetheless, clones expressing cell surface immunoglobulin expressed apparently normal mRNA for immunoglobulin mu and light chain and contained apparently normal immunoglobulin heavy and light chain gene rearrangements. Furthermore, no evidence for chromosomal deletions or aberrations of the Ig loci was detected among either sIg+ or sIg- REV-T(CSV)-transformed clones.

  16. Human T-cell leukemia virus type 1 (HTLV-1) Tax1 oncoprotein but not HTLV-2 Tax2 induces the expression of OX40 ligand by interacting with p52/p100 and RelB.

    PubMed

    Motai, Yosuke; Takahashi, Masahiko; Takachi, Takayuki; Higuchi, Masaya; Hara, Toshifumi; Mizuguchi, Mariko; Aoyagi, Yutaka; Terai, Shuji; Tanaka, Yuetsu; Fujii, Masahiro

    2016-02-01

    Human T-cell leukemia virus type 1 (HTLV-1) is a causative retrovirus of adult T-cell leukemia and HTLV-1-associated myelopathy. Unlike HTLV-1, the same group of retrovirus HTLV-2 has not been found to be associated with these diseases. HTLV-1 and HTLV-2 encode transforming proteins Tax1 and Tax2, and a few distinct activities of Tax1 from those of Tax2 have been proposed to contribute to the HTLV-1-specific pathogenesis of disease. One significant difference of Tax1 from Tax2 is the activation of transcription factor NF-κB2/p100/p52. We found that Tax1 but not Tax2 induces the expression of OX40 ligand (OX40L) in a human T-cell line. To induce the OX40L expression, Tax1 but not Tax2 was observed to interact with NF-κB2/p100/p52 and RelB and the distinct interaction activity was mediated by the Tax1 amino acid region of 225-232. In addition, Tax1 but not Tax2 or Tax1/225-232 interacted with p65, p50, and c-Rel; however, the interactions were much less than those noted with NF-κB2/p100/p52 and RelB. OX40L is a T-cell costimulatory molecule of the tumor necrosis factor family, and its signal plays a critical role in establishing adaptive immunity by inducing the polarized differentiation of T-cells to cells such as T helper type 2 and T follicular helper cells. Therefore, the present findings suggest that Tax1 might alter the immune response to HTLV-1 and/or differentiation of HTLV-1-infected T-cells via OX40L induction, thereby acting as a factor mediating the distinct phenotypes and pathogenesis of HTLV-1 from that of HTLV-2.

  17. 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.

  18. Replacement of the human cytomegalovirus promoter with fish enhancer and core elements to control the expression of the G gene of viral haemorrhagic septicemia virus (VHSV).

    PubMed

    Martinez-Lopez, A; Chinchilla, B; Encinas, P; Gomez-Casado, E; Estepa, A; Coll, J M

    2012-12-15

    This work explores some of the possibilities to replace human cytomegalovirus (CMV) core and/or enhancer promoter control elements to create new expression vectors for use with fish. The work is relevant to fish vaccination, since DNA vaccines use eukaryotic expression plasmids controlled by the human cytomegalovirus (CMV) promoter to be effective against novirhabdoviruses, such as viral haemorrhagic septicemia virus (VHSV), one of the most devastating fish viral European diseases. To reduce possible homologous recombination with fish genome, core and enhancer sequences from fish origin, such as trout interferon-inducible myxovirus protein (Mx), zebrafish retrovirus long terminal repeat (LTR) and carp β-actin (AE6), were combined with those of CMV to design alternative hybrid promoters. The substitution of CMV core and/or enhancer with the corresponding elements of Mx or the LTR core maintained a similar in vitro protein G expression level than that obtained by using the CMV promoter. Vectors using the dsRNA-inducible Mx enhancer followed either by the LTR or the AE6 cores showed the highest in vitro protein G expression levels. Furthermore, synthetic constructs using the Mx enhancer maintained their polyI:C induction capabilities despite the core used. Some of these hybrid promoters might contribute to the development of all-fish-vectors for DNA vaccines while others might be useful for more basic studies.

  19. PRMT5 Is Upregulated in HTLV-1-Mediated T-Cell Transformation and Selective Inhibition Alters Viral Gene Expression and Infected Cell Survival.

    PubMed

    Panfil, Amanda R; Al-Saleem, Jacob; Howard, Cory M; Mates, Jessica M; Kwiek, Jesse J; Baiocchi, Robert A; Green, Patrick L

    2015-12-30

    Human T-cell leukemia virus type-1 (HTLV-1) is a tumorigenic retrovirus responsible for development of adult T-cell leukemia/lymphoma (ATLL). This disease manifests after a long clinical latency period of up to 2-3 decades. Two viral gene products, Tax and HBZ, have transforming properties and play a role in the pathogenic process. Genetic and epigenetic cellular changes also occur in HTLV-1-infected cells, which contribute to transformation and disease development. However, the role of cellular factors in transformation is not completely understood. Herein, we examined the role of protein arginine methyltransferase 5 (PRMT5) on HTLV-1-mediated cellular transformation and viral gene expression. We found PRMT5 expression was upregulated during HTLV-1-mediated T-cell transformation, as well as in established lymphocytic leukemia/lymphoma cell lines and ATLL patient PBMCs. shRNA-mediated reduction in PRMT5 protein levels or its inhibition by a small molecule inhibitor (PRMT5i) in HTLV-1-infected lymphocytes resulted in increased viral gene expression and decreased cellular proliferation. PRMT5i also had selective toxicity in HTLV-1-transformed T-cells. Finally, we demonstrated that PRMT5 and the HTLV-1 p30 protein had an additive inhibitory effect on HTLV-1 gene expression. Our study provides evidence for PRMT5 as a host cell factor important in HTLV-1-mediated T-cell transformation, and a potential target for ATLL treatment.

  20. Nef from primary isolates of human immunodeficiency virus type 1 suppresses surface CD4 expression in human and mouse T cells.

    PubMed Central

    Anderson, S; Shugars, D C; Swanstrom, R; Garcia, J V

    1993-01-01

    The human immunodeficiency virus type 1 (HIV-1) nef gene was originally described as a negative regulator of transcription from the viral long terminal repeat promoter. This observation has been disputed, and the function of Nef remains unclear. In vivo experiments have indicated that an intact nef gene is required for disease progression in macaques infected with simian immunodeficiency virus, suggesting a role for Nef in the pathogenesis of AIDS. We and others have previously shown that expression of Nef in cells bearing surface CD4 results in a sustained decrease in surface CD4 expression. This was demonstrated for Nef from two laboratory strains of HIV-1, Bru and SF2. Because both of these isolates were passaged in vitro prior to molecular cloning and in vitro passage can result in mutations which might alter nef gene function, we have analyzed two primary isolates of Nef for their ability to suppress cell surface CD4 expression. The nef genes of HIV-1 isolates from two patients with fewer than 200 CD4+ T cells per mm3 of blood were introduced into human and mouse T-cell lines by retrovirus-mediated gene transfer. Expression of Nef from both isolates correlated with a decrease in surface expression of both human and mouse CD4. To determine whether the ability to suppress surface CD4 expression is a general function of Nef, we also tested an artificially generated consensus nef gene derived from analysis of 54 patient isolates of HIV-1. Expression of the consensus Nef protein also correlated with decreased cell surface CD4 expression in both mouse and human T-cell lines. These results suggest that the ability to suppress cell surface CD4 expression is an intrinsic feature of HIV-1 Nef. Images PMID:8331733

  1. 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.

  2. The HTLV-1 HBZ protein inhibits cyclin D1 expression through interacting with the cellular transcription factor CREB.

    PubMed

    Ma, Yunyun; Zheng, Shangen; Wang, Yuanyuan; Zang, Wenqiao; Li, Min; Wang, Na; Li, Ping; Jin, Jing; Dong, Ziming; Zhao, Guoqiang

    2013-10-01

    Human T cell leukemia virus type 1 (HTLV-1) is an oncogenic retrovirus that can cause adult T-cell leukemia (ATL) and other diseases. The HTLV-1 bZIP factor (HBZ), which is encoded by an mRNA of the opposite polarity of the viral genomic RNA, interacts with several transcription factors and is involved in T cell proliferation, viral gene transcription and cellular transformation. Cyclin D1 is a pivotal regulatory protein involved in cell cycle progression, and its depressed expression correlates with cell cycle prolongation or arrested at the G1/S transition. In our present study, we observed that HBZ expression suppressed cyclin D1 level. To investigate the role of HBZ on cyclin D1 depression, we transduced HBZ with lentivirus vector into 293T cells, CEM cells and Jurkat cells. The results of Western blot, RT-PCR and luciferase assays showed that transcriptional activity of the cyclin D1 promoter was suppressed by the bZIP domain of HBZ (HBZ-bZIP) through cyclic AMP response element (CRE) site. Immunoprecipitation and GST pull-down assays showed the binding of HBZ-bZIP to CRE-binding protein (CREB), which confirmed that the cyclin D1 promoter activity inhibition via the CRE-site was mediated by HBZ-bZIP. The results suggested that HBZ suppressed cyclin D1 transcription through interactions with CREB and along with other viral protein, HBZ may play a causal role for leukemogenesis.

  3. Chip, a widely expressed chromosomal protein required for segmentation and activity of a remote wing margin enhancer in Drosophila

    PubMed Central

    Morcillo, Patrick; Rosen, Christina; Baylies, Mary K.; Dorsett, Dale

    1997-01-01

    The mechanisms allowing remote enhancers to regulate promoters several kilobase pairs away are unknown but are blocked by the Drosophila suppressor of Hairy-wing protein (Suhw) that binds to gypsy retrovirus insertions between enhancers and promoters. Suhw bound to a gypsy insertion in the cut gene also appears to act interchromosomally to antagonize enhancer–promoter interactions on the homologous chromosome when activity of the Chip gene is reduced. This implicates Chip in enhancer–promoter communication. We cloned Chip and find that it encodes a homolog of the recently discovered mouse Nli/Ldb1/Clim-2 and Xenopus Xldb1 proteins that bind nuclear LIM domain proteins. Chip protein interacts with the LIM domains in the Apterous homeodomain protein, and Chip interacts genetically with apterous, showing that these interactions are important for Apterous function in vivo. Importantly, Chip also appears to have broad functions beyond interactions with LIM domain proteins. Chip is present in all nuclei examined and at numerous sites along the salivary gland polytene chromosomes. Embryos without Chip activity lack segments and show abnormal gap and pair–rule gene expression, although no LIM domain proteins are known to regulate segmentation. We conclude that Chip is a ubiquitous chromosomal factor required for normal expression of diverse genes at many stages of development. We suggest that Chip cooperates with different LIM domain proteins and other factors to structurally support remote enhancer–promoter interactions. PMID:9334334

  4. Subgroup J avian leukosis virus infection of chicken dendritic cells induces apoptosis via the aberrant expression of microRNAs.

    PubMed

    Liu, Di; Dai, Manman; Zhang, Xu; Cao, Weisheng; Liao, Ming

    2016-02-01

    Subgroup J avian leukosis virus (ALV-J) is an oncogenic retrovirus that causes immunosuppression and enhances susceptibility to secondary infection. The innate immune system is the first line of defense in preventing bacterial and viral infections, and dendritic cells (DCs) play important roles in innate immunity. Because bone marrow is an organ that is susceptible to ALV-J, the virus may influence the generation of bone marrow-derived DCs. In this study, DCs cultured in vitro were used to investigate the effects of ALV infection. The results revealed that ALV-J could infect these cells during the early stages of differentiation, and infection of DCs with ALV-J resulted in apoptosis. miRNA sequencing data of uninfected and infected DCs revealed 122 differentially expressed miRNAs, with 115 demonstrating upregulation after ALV-J infection and the other 7 showing significant downregulation. The miRNAs that exhibited the highest levels of upregulation may suppress nutrient processing and metabolic function. These results indicated that ALV-J infection of chicken DCs could induce apoptosis via aberrant microRNA expression. These results provide a solid foundation for the further study of epigenetic influences on ALV-J-induced immunosuppression.

  5. Cloning of the rat ecotropic retroviral receptor and studies of its expression in intestinal tissues

    SciTech Connect

    Puppi, M.; Henning, S.J.

    1995-05-01

    A long-term goal of our laboratory is to establish a rat model to study the feasibility of using the intestinal tract as a site for somatic gene therapy. As a step toward that goal, the current study reports the cloning of the rat ecotropic retroviral receptor (EcoR) cDNA and the study of various aspects of its expression in the intestinal cDNA library with mouse EcoR cDNA. A clone of approximately 7 kb, designated MP10, was obtained. Partial sequencing of MP10 from the 5{prime} end revealed a level of similarity of 92% compared with mouse EcoR. The presence of a 5{prime} untranslated region and a 3{prime} poly(A)tract, together with the overall size of the cDNA, suggest that is very close to being a full-length cDNA for this large transcript. Northern blots with MP10 showed an RNA of approximately 7.9 kb present along the entire length of the small intestine and somewhat less abundant in the colon. Developmental studies showed high levels of EcoR in fetal rat intestine, a decline in the early postnatal period, then a gradual rise to adulthood. Caco-2 cells were used to assess the expression of EcoR in proliferating compared with differentiated intestinal epithelial cells. EcoR mRNA was found to be very much more abundant in nondifferentiated cells and declined to low levels as the cells underwent spontaneous differentiation. These patterns of EcoR expression indicate that ecotropic retroviruses should be suitable vectors with which to attempt gene transfer into the intestinal epithelium. In addition, since the endogenous role of EcoR is as the y{sup +} cationic amino acid transporter, these data have significance for understanding patterns of amino acid transport in the intestinal epithelium. 37 refs., 4 figs.

  6. Molecular Evolution of the Porcine Type I Interferon Family: Subtype-Specific Expression and Antiviral Activity

    PubMed Central

    Sang, Yongming; Bergkamp, Joseph; Blecha, Frank

    2014-01-01

    Type I interferons (IFNs), key antiviral cytokines, evolve to adapt with ever-changing viral threats during vertebrate speciation. Due to novel pathogenic pressure associated with Suidae speciation and domestication, porcine IFNs evolutionarily engender both molecular and functional diversification, which have not been well addressed in pigs, an important livestock species and animal model for biomedical sciences. Annotation of current swine genome assembly Sscrofa10.2 reveals 57 functional genes and 16 pseudogenes of type I IFNs. Subfamilies of multiple IFNA, IFNW and porcine-specific IFND genes are separated into four clusters with ∼60 kb intervals within the IFNB/IFNE bordered region in SSC1, and each cluster contains mingled subtypes of IFNA, IFNW and IFND. Further curation of the 57 functional IFN genes indicates that they include 18 potential artifactual duplicates. We performed phylogenetic construction as well as analyses of gene duplication/conversion and natural selection and showed that porcine type I IFN genes have been undergoing active diversification through both gene duplication and conversion. Extensive analyses of the non-coding sequences proximal to all IFN coding regions identified several genomic repetitive elements significantly associated with different IFN subtypes. Family-wide studies further revealed their molecular diversity with respect to differential expression and restrictive activity on the resurgence of a porcine endogenous retrovirus. Based on predicted 3-D structures of representative animal IFNs and inferred activity, we categorized the general functional propensity underlying the structure-activity relationship. Evidence indicates gene expansion of porcine type I IFNs. Genomic repetitive elements that associated with IFN subtypes may serve as molecular signatures of respective IFN subtypes and genomic mechanisms to mediate IFN gene evolution and expression. In summary, the porcine type I IFN profile has been phylogenetically

  7. A simple and efficient procedure for generating stable expression libraries by cDNA cloning in a retroviral vector.

    PubMed Central

    Rayner, J R; Gonda, T J

    1994-01-01

    cDNA expression cloning is a powerful method for the rescue and identification of genes that are able to confer a readily identifiable phenotype on specific cell types. Retroviral vectors provide several advantages over DNA-mediated gene transfer for the introduction of expression libraries into eukaryotic cells since they can be used to express genes in a wide range of cell types, including those that form important experimental systems such as the hemopoietic system. We describe here a straightforward and efficient method for generating expression libraries by using a murine retroviral vector. Essentially, the method involves the directional cloning of cDNA into the retroviral vector and the generation of pools of stable ecotropic virus producing cells from this DNA. The cells so derived constitute the library, and the virus they yield is used to infect appropriate target cells for subsequent functional screening. We have demonstrated the feasibility of this procedure by constructing several large retroviral libraries (10(5) to 10(6) individual clones) and then using one of these libraries to isolate cDNAs for interleukin-3 and granulocyte-macrophage colony-stimulating factor on the basis of the ability of these factors to confer autonomous growth on the factor-dependent hemopoietic cell line FDC-P1. Moreover, the frequency at which these factor-independent clones were isolated approximated the frequency at which they were represented in the original plasmid library. These results suggest that expression cloning with retroviruses is a practical and efficient procedure and should be a valuable method for the isolation of important regulatory genes. Images PMID:8289827

  8. Regulation of expression driven by human immunodeficiency virus type 1 and human T-cell leukemia virus type I long terminal repeats in pluripotential human embryonic cells

    SciTech Connect

    Maio, J.; Brown, F.L. )

    1988-04-01

    Human pluripotential embryonic teratocarcinoma cells differentially expressed gene activity controlled by the human immunodeficiency virus type 1 (HIV-1) and human T-cell leukemia virus type I (HTLV-I) long terminal repeats (LTRs) when differentiation was induced by the morphogen all-trans retinoic acid. The alterations occurred after commitment and before the appearance of the multiple cell types characteristic of these pluripotential cells. After commitment, gene activity controlled by the HIV-1 LTR markedly increased, whereas that controlled by the HTLV-I LTR decreased. Steady-state mRNA levels and nuclear run-on transcription indicated that the increased HIV-1-directed activity during differentiation occurred posttranscriptionally, whereas the decreased HTLV-I activity was at the transcriptional level. Phorbol esters did not cause commitment but strongly enhanced expression by both viral LTRs at the transcriptional level. Differentiating cells gradually lost the ability to respond to phorbol ester stimulation. Experiments with a deletion mutant of the HIV-1 LTR suggested that this was due to imposition of negative regulation during differentiation that was not reversed by phorbol ester induction. Cycloheximide, with or without phorbol ester, slightly stimulated HIV-1-directed activity at the transcriptional level and massively increased the amounts of steady-state mRNA by posttranscriptional superinduction. It appeared, however, that new nuclear protein synthesis was required for maximal transcriptional stimulation by phorbol esters. Thus, changing cellular regulatory mechanisms influenced human retrovirus expression during human embryonic cell differentiation.

  9. Expression of the human beta-globin gene after retroviral transfer into murine erythroleukemia cells and human BFU-E cells.

    PubMed Central

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

    1988-01-01

    Replication-defective amphotropic retrovirus vectors containing either the human beta-globin gene with introns or an intronless beta-globin minigene were constructed and used to study beta-globin expression following gene transfer into hematopoietic cells. The beta-globin genes were marked by introducing a 6-base-pair insertion into the region corresponding to the 5' untranslated region of the beta-globin mRNA to allow detection of RNA encoded by the new gene in human cells expressing normal human beta-globin RNA. Introduction of a virus containing the beta-globin gene with introns into murine erythroleukemia cells resulted in inducible expression of human beta-globin RNA and protein, while the viruses containing the minigene were inactive. The introduced human beta-globin gene was 6 to 110% as active as the endogenous mouse beta maj-globin genes in six randomly chosen cell clones. Introduction of the viruses into human BFU-E cells, followed by analysis of marked and unmarked globin RNAs in differentiated erythroid colonies, revealed that the introduced beta-globin gene was about 5% as active as the endogenous genes in these normal human erythroid cells and that again the minigene was inactive. These data are discussed in terms of the potential treatment of genetic disease by gene therapy. Images PMID:3288863

  10. Expression of Endogenous Retroviral Genes in Leukemic Guinea Pig Cells

    PubMed Central

    Davis, A. R.; Nayak, D. P.

    1977-01-01

    The expression of guinea pig retrovirus (5-bromodeoxyuridine[BUdR]-induced GPV) was studied in guinea pig L2C leukemic lymphoblasts by use of molecular hybridization of viral complementary DNA (cDNA) to cellular RNA. It was found that L2C leukemic lymphoblasts, leukemic spleen, and BUdR-induced virus-producing cells contain virus-specific RNA: 0.05% (800 to 960 copies per cell), 0.02% (360 copies per cell), and 0.3% (5,120 copies per cell), respectively. Adult normal liver and spleen, on the other hand, contain less than 0.2 copy of viral RNA per cell. Both BUdR-induced cells and L2C leukemic lymphoblasts contained 14S, 22S, 35S, and 70S RNA species of total and cytoplasmic virus-specific RNA as determined by sucrose velocity gradient analysis and hybridization of sucrose gradient fractions to cDNA. Virus-specific mRNA was identified in both BUdR-induced cells and L2C leukemic lymphoblasts by the criterion that it cosedimented with purified polyribosomes in a sucrose gradient and that it changed to a lower sedimentation value if polyribosomes were disaggregated with EDTA prior to centrifugation. Virus-specific mRNA obtained from either the polyribosome region of purified polyribosomes or the released messenger region of EDTA-disaggregated purified polyribosomes consisted of 14S, 20S, and 35S species in both BUdR-induced cells and L2C leukemic lymphoblasts. Hybridization of cDNA to the RNA of L2C leukemic lymphoblasts and BUdR-induced cells was essentially complete. Additionally, leukemic lymphoblast RNA could displace 95% of the hybridization of BUdR-induced GPV 70S RNA to guinea pig DNA. The midpoints of thermal denaturation of hybrids formed between GPV cDNA and the RNA of either L2C leukemic lymphoblasts or the 70S RNA of BUdR-induced GPV were both 89°C in 2× concentrated 0.15 M NaCl plus 0.015 M sodium citrate. These results show that BUdR-induced GPV genes are essentially completely expressed in L2C leukemic lymphoblasts and that virus-specific mRNA is

  11. Flow cytometry analysis of adenosine deaminase (ADA) expression: a simple and reliable tool for the assessment of ADA-deficient patients before and after gene therapy.

    PubMed

    Otsu, Makoto; Hershfield, Michael S; Tuschong, Laura M; Muul, Linda M; Onodera, Masafumi; Ariga, Tadashi; Sakiyama, Yukio; Candotti, Fabio

    2002-02-10

    Clinical gene therapy trials for adenosine deaminase (ADA) deficiency have shown limited success of corrective gene transfer into autologous T lymphocytes and CD34(+) cells. In these trials, the levels of gene transduction and expression in hematopoietic cells have been assessed by DNA- or RNA-based assays and measurement of ADA enzyme activity. Although informative, these methods are rarely applied to clonal analysis. The results of these assays therefore provide best estimates of transduction efficiency and gene expression in bulk populations based on the assumption that gene transfer and expression are uniformly distributed among transduced cells. As a useful additional tool for evaluation of ADA gene expression, we have developed a flow cytometry (fluorescence-activated cell sorting, FACS) assay capable of estimating the levels of intracellular ADA on a single-cell basis. We validated this technique with T cell lines and peripheral blood mononuclear cells (PBMCs) from ADA-deficient patients that showed severely reduced levels of ADA expression (ADA-dull) by FACS and Western blot analyses. After retrovirus-mediated ADA gene transfer, these cells showed clearly distinguishable populations exhibiting ADA expression (ADA-bright), thus allowing estimation of transduction efficiency. By mixing ADA-deficient and normal cells and using enzymatic amplification, we determined that our staining procedure could detect as little as 5% ADA-bright cells. This technique, therefore, will be useful to quickly assess the expression of ADA in hematopoietic cells of severe combined immunodeficient patients and represents an important tool for the follow-up of patients treated in clinical gene transfer protocols.

  12. [Construction of mouse VCAM-1 expression vector and establishment of stably transfected MSC line C3H10T1/2].

    PubMed

    Chen, Hui; Zhu, Heng; Chu, Ya-Nan; Xu, Fen-Fen; Liu, Yuan-Lin; Tang, Bo; Li, Xi-Mei; Hu, Liang-Ding; Zhang, Yi

    2014-10-01

    This study was aimed to construct the mouse VCAM-1 expression vector, to establish the stably transfected MSC line and to investigate the effect of VCAM-1-modified mesenchymal stem cells (MSC) on the immunological characteristics of MSC. The cDNA of murine VCAM-1 gene was amplified by RT-PCR from the total RNA isolated from the mouse spleen; then the cDNA was inserted into the retrovirus vector PMSCVmigr-1; the recombinant plasmid was confirmed by restriction endonuclease experiments and sequencing, then designated as PMSCVmigr-1-mVCAM-1; the recombinant plasmid PMSCVmigr-1-mVCAM-1 was transfected into 293 cells by lipofecamin and the supernatant was collected to transfect MSC cell line (C3H10T1/2). Moreover, VCAM-1 expression on MSC was evaluated by FACS. Furthermore, the inhibitory effect of VCAM-1-MSC on lymphocytic transformation was tested by (3)H-TdR incorporation assay. The results indicated that the successful construction of recombinant retroviral expression plasmid of mouse VCAM-1 was confirmed by digesting and sequancing. After transfection of MSC with retroviral supernaptant, the high expression of VCAM-1 on MSC could be detected by flow cytometry. The MSC high expressing VCAM-1 could significantly inhibit the proliferation of Con A-inducing lymphocytes in dose-depentent marrer. It is concluded that recombinant retroviral encoding VCAM-1 (PMSCVmigr-1-mVCAM-1) has been successfully constructed and mouse VCAM-1 has been stably expressed in C3H10T1/2. MSC over-expressing VCAM-1 show more potent immunosuppressive effect on cellular immune reaction in vitro. Our data laid a foundation for the subsequent studying the effect of VCAM-1 transfecting into MSC on immune related disease study.

  13. A Versatile Viral System for Expression and Depletion of Proteins in Mammalian Cells

    PubMed Central

    Campeau, Eric; Ruhl, Victoria E.; Rodier, Francis; Smith, Corey L.; Rahmberg, Brittany L.; Fuss, Jill O.; Campisi, Judith; Yaswen, Paul; Cooper, Priscilla K.; Kaufman, Paul D.

    2009-01-01

    The ability to express or deplete proteins in living cells is crucial for the study of biological processes. Viral vectors are often useful to deliver DNA constructs to cells that are difficult to transfect by other methods. Lentiviruses have the additional advantage of being able to integrate into the genomes of non-dividing mammalian cells. However, existing viral expression systems generally require different vector backbones for expression of cDNA, small hairpin RNA (shRNA) or microRNA (miRNA) and provide limited drug selection markers. Furthermore, viral backbones are often recombinogenic in bacteria, complicating the generation and maintenance of desired clones. Here, we describe a collection of 59 vectors that comprise an integrated system for constitutive or inducible expression of cDNAs, shRNAs or miRNAs, and use a wide variety of drug selection markers. These vectors are based on the Gateway technology (Invitrogen) whereby the cDNA, shRNA or miRNA of interest is cloned into an Entry vector and then recombined into a Destination vector that carries the chosen viral backbone and drug selection marker. This recombination reaction generates the desired product with >95% efficiency and greatly reduces the frequency of unwanted recombination in bacteria. We generated Destination vectors for the production of both retroviruses and lentiviruses. Further, we characterized each vector for its viral titer production as well as its efficiency in expressing or depleting proteins of interest. We also generated multiple types of vectors for the production of fusion proteins and confirmed expression of each. We demonstrated the utility of these vectors in a variety of functional studies. First, we show that the FKBP12 Destabilization Domain system can be used to either express or deplete the protein of interest in mitotically-arrested cells. Also, we generate primary fibroblasts that can be induced to senesce in the presence or absence of DNA damage. Finally, we

  14. Genetic modification of a chicken expression system for the galactosylation of therapeutic proteins produced in egg white.

    PubMed

    Mizutani, Akifumi; Tsunashima, Hiroyuki; Nishijima, Ken-ichi; Sasamoto, Takako; Yamada, Yuki; Kojima, Yasuhiro; Motono, Makoto; Kojima, Jun; Inayoshi, Yujin; Miyake, Katsuhide; Park, Enoch Y; Iijima, Shinji

    2012-02-01

    As a tool for large scale production of recombinant proteins, chickens have advantages such as high productivity and low breeding costs compared to other animals. We previously reported the production of erythropoietin, the tumor necrosis factor receptor fused to an Fc fragment, and an Fc-fused single-chain Fv antibody in eggs laid by genetically manipulated chickens. In egg white, however, the incomplete addition of terminal sugars such as sialic acid and galactose was found on N-linked glycans of exogenously expressed proteins. This could be a draw back to the use of transgenic chickens since the loss of these terminal sugars may affect the functions and stability of recombinant proteins purified from chicken egg white for pharmaceutical usage. To overcome this problem, we studied galactosyltransferase (GalT) activity in the magnum where the majority of egg-white proteins are secreted. In the magnum, lower β1,4-GalT1 expression and poor galactose-transfer activity were observed. Thus, we supposed that the lack of GalT1 activity may partly cause the incomplete glycosylation of egg-white proteins, and generated genetically manipulated chickens expressing GalT1 by retrovirus-mediated gene transfer. In a Golgi fraction prepared from magnum cells of the genetically manipulated chickens, significant GalT activity was detected. The series of analyses revealed a considerable improvement in the galactosylation of native egg-white proteins as well as an exogenously expressed single-chain Fv antibody fused to an Fc fragment. We conclude that chickens with genetically modified GalT activity in the magnum could be an attractive platform for producing galactosylated therapeutics.

  15. Establishment of mouse leukemia cell lines expressing human CD4/CCR5 using lentiviral vectors.

    PubMed

    Li, Ya-Jing; ZhuGe, Fu-Yan; Zeng, Chang-Chun; He, Jin-Yang; Tan, Ning; Liang, Juan

    2017-04-01

    A low-cost rodent model of HIV infection and which presents high application value is an effective tool to investigate HIV infection and pathogenesis. However, development of such a small animal model has been hampered by the unsuitability of rodent cells for HIV-1 replication given that the retrovirus HIV-1 has high selectivity to its host cell. Our study used the mouse leukemia cell lines L615 and L1210 that were induced by murine leukemia virus and transfected with hCD4/CCR5 loaded-lentiviral vector. Lentiviral vectors containing the genes hCD4/CCR5 under the transcriptional control of cytomegalovirus promoter were designed. Transfection efficiencies of human CD4 and CCR5 in L615 and L1210 cells were analyzed by quantitative real-time polymerase chain reaction (RT-PCR) and Western blot assay. Results showed that hCD4 and CCR5 proteins were expressed on the cell surface, demonstrating that the L615 and L1210 cells were humanized and that they possess the characteristics necessary for HIV infection of human host cells. Moreover, the sensitivity of human CD4/CCR5 transgenic mouse cells to HIV infection was confirmed by RT-PCR and ELISA. Mouse leukemia cell lines that could express hCD4 and CCR5 were thus established to facilitate normal entry of HIV-1 so that a human CD4/CCR5 transgenic mice cell model can be used to investigate the transmission and pathogenesis of HIV/AIDS and potential antiviral drugs against this disease.

  16. Interleukin-4 Expressed By Neoplastic Cells Provokes an Anti-Metastatic Myeloid Immune Response

    PubMed Central

    Zhang, Connie S.; Kim, Hyeyeon; Mullins, Graeme; Tyryshkin, Kathrin; LeBrun, David P.; Elliott, Bruce E.; Greer, Peter A.

    2016-01-01

    Objective Interleukin-4 (IL-4) can induce macrophages to undergo alternative activation and polarize toward an M2-like or wound healing phenotype. Tumor associated macrophages (TAMs) are thought to assume M2-like properties, and it has been suggested they promote tumor growth and metastasis through effects on the tumor stroma, including extracelluar matrix remodeling and angiogenesis. IL-4 also promotes macrophage survival and formation of multinucleated giant cells, which have enhanced phagocytic behavior. This study was designed to explore the effect of cancer cell derived IL-4 on the tumor immune stroma and metastasis. Methods The metastatic mouse mammary carcinoma cell line AC2M2 was transduced with control or IL-4 encoding retroviruses and employed in orthotopic engraftment models. Tumor growth and metastasis were assessed. The cellular composition and biomarker expression of tumors were examined by immunohistochemical staining and flow cytometry; the transcriptome of the immune stroma was analyzed by nanoString based transcript quantitation; and in vivo and in vitro interactions between cancer cells and macrophages were assessed by flow cytometry and co-culture with video-time lapse microscopy, respectively. Results Unexpectedly, tumors from IL-4 expressing AC2M2 engrafted cells grew at reduced rates, and most surprising, they lost all metastatic potential relative to tumors from control AC2M2 cells. Myeloid cell numbers were not increased in IL-4 expressing tumors, but their expression of the M2 marker arginase I was elevated. Transcriptome analysis revealed an immune signature consistent with IL-4 induced M2 polarization of the tumor microenvironment and a generalized increase in myeloid involvement in the tumor stroma. Flow cytometry analysis indicated enhanced cancer cell phagocytosis by TAMs from IL-4 expressing tumors, and co-culture studies showed that IL-4 expressing cancer cells supported the survival and promoted the in vitro phagocytic behavior of

  17. Increased expression with differential subcellular location of cytidine deaminase APOBEC3G in human CD4(+) T-cell activation and dendritic cell maturation.

    PubMed

    Oliva, Harold; Pacheco, Rodrigo; Martinez-Navio, José M; Rodríguez-García, Marta; Naranjo-Gómez, Mar; Climent, Núria; Prado, Carolina; Gil, Cristina; Plana, Montserrat; García, Felipe; Miró, José M; Franco, Rafael; Borras, Francesc E; Navaratnam, Naveenan; Gatell, José M; Gallart, Teresa

    2016-08-01

    APOBEC3G (apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 3G; A3G) is an innate defense protein showing activity against retroviruses and retrotransposons. Activated CD4(+) T cells are highly permissive for HIV-1 replication, whereas resting CD4(+) T cells are refractory. Dendritic cells (DCs), especially mature DCs, are also refractory. We investigated whether these differences could be related to a differential A3G expression and/or subcellular distribution. We found that A3G mRNA and protein expression is very low in resting CD4(+) T cells and immature DCs, but increases strongly following T-cell activation and DC maturation. The Apo-7 anti-A3G monoclonal antibody (mAb), which was specifically developed, confirmed these differences at the protein level and disclosed that A3G is mainly cytoplasmic in resting CD4(+) T cells and immature DCs. Nevertheless, A3G translocates to the nucleus in activated-proliferating CD4(+) T cells, yet remaining cytoplasmic in matured DCs, a finding confirmed by immunoblotting analysis of cytoplasmic and nuclear fractions. Apo-7 mAb was able to immunoprecipitate endogenous A3G allowing to detect complexes with numerous proteins in activated-proliferating but not in resting CD4(+) T cells. The results show for the first time the nuclear translocation of A3G in activated-proliferating CD4(+) T cells.

  18. 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.

  19. High expression Zymomonas promoters

    DOEpatents

    Viitanen, Paul V.; Tao, Luan; Zhang, Yuying; Caimi, Perry G.; McCole, Laura : Zhang, Min; Chou, Yat-Chen; McCutchen, Carol M.; Franden, Mary Ann

    2011-08-02

    Identified are mutants of the promoter of the Z. mobilis glyceraldehyde-3-phosphate dehydrogenase gene, which direct improved expression levels of operably linked heterologous nucleic acids. These are high expression promoters useful for expression of chimeric genes in Zymomonas, Zymobacter, and other related bacteria.

  20. [Rapid and efficient expression of foreign genes in mammalian cells by baculovirus vectors].

    PubMed

    Cheng, Tong; Xu, Chen-Yu; Wang, Ying-Bin; Chen, Min; Wu, Ting; Xie, Xiao-Yan; Zhang, Jun; Xia, Ning-Shao

    2003-09-01

    culture supernatant (BacV-CMV-EGFPA, 1.2 x 10(7) pfu/mL) serially diluted by DMEM culture medium containing 10% FBS and the transduction times ranged from 1 to 24 hours. Twenty-four hours post transduction the efficiencies of gene-transfer and expression were analyzed by FCM. Results show that incubating the target cells with the 1:1 diluted culture supernatant (moi = 50) for 12 hours in 37 degrees C CO2 incubator would achieve the highest infection and expression efficiency with the least impairment on cell viability. We compared the gene-transfer and expression efficiency of recombinant baculovirus in HepG2 and CV1 cells with lipofectAMINE and recombinant retrovirus system, results show that under the similar conditions the recombinant baculovirus could achieve the highest gene-transfer and expression efficiency than the other two systems. So we can draw a conclusion that directly incubating the mammalian cells with the culture supernantant of the infected Sf9 cells could serve as a very convenient way for rapid and efficient expression of foreign gene in mammalian cells.

  1. Protein expression-yeast.

    PubMed

    Nielsen, Klaus H

    2014-01-01

    Yeast is an excellent system for the expression of recombinant eukaryotic proteins. Both endogenous and heterologous proteins can be overexpressed in yeast (Phan et al., 2001; Ton and Rao, 2004). Because yeast is easy to manipulate genetically, a strain can be optimized for the expression of a specific protein. Many eukaryotic proteins contain posttranslational modifications that can be performed in yeast but not in bacterial expression systems. In comparison with mammalian cell culture expression systems, growing yeast is both faster and less expensive, and large-scale cultures can be performed using fermentation. While several different yeast expression systems exist, this chapter focuses on the budding yeast Saccharomyces cerevisiae and will briefly describe some options to consider when selecting vectors and tags to be used for protein expression. Throughout this chapter, the expression and purification of yeast eIF3 is shown as an example alongside a general scheme outline.

  2. Multiple sclerosis-associated virus-related pol sequences found both in multiple sclerosis and healthy donors are more frequently expressed in multiple sclerosis patients.

    PubMed

    Nowak, Jerzy; Januszkiewicz, Danuta; Pernak, Monika; Liweń, Izabela; Zawada, Mariola; Rembowska, Jolanta; Nowicka, Karina; Lewandowski, Krzysztof; Hertmanowska, Hanna; Wender, Mieczyslaw

    2003-02-01

    In the etiopathogenesis of multiple sclerosis (MS), both genetic and environmental factors play an important role. Among environmental factors, viral infections are most likely connected with the etiology of MS. There are many evidence suggesting possible involvement of retroviruses in the development of autoimmune diseases including MS. Multiple sclerosis-associated retrovirus (MSRV) seems to be the important candidate for viral etiology of MS. The aim of the study was to analyze MSRV pol sequences in patients with MS. As control, groups of myasthenia gravis, Parkinson's disease, and migraine patients, and healthy individuals have been studied. The MSRV pol sequences have been analyzed in RNA isolated from the serum and in DNA and RNA of peripheral blood lymphocytes from untreated MS patients and control groups. The MSRV pol sequences have been detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and PCR technique, using specific oligonucleotide primers. In the serum RNA (cDNA), MSRV pol sequences have been identified in 31/32 MS patients. MSRV pol sequences were detected in serum cDNA of 9/17 myasthenia gravis patients, 7/16 Parkinson's disease patients, 10/21 migraine patients, and 13/27 healthy individuals. MSRV pol sequences were observed also in RNA from lymphocytes of all MS patients, 12/17 myasthenia gravis patients, 9/16 Parkinson's disease patients, 14/21 migraine patients, and 18/27 healthy donors. In the DNA from peripheral blood lymphocytes of all studied patients and healthy individuals, MSRV pol sequences have been found. The observed pattern of fiber-fluorescence in situ hybridization (FISH) signals suggests the presence of multiple copies of MSRV pol sequences, most likely tandemly dispersed in the genome. It can be concluded that MSRV pol sequences are endogenous, widespread in lymphocytes DNA, and transcribed into RNA of MS patients as well as of other studied patients and healthy individuals. However, more frequent expression of

  3. Holistic facial expression classification

    NASA Astrophysics Data System (ADS)

    Ghent, John; McDonald, J.

    2005-06-01

    This paper details a procedure for classifying facial expressions. This is a growing and relatively new type of problem within computer vision. One of the fundamental problems when classifying facial expressions in previous approaches is the lack of a consistent method of measuring expression. This paper solves this problem by the computation of the Facial Expression Shape Model (FESM). This statistical model of facial expression is based on an anatomical analysis of facial expression called the Facial Action Coding System (FACS). We use the term Action Unit (AU) to describe a movement of one or more muscles of the face and all expressions can be described using the AU's described by FACS. The shape model is calculated by marking the face with 122 landmark points. We use Principal Component Analysis (PCA) to analyse how the landmark points move with respect to each other and to lower the dimensionality of the problem. Using the FESM in conjunction with Support Vector Machines (SVM) we classify facial expressions. SVMs are a powerful machine learning technique based on optimisation theory. This project is largely concerned with statistical models, machine learning techniques and psychological tools used in the classification of facial expression. This holistic approach to expression classification provides a means for a level of interaction with a computer that is a significant step forward in human-computer interaction.

  4. Interactions of the Cytoplasmic Domains of Human and Simian Retroviral Transmembrane Proteins with Components of the Clathrin Adaptor Complexes Modulate Intracellular and Cell Surface Expression of Envelope Glycoproteins

    PubMed Central

    Berlioz-Torrent, Clarisse; Shacklett, Barbara L.; Erdtmann, Lars; Delamarre, Lelia; Bouchaert, Isabelle; Sonigo, Pierre; Dokhelar, Marie Christine; Benarous, Richard

    1999-01-01

    The cytoplasmic domains of the transmembrane (TM) envelope proteins (TM-CDs) of most retroviruses have a Tyr-based motif, YXXØ, in their membrane-proximal regions. This signal is involved in the trafficking and endocytosis of membrane receptors via clathrin-associated AP-1 and AP-2 adaptor complexes. We have used CD8-TM-CD chimeras to investigate the role of the Tyr-based motif of human immunodeficiency virus type 1 (HIV-1), simian immunodeficiency virus (SIV), and human T-leukemia virus type 1 (HTLV-1) TM-CDs in the cell surface expression of the envelope glycoprotein. Flow cytometry and confocal microscopy studies showed that this motif is a major determinant of the cell surface expression of the CD8-HTLV chimera. The YXXØ motif also plays a key role in subcellular distribution of the envelope of lentiviruses HIV-1 and SIV. However, these viruses, which encode TM proteins with a long cytoplasmic domain, have additional determinants distal to the YXXØ motif that participate in regulating cell surface expression. We have also used the yeast two-hybrid system and in vitro binding assays to demonstrate that all three retroviral YXXØ motifs interact with the μ1 and μ2 subunits of AP complexes and that the C-terminal regions of HIV-1 and SIV TM proteins interact with the β2 adaptin subunit. The TM-CDs of HTLV-1, HIV-1, and SIV also interact with the whole AP complexes. These results clearly demonstrate that the cell surface expression of retroviral envelope glycoproteins is governed by interactions with adaptor complexes. The YXXØ-based signal is the major determinant of this interaction for the HTLV-1 TM, which contains a short cytoplasmic domain, whereas the lentiviruses HIV-1 and SIV have additional determinants distal to this signal that are also involved. PMID:9882340

  5. Ping-pong amplification of a retroviral vector achieves high-level gene expression: human growth hormone production.

    PubMed Central

    Kozak, S L; Kabat, D

    1990-01-01

    Retroviral vectors offer major advantages for gene transfer studies but have not been useful for producing proteins in large quantities. This deficiency has resulted in part from interference to superinfection, which limits the numbers of active proviruses in cells. Recently, we found that these vectors amplify when they are added as calcium phosphate precipitates to cocultures of cells that package retroviruses into ecotropic and amphotropic host range envelopes. Helper-free virions from either cell type can infect the other without interference, resulting in theoretically limitless back-and-forth (ping-pong) vector replication. In initial studies, however, amplifications of a vector that contained the human growth hormone gene ceased when the hormone produced was 0.3% or less of cellular protein synthesis. This limit was caused by two factors. First, recombinant shutoff viruses that are replication defective and encode envelope glycoproteins form at a low probability during any round of the vector replication cycle and these spread in cocultures, thereby establishing interference. Single cells in shutoff cocultures therefore synthesize both ecotropic and amphotropic envelope glycoproteins, and they release promiscuous (presumably hybrid) virions. The probability of forming shutoff viruses before the vector had amplified to a high multiplicity was reduced by using small cocultures. Second, cells with large numbers of proviruses are unhealthy and their proviral expression can be unstable. Stable expresser cell clones were obtained by selection. Thereby, cell lines were readily obtained that stably produce human growth hormone as 4 to 6% of the total protein synthesis. A ping-pong retroviral vector can be used for high-level protein production in vertebrate cells. Images PMID:2352330

  6. Expression of human beta-hexosaminidase alpha-subunit gene (the gene defect of Tay-Sachs disease) in mouse brains upon engraftment of transduced progenitor cells.

    PubMed

    Lacorazza, H D; Flax, J D; Snyder, E Y; Jendoubi, M

    1996-04-01

    In humans, beta-hexosaminidase alpha-subunit deficiency prevents the formation of a functional beta-hexosaminidase A heterodimer resulting in the severe neurodegenerative disorder, Tay-Sachs disease. To explore the feasibility of using ex vivo gene transfer in this lysosomal storage disease, we produced ecotropic retroviruses encoding the human beta-hexosaminidase alpha-subunit cDNA and transduced multipotent neural cell lines. Transduced progenitors stably expressed and secreted high levels of biologically active beta-hexosaminidase A in vitro and cross-corrected the metabolic defect in a human Tay-Sachs fibroblasts cell line in vitro. These genetically engineered CNS progenitors were transplanted into the brains of both normal fetal and newborn mice. Engrafted brains, analyzed at various ages after transplant, produced substantial amounts of human beta-hexosaminidase alpha-subunit transcript and protein, which was enzymatically active throughout the brain at a level reported to be therapeutic in Tay-Sachs disease. These results have implications for treating neurologic diseases characterized by inherited single gene mutations.

  7. 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.

  8. T-bet over-expression regulates aryl hydrocarbon receptor-mediated T helper type 17 differentiation through an interferon (IFN)γ-independent pathway.

    PubMed

    Yokosawa, M; Kondo, Y; Tahara, M; Iizuka-Koga, M; Segawa, S; Kaneko, S; Tsuboi, H; Yoh, K; Takahashi, S; Matsumoto, I; Sumida, T

    2017-04-01

    Various transcription factors are also known to enhance or suppress T helper type 17 (Th17) differentiation. We have shown previously that the development of collagen-induced arthritis was suppressed in T-bet transgenic (T-bet Tg) mice, and T-bet seemed to suppress Th17 differentiation through an interferon (IFN)-γ-independent pathway, although the precise mechanism remains to be clarified. The present study was designed to investigate further the mechanisms involved in the regulation of Th17 differentiation by T-bet over-expression, and we found the new relationship between T-bet and aryl hydrocarbon receptor (AHR). Both T-bet Tg mice and IFN-γ(-/-) -over-expressing T-bet (T-bet Tg/IFN-γ(-/-) ) mice showed inhibition of retinoic acid-related orphan receptor (ROR)γt expression and IL-17 production by CD4(+) T cells cultured under conditions that promote Th-17 differentiation, and decreased IL-6 receptor (IL-6R) expression and signal transducer and activator of transcription-3 (STAT-3) phosphorylation in CD4(+) T cells. The mRNA expression of ahr and rorc were suppressed in CD4(+) T cells cultured under Th-17 conditions from T-bet Tg mice and T-bet Tg/IFN-γ(-/-) mice. CD4(+) T cells of wild-type (WT) and IFN-γ(-/-) mice transduced with T-bet-expressing retrovirus also showed inhibition of IL-17 production, whereas T-bet transduction had no effect on IL-6R expression and STAT-3 phosphorylation. Interestingly, the mRNA expression of ahr and rorc were suppressed in CD4(+) T cells with T-bet transduction cultured under Th17 conditions. The enhancement of interleukin (IL)-17 production from CD4(+) T cells by the addition of AHR ligand with Th17 conditions was cancelled by T-bet over-expression. Our findings suggest that T-bet over-expression-induced suppression of Th17 differentiation is mediated through IFN-γ-independent AHR suppression.

  9. Viral vectors: from virology to transgene expression

    PubMed Central

    Bouard, D; Alazard-Dany, N; Cosset, F-L

    2009-01-01

    In the late 1970s, it was predicted that gene therapy would be applied to humans within a decade. However, despite some success, gene therapy has still not become a routine practise in medicine. In this review, we will examine the problems, both experimental and clinical, associated with the use of viral material for transgenic insertion. We shall also discuss the development of viral vectors involving the most important vector types derived from retroviruses, adenoviruses, herpes simplex viruses and adeno-associated viruses. This article is part of a themed section on Vector Design and Drug Delivery. For a list of all articles in this section see the end of this paper, or visit: http://www3.interscience.wiley.com/journal/121548564/issueyear?year=2009 PMID:18776913

  10. Cloning of the rat ecotropic retroviral receptor and studies of its expression in intestinal tissues.

    PubMed

    Puppi, M; Henning, S J

    1995-05-01

    A long-term goal of our laboratory is to establish a rat model to study the feasibility of using the intestinal tract as a site for somatic gene therapy. As a step toward that goal, the current study reports the cloning of the rat ecotropic retroviral receptor (EcoR) cDNA and the study of various aspects of its expression in the intestinal tissues. The cDNA for rat EcoR was cloned by screening a size-selected rat intestinal cDNA library with mouse EcoR cDNA. A clone of approximately 7 kb, designated MP10, was obtained. Partial sequencing of MP10 from the 5' end revealed a level of similarity of 92% compared with mouse EcoR. The presence of a 5' untranslated region and a 3' poly(A) tract, together with the overall size of the cDNA, suggest that is very close to being a full-length cDNA for this large transcript. Northern blots with MP10 showed an RNA of approximately 7.9 kb present along the entire length of the small intestine and somewhat less abundant in the colon. Developmental studies showed high levels of EcoR in fetal rat intestine, a decline in the early postnatal period, then a gradual rise to adulthood. Caco-2 cells were used to assess the expression of EcoR in proliferating compared with differentiated intestinal epithelial cells. EcoR mRNA was found to be very much more abundant in nondifferentiated cells and declined to low levels as the cells underwent spontaneous differentiation. These patterns of EcoR expression indicate that ecotropic retroviruses should be suitable vectors with which to attempt gene transfer into the intestinal epithelium. In addition, since the endogenous role of EcoR is as the y+ cationic amino acid transporter, these data have significance for understanding patterns of amino acid transport in the intestinal epithelium.

  11. Guidelines on Open Expression.

    ERIC Educational Resources Information Center

    Pennsylvania Univ., Philadelphia.

    These Guidelines on open expression at the University of Pennsylvania include: (1) a statement of principles, expressing support for freedom of thought, inquiry, speech and lawful assembly, and for the need to ensure continuing openness and effectiveness of channels of communication; (2) a description of the newly created Committee on Open…

  12. Facial expression and sarcasm.

    PubMed

    Rockwell, P

    2001-08-01

    This study examined facial expression in the presentation of sarcasm. 60 responses (sarcastic responses = 30, nonsarcastic responses = 30) from 40 different speakers were coded by two trained coders. Expressions in three facial areas--eyebrow, eyes, and mouth--were evaluated. Only movement in the mouth area significantly differentiated ratings of sarcasm from nonsarcasm.

  13. Darwin and Emotion Expression

    ERIC Educational Resources Information Center

    Hess, Ursula; Thibault, Pascal

    2009-01-01

    In his book "The Expression of the Emotions in Man and Animals," Charles Darwin (1872/1965) defended the argument that emotion expressions are evolved and adaptive (at least at some point in the past) and serve an important communicative function. The ideas he developed in his book had an important impact on the field and spawned rich domains of…

  14. EXPRESS Rack Mockup

    NASA Technical Reports Server (NTRS)

    2002-01-01

    The EXPRESS Rack is a standardized payload rack system that transports, stores, and supports experiments aboard the International Space Station (ISS). EXPRESS stands for EXpedite the PRocessing of Experiments to the Space Station, reflecting the fact that this system was developed specifically to maximize the Station's research capabilities. The EXPRESS Rack system supports science payloads in several disciplines, including biology, chemistry, physics, ecology, and medicine. With the EXPRESS Rack, getting experiments to space has never been easier or more affordable. With its standardized hardware interfaces and streamlined approach, the EXPRESS Rack enables quick, simple integration of multiple payloads aboard the ISS. The system is comprised of elements that remain on the ISS, as well as elements that travel back and forth between the ISS and Earth via the Space Shuttle. The Racks stay on orbit continually, while experiments are exchanged in and out of the EXPRESS Racks as needed, remaining on the ISS for three months to several years, depending on the experiment's time requirements. A refrigerator-sized Rack can be divided into segments, as large as half of an entire rack or as small as a bread box. Payloads within EXPRESS Racks can operate independently of each other, allowing for differences in temperature, power levels, and schedules. Experiments contained within EXPRESS Racks may be controlled by the ISS crew or remotely by the Payload Rack Officer at the Payload Operations Center at the Marshall Space Flight Center (MSFC). The EXPRESS Rack system was developed by MSFC and built by the Boeing Co. in Huntsville, Alabama. Eight EXPRESS Racks are being built for use on the ISS.

  15. Long-term Persistence of CD4+ but Rapid Disappearance of CD8+ T Cells Expressing an MHC Class I-restricted TCR of Nanomolar Affinity

    PubMed Central

    Engels, Boris; Chervin, Adam S; Sant, Andrea J; Kranz, David M; Schreiber, Hans

    2012-01-01

    Most T cells have T cell receptors (TCR) of micromolar affinity for peptide-major histocompatibility complex (MHC) ligands, but genetic engineering can generate TCRs of nanomolar affinity. The affinity of the TCR used, m33, for its cognate non-self peptide–MHC-I complex (SIYRYYGL-Kb) is 1,000-fold higher than of the wild-type TCR 2C. The affinity of m33 for the self-peptide dEV-8 on Kb is only twofold higher. Mouse CD8+ T cells transduced with an m33-encoding retrovirus showed binding of SIY-Kb and potent function in vitro, but in vivo these T cells disappeared within hours after transfer into syngeneic hosts without causing graft-versus-host disease (GVHD). Accordingly, in cases where such CD8-dependent self-reactivity might occur in human adoptive T cell therapies, our results show that a peripheral T-cell deletion mechanism could operate to avoid reactions with the host. In contrast to CD8+ T cells, we show that CD4+ T cells expressing m33 survived for months in vivo. Furthermore, the m33-transduced CD4+ T cells were able to mediate antigen-specific rejection of 6-day-old tumors. Together, we show that CD8+ T cell expressing a MHC class I-restricted high-affinity TCR were rapidly deleted whereas CD4+ T cells expressing the same TCR survived and provided function while being directed against a class I-restricted antigen. PMID:22233579

  16. Insights into gene expression changes impacting B-cell transformation: cross-species microarray analysis of bovine leukemia virus tax-responsive genes in ovine B cells.

    PubMed

    Klener, Pavel; Szynal, Maud; Cleuter, Yvette; Merimi, Makram; Duvillier, Hugues; Lallemand, Françoise; Bagnis, Claude; Griebel, Philip; Sotiriou, Christos; Burny, Arsène; Martiat, Philippe; Van den Broeke, Anne

    2006-02-01

    Large-animal models for leukemia have the potential to aid in the understanding of networks that contribute to oncogenesis. Infection of cattle and sheep with bovine leukemia virus (BLV), a complex retrovirus related to human T-cell leukemia virus type 1 (HTLV-1), is associated with the development of B-cell leukemia. Whereas the natural disease in cattle is characterized by a low tumor incidence, experimental infection of sheep leads to overt leukemia in the majority of infected animals, providing a model for studying the pathogenesis associated with BLV and HTLV-1. Tax(BLV), the major oncoprotein, initiates a cascade of events leading toward malignancy, although the basis of transformation is not fully understood. We have taken a cross-species ovine-to-human microarray approach to identify Tax(BLV)-responsive transcriptional changes in two sets of cultured ovine B cells following retroviral vector-mediated delivery of Tax(BLV). Using cDNA-spotted microarrays comprising 10,336 human genes/expressed sequence tags, we identified a cohort of differentially expressed genes, including genes related to apoptosis, DNA transcription, and repair; proto-oncogenes; cell cycle regulators; transcription factors; small Rho GTPases/GTPase-binding proteins; and previously reported Tax(HTLV-1)-responsive genes. Interestingly, genes known to be associated with human neoplasia, especially B-cell malignancies, were extensively represented. Others were novel or unexpected. The results suggest that Tax(BLV) deregulates a broad network of interrelated pathways rather than a single B-lineage-specific regulatory process. Although cross-species approaches do not permit a comprehensive analysis of gene expression patterns, they can provide initial clues for the functional roles of genes that participate in B-cell transformation and pinpoint molecular targets not identified using other methods in animal models.

  17. Cellular and species resistance to murine amphotropic, gibbon ape, and feline subgroup C leukemia viruses is strongly influenced by receptor expression levels and by receptor masking mechanisms.

    PubMed

    Tailor, C S; Nouri, A; Kabat, D

    2000-10-01

    Chinese hamster ovary (CHO) cells are resistant to infections by gibbon ape leukemia virus (GALV) and amphotropic murine leukemia virus (A-MLV) unless they are pretreated with tunicamycin, an inhibitor of N-linked glycosylation. These viruses use the related sodium-phosphate symporters Pit1 and Pit2, respectively, as receptors in nonhamster cells, and evidence has suggested that the corresponding transporters of CHO cells may be masked by tunicamycin-sensitive secreted inhibitors. Although the E36 line of Chinese hamster cells was reported to secrete the putative Pit2 inhibitor and to be sensitive to the inhibitory CHO factors, E36 cells are highly susceptible to both GALV and A-MLV in the absence of tunicamycin. Moreover, expression of E36 Pit2 in CHO cells conferred tunicamycin-independent susceptibilities to both viruses. Based on the latter results, it was suggested that E36 Pit2 must functionally differ from the endogenous Pit2 of CHO cells. To test these ideas, we analyzed the receptor properties of CHO Pit1 and Pit2 in CHO cells. Surprisingly, and counterintuitively, transfection of a CHO Pit2 expression vector into CHO cells conferred strong susceptibility to both GALV and A-MLV, and similar overexpression of CHO Pit1 conferred susceptibility to GALV. Thus, CHO Pit2 is a promiscuous functional receptor for both viruses, and CHO Pit1 is a functional receptor for GALV. Similarly, we found that the natural resistance of Mus dunni tail fibroblasts to subgroup C feline leukemia viruses (FeLV-C) was eliminated simply by overexpression of the endogenous FeLV-C receptor homologue. These results demonstrate a novel and simple method to unmask latent retroviral receptor activities that occur in some cells. Specifically, resistances to retroviruses that are caused by subthreshold levels of receptor expression or by stoichiometrically limited masking or interference mechanisms can be efficiently overcome simply by overexpressing the endogenous receptors in the same

  18. Dense array expressions

    NASA Astrophysics Data System (ADS)

    Wilson, Joseph N.; Chen, LiangMing

    1999-10-01

    Various researchers have realized the value of implementing loop fusion to evaluate dense (pointwise) array expressions. Recently, the method of template metaprogramming in C++ has been used to significantly speed-up the evaluation of array expressions, allowing C++ programs to achieve performance comparable to or better than FORTRAN for numerical analysis applications. Unfortunately, the template metaprogramming technique suffers from several limitations in applicability, portability, and potential performance. We present a framework for evaluating dense array expressions in object-oriented programming languages. We demonstrate how this technique supports both common subexpression elimination and threaded implementation and compare its performance to object-library and hand-generated code.

  19. Murine cytomegalovirus infection of mouse macrophages stimulates early expression of suppressor of cytokine signaling (SOCS)1 and SOCS3

    PubMed Central

    Alston, Christine I.; Dix, Richard D.

    2017-01-01

    Human cytomegalovirus (HCMV) is a species-specific β-herpesvirus that infects for life up to 80% of the world’s population and causes severe morbidity in at-risk immunocompromised populations. Suppressors of cytokine signaling (SOCS)1 and SOCS3 are host proteins that act as inducible negative feedback regulators of cytokine signaling and have been implicated in several ocular diseases and viral infections. We recently found in our mouse model of experimental cytomegalovirus retinitis that subretinally-injected murine cytomegalovirus (MCMV) stimulates ocular SOCS1 and SOCS3 during retrovirus-induced immune suppression of murine AIDS (MAIDS), and that infiltrating macrophages are prominent cellular sources of retinal SOCS1 and SOCS3 expression. Herein we investigate possible virologic mechanisms whereby MCMV infection may stimulate SOCS1 and/or SOCS3 expression in cell culture. We report that infection of IC-21 mouse macrophages with MCMV propagated through the salivary glands of BALB/c mice, but not from tissue culture in C57BL/6 fibroblasts, transiently stimulates SOCS1 and SOCS3 mRNA transcripts, but not SOCS5 mRNA. Viral tegument proteins are insufficient for this stimulation, as replication-deficient UV-inactivated MCMV fails to stimulate SOCS1 or SOCS3 in IC-21 macrophages. By contrast, infection of murine embryonic fibroblasts (MEFs) with either productive MCMV or UV-inactivated MCMV significantly stimulates SOCS1 and SOCS3 mRNA expression early after infection. Treatment of MCMV-infected IC-21 mouse macrophages with the antiviral drug ganciclovir significantly decreases MCMV-stimulated SOCS3 expression at 3 days post-infection. These data suggest cell type-specific, different roles for viral immediate early or early gene expression and/or viral tegument proteins in the early stimulation of SOCS1 and SOCS3 during MCMV infection. Furthermore, putative biphasic stimulation of SOCS3 during late MCMV infection of IC-21 mouse macrophages may occur by divergent

  20. Deciphering the Code for Retroviral Integration Target Site Selection

    PubMed Central

    Santoni, Federico Andrea; Hartley, Oliver; Luban, Jeremy

    2010-01-01

    Upon cell invasion, retroviruses generate a DNA copy of their RNA genome and integrate retroviral cDNA within host chromosomal DNA. Integration occurs throughout the host cell genome, but target site selection is not random. Each subgroup of retrovirus is distinguished from the others by attraction to particular features on chromosomes. Despite extensive efforts to identify host factors that interact with retrovirion components or chromosome features predictive of integration, little is known about how integration sites are selected. We attempted to identify markers predictive of retroviral integration by exploiting Precision-Recall methods for extracting information from highly skewed datasets to derive robust and discriminating measures of association. ChIPSeq datasets for more than 60 factors were compared with 14 retroviral integration datasets. When compared with MLV, PERV or XMRV integration sites, strong association was observed with STAT1, acetylation of H3 and H4 at several positions, and methylation of H2AZ, H3K4, and K9. By combining peaks from ChIPSeq datasets, a supermarker was identified that localized within 2 kB of 75% of MLV proviruses and detected differences in integration preferences among different cell types. The supermarker predicted the likelihood of integration within specific chromosomal regions in a cell-type specific manner, yielding probabilities for integration into proto-oncogene LMO2 identical to experimentally determined values. The supermarker thus identifies chromosomal features highly favored for retroviral integration, provides clues to the mechanism by which retrovirus integration sites are selected, and offers a tool for predicting cell-type specific proto-oncogene activation by retroviruses. PMID:21124862

  1. Disorder of written expression

    MedlinePlus

    ... expression is a childhood condition that involves poor writing skills. Causes Specific learning disorder with impairment in ... and punctuation Poor handwriting Poor spelling Poorly organized writing Exams and Tests Other causes of learning disabilities ...

  2. Visualization of the human CD4{sup +} T-cell response in humanized HLA-DR4-expressing NOD/Shi-scid/γc{sup null} (NOG) mice by retrogenic expression of the human TCR gene

    SciTech Connect

    Takahashi, Takeshi Katano, Ikumi; Ito, Ryoji; Ito, Mamoru

    2015-01-02

    Highlights: • β-Lactoglobulin (BLG) specific TCR genes were introduced to human HSC by retrovirus. • Human HSC with BLG-specific TCR were transplanted into NOG-HLA-DR4 I-A{sup −/−} mice. • BLG-specific TCR induced positive selection of thymocytes. • BLG-specific TCR positive CD4{sup +} T cells mediated immune responses in humanized mice. - Abstract: The development of severe immunodeficient mouse strains containing various human genes, including cytokines or HLA, has enabled the reconstitution of functional human immune systems after transplantation of human hematopoietic stem cells (HSC). Accumulating evidence has suggested that HLA-restricted antigen-specific human T-cell responses can be generated in these humanized mice. To directly monitor immune responses of human CD4{sup +} T cells, we introduced β-lactoglobulin (BLG)-specific T cell receptor (TCR) genes derived from CD4{sup +} T-cell clones of cow-milk allergy patients into HSCs, and subsequently transplanted them into NOG-HLA-DR4 transgenic/I-Aβ deficient mice (NOG-DR4/I-A{sup o}). In the thymus, thymocytes with BLG-specific TCR preferentially differentiated into CD4{sup +}CD8{sup −} single-positive cells. Adoptive transfer of mature CD4{sup +} T cells expressing the TCR into recipient NOG-DR4/I-A{sup o} mice demonstrated that human CD4{sup +} T cells proliferated in response to antigenic stimulation and produced IFN-γ in vivo, suggesting that functional T-cell reactions (especially Th1-skewed responses) were induced in humanized mice.

  3. Memory beyond expression.

    PubMed

    Delorenzi, A; Maza, F J; Suárez, L D; Barreiro, K; Molina, V A; Stehberg, J

    2014-01-01

    The idea that memories are not invariable after the consolidation process has led to new perspectives about several mnemonic processes. In this framework, we review our studies on the modulation of memory expression during reconsolidation. We propose that during both memory consolidation and reconsolidation, neuromodulators can determine the probability of the memory trace to guide behavior, i.e. they can either increase or decrease its behavioral expressibility without affecting the potential of persistent memories to be activated and become labile. Our hypothesis is based on the findings that positive modulation of memory expression during reconsolidation occurs even if memories are behaviorally unexpressed. This review discusses the original approach taken in the studies of the crab Neohelice (Chasmagnathus) granulata, which was then successfully applied to test the hypothesis in rodent fear memory. Data presented offers a new way of thinking about both weak trainings and experimental amnesia: memory retrieval can be dissociated from memory expression. Furthermore, the strategy presented here allowed us to show in human declarative memory that the periods in which long-term memory can be activated and become labile during reconsolidation exceeds the periods in which that memory is expressed, providing direct evidence that conscious access to memory is not needed for reconsolidation. Specific controls based on the constraints of reminders to trigger reconsolidation allow us to distinguish between obliterated and unexpressed but activated long-term memories after amnesic treatments, weak trainings and forgetting. In the hypothesis discussed, memory expressibility--the outcome of experience-dependent changes in the potential to behave--is considered as a flexible and modulable attribute of long-term memories. Expression seems to be just one of the possible fates of re-activated memories.

  4. Immunotherapies for Targeting Ancient Retrovirus during Breast Cancer

    DTIC Science & Technology

    2014-03-01

    fluorescent protein, EGFP)11,12. Irradiated aAPCs were supplemented every 7 days to stimulate the growth of HERV-K-specific CAR+ T cells. PBMCs without...K CAR. The CAR+ T cells were stained with the Fc antibody every week before the supplementing the culture with irradiated aAPCs. Flow data revealed...enhanced green fluorescent protein, EGFP)7, 8. Irradiated aAPCs were supplemented every 7 days to stimulate the growth of HERV-K-specific CAR+ T

  5. Retrovirus Epidemiology Donor Study-II (REDS-II)

    ClinicalTrials.gov

    2016-04-14

    Acquired Immunodeficiency Syndrome; Blood Donors; Blood Transfusion; HIV Infections; HIV-1; HIV-2; HTLV-I; HTLV-II; Retroviridae Infections; Hepatitis, Viral, Human; Hepatitis B; Hepacivirus; West Nile Virus

  6. Pluripotency and the endogenous retrovirus HERVH: Conflict or serendipity?

    PubMed

    Izsvák, Zsuzsanna; Wang, Jichang; Singh, Manvendra; Mager, Dixie L; Hurst, Laurence D

    2016-01-01

    Remnants of ancient retroviral infections during evolution litter all mammalian genomes. In modern humans, such endogenous retroviral (ERV) sequences comprise at least 8% of the genome. While ERVs and other types of transposable elements undoubtedly contribute to the genomic "junk yard", functions for some ERV sequences have been demonstrated, with growing evidence that ERVs can be important players in gene regulatory processes. Here we focus on one particular large family of human ERVs, termed HERVH, which several recent studies suggest has a key regulatory role in human pluripotent stem cells. Remarkably, this is not the first instance of an ERV controlling pluripotency. We speculate as to why this convergent evolution might have come about, suggesting that it may reflect selection on the virus to extend the time available for transposition. Alternatively it may reflect serendipity alone.

  7. The mongoose, the pheasant, the pox, and the retrovirus.

    PubMed

    Etienne, Lucie; Emerman, Michael

    2013-01-01

    Paleovirology is the study of ancient viruses. The existence of a paleovirus can sometimes be detected by virtue of its accidental insertion into the germline of different animal species, which allows one to date when the virus actually existed. However, the ancient and the modern often connect, as modern viruses have unexpected origins that can be traced to ancient infections. The genomes of two species of mongooses and an egg-laying mammal called an echidna show that a virus currently present in poultry, the reticuloendotheliosis virus (REV), is actually of ancient exotic mammalian origin. REV apparently spread to poultry through a circuitous route involving the isolation of malaria parasites from a pheasant from Borneo housed at the Bronx Zoo that was contaminated with REV. Repeated passage of this virus in poultry adapted the virus to its new host. At some point, the virus got inserted into another virus, called fowlpox virus, which has spread back into the wild. Although REV may still exist somewhere in a mammalian host, its modern form links an 8 million-year-old infection of the ancestor of a mongoose to a virus that now is circulating in wild birds through malaria studies in the mid-20(th) century. These lessons of ancient and modern viruses have implications for modern human pandemics from viral reservoirs and for human interventions that may come with unintended consequences.

  8. Characterization of African Human Retroviruses Related to HTLV-111/LAV

    DTIC Science & Technology

    1990-10-02

    AND DEVELOPMENT COMMAND Fort Detrick, Frederick, Maryland 21702-5012 Contract No. DAMD17-87-C-7072 Harvard School of Public Health Boston, MA 02115...Harvard School of Public (If applicable) Health 1 6c. ADDRESS (City, State, and ZIP Code) 7b. ADDRESS (City, State, and ZIP Code) Boston, MA 02115 Ba...Natural History of HIV-2 In Senegal in 1966, prostitution was legalized in conjunction with a nationally supported program for Sexually Transmitted

  9. [New human retroviruses: HTLV-3 and HTLV-4].

    PubMed

    Mahieux, R; Gessain, A

    2005-11-01

    Human T cell leukemia/lymphoma virus Type 1 and 2 (HTLV-1 and HTLV-2), together with their simian counterparts (STLV-1, STLV-2 and STLV-3), belong to the Primate T lymphotropic viruses group (PTLV). HTLV-1 infects 15 to 20 million people worldwide, while STLV-1 is endemic in a number of simian species living in the Old World. Due to the high percentage of homologies between HTLV-1 and STLV-1 strains, it has now been widely accepted that most HTLV-1 subtypes arose from interspecies transmission between monkeys and humans. On the opposite, there is no close human homolog of the two STLV-2 strains that have been discovered in African bonobos chimpanzees. These results suggest that the interspecies transmission that lead to the present day HTLV-2 must have occurred in a distant past. STLV-3 viruses are very divergent, both from HTLV-1 and from HTLV-2. They are endemic in several monkey species that live in west, central and east Africa. Recently, two laboratories independently reported the discovery of the human homolog (HTLV-3) of STLV-3 in two inhabitants from south Cameroon whose sera exhibited HTLV indeterminate serologies. Together with STLV-3, these two viruses belong therefore to the PTLV-3 group. In addition, a fourth HTLV type (HTLV-4) was also discovered in the same geographical area. Current studies are aimed at determining the molecular characterization of these viruses. In particular, the possible oncogenic properties of their viral transactivator Tax is being investigated, as well as their modes of transmission and their possible association with human diseases.

  10. Synthetically functionalized retroviruses produced from the bioorthogonally engineered cell surface.

    PubMed

    Wong, Shirley; Kwon, Young Jik

    2011-02-16

    Conjugation of desired molecules onto retroviral surfaces through the ease of the bioorthogonal functionalization method was demonstrated. Oxidation of surface sialic acids using periodate and further p-anisidine-catalyzed conjugation with aminooxy-bearing molecules were used to directly label retroviral envelope with a fluorescent dye. The retroviral particles that were produced from a bioorthogonally functionalized virus producing cell surface and further tethered with magnetic nanoparticles were efficiently purified by simple magnetic column separation and capable of magnet-directed transduction.

  11. Genetic modification of lymphocytes by retrovirus-based vectors.

    PubMed

    Suerth, Julia D; Schambach, Axel; Baum, Christopher

    2012-10-01

    The genetic modification of lymphocytes is an important topic in the emerging field of gene therapy. Many clinical trials targeting immunodeficiency syndromes or cancer have shown therapeutic benefit; further applications address inflammatory and infectious disorders. Retroviral vector development requires a detailed understanding of the interactions with the host. Most researchers have used simple gammaretroviral vectors to modify lymphocytes, either directly or via hematopoietic stem and progenitor cells. Lentiviral, spumaviral (foamyviral) and alpharetroviral vectors were designed to reduce the necessity for cell stimulation and to utilize potentially safer integration properties. Novel surface modifications (pseudotyping) and transgenes, built using synthetic components, expand the retroviral toolbox, altogether promising increased specificity and potency. Product consistency will be an important criterion for routine clinical use.

  12. Selectivity of face aftereffects for expressions and anti-expressions.

    PubMed

    Juricevic, Igor; Webster, Michael A

    2012-01-01

    Adapting to a facial expression can alter the perceived expression of subsequently viewed faces. However, it remains unclear whether this adaptation affects each expression independently or transfers from one expression to another, and whether this transfer impedes or enhances responses to a different expression. To test for these interactions, we probed the basic expressions of anger, fear, happiness, sadness, surprise, and disgust, adapting to one expression and then testing on all six. Each expression was varied in strength by morphing it with a common neutral facial expression. Observers determined the threshold level required to correctly identify each expression, before or after adapting to a face with a neutral or intense expression. The adaptation was strongly selective for the adapting category; responses to the adapting expression were reduced, while other categories showed little consistent evidence of either suppression or facilitation. In a second experiment we instead compared adaptation to each expression and its anti-expression. The latter are defined by the physically complementary facial configuration, yet appear much more ambiguous as expressions. In this case, for most expressions the opposing faces produced aftereffects of opposite sign in the perceived expression. These biases suggest that the adaptation acts in part by shifting the perceived neutral point for the facial configuration. This is consistent with the pattern of renormalization suggested for adaptation to other facial attributes, and thus may reflect a generic level of configural coding. However, for most categories aftereffects were stronger for expressions than anti-expressions, pointing to the possible influence of an additional component of the adaptation at sites that explicitly represent facial expressions. At either level our results are consistent with other recent work in suggesting that the six expressions are defined by dimensions that are largely independently

  13. Human T-cell leukemia virus type 1 expressing nonoverlapping tax and rex genes replicates and immortalizes primary human T lymphocytes but fails to replicate and persist in vivo.

    PubMed

    Younis, Ihab; Yamamoto, Brenda; Phipps, Andrew; Green, Patrick L

    2005-12-01

    Human T-cell leukemia virus type 1 (HTLV-1) is an oncogenic retrovirus associated primarily with adult T-cell leukemia and neurological disease. HTLV-1 encodes the positive trans-regulatory proteins Tax and Rex, both of which are essential for viral replication. Tax activates transcription initiation from the viral long terminal repeat and modulates the transcription or activity of a number of cellular genes. Rex regulates gene expression posttranscriptionally by facilitating the cytoplasmic expression of incompletely spliced viral mRNAs. Tax and Rex mutants have been identified that have defective activities or impaired biochemical properties associated with their function. To ultimately determine the contribution of specific protein activities on viral replication and cellular transformation of primary T cells, mutants need to be characterized in the context of an infectious molecular clone. Since the tax and rex genes are in partially overlapping reading frames, mutation in one gene frequently disrupts the other, confounding interpretation of mutational analyses in the context of the virus. Here we generated and characterized a unique proviral clone (H1IT) in which the tax and rex genes were separated by expressing Tax from an internal ribosome entry site. We showed that H1IT expresses both functional Tax and Rex. In short- and long-term coculture assays, H1IT was competent to infect and immortalize primary human T cells similar to wild-type HTLV-1. In contrast, H1IT failed to efficiently replicate and persist in inoculated rabbits, thus emphasizing the importance of temporal and quantitative regulation of specific mRNA for viral survival in vivo.

  14. Antitumor activity of chimeric immunoreceptor gene-modified Tc1 and Th1 cells against autologous carcinoembryonic antigen-expressing colon cancer cells.

    PubMed

    Sasaki, Takeshi; Ikeda, Hiroaki; Sato, Masayoshi; Ohkuri, Takayuki; Abe, Hiroyuki; Kuroki, Masahide; Onodera, Masafumi; Miyamoto, Masaki; Kondo, Satoshi; Nishimura, Takashi

    2006-09-01

    To generate tumor-specific and interferon (IFN)-gamma-producing Tc1 and Th1 cells applicable for many cancer patients, we previously developed a protocol for generating carcinoembryonic antigen (CEA)-specific Tc1 and Th1 cells from healthy human T cells by transduction with a lentivirus containing a chimeric immunoglobulin T-cell receptor (cIgTCR) gene composed of single-chain variable fragments from an anti-CEA-specific monoclonal antibody fused to an intracellular signaling domain of CD28 and CD3zeta. These cells, designated Tc1-T and Th1-T bodies, respectively, showed strong antitumor activity against CEA-expressing tumor cells in RAG2-/- mice when both of them were transferred. However, it remains unclear whether it is possible to generate Tc1-T and Th1-T bodies from cancer patients with defective T-cell function because of significant immunosuppression. Here, we prepared Tc1-T and Th1-T bodies from T cells of a colon cancer patient, and asked whether these T bodies can exert effective T-cell function against autologous tumor cells. These T bodies showed high cytotoxicity and produced IFN-gamma in response to CEA-expressing autologous tumor cells, even in the presence of soluble CEA. It was also demonstrated that Th1-T bodies supported the survival of Tc1-T bodies through cell-to-cell interactions. Furthermore, our protocol utilized retrovirus for cIgTCR transduction to achieve better induction efficiency compared to lentivirus-mediated transduction. Taken together, our findings here indicate that retrovirally transduced Tc1-T and Th1-T bodies will become a promising strategy for adoptive immunotherapy of human cancer.

  15. Facial expressions recognition with an emotion expressive robotic head

    NASA Astrophysics Data System (ADS)

    Doroftei, I.; Adascalitei, F.; Lefeber, D.; Vanderborght, B.; Doroftei, I. A.

    2016-08-01

    The purpose of this study is to present the preliminary steps in facial expressions recognition with a new version of an expressive social robotic head. So, in a first phase, our main goal was to reach a minimum level of emotional expressiveness in order to obtain nonverbal communication between the robot and human by building six basic facial expressions. To evaluate the facial expressions, the robot was used in some preliminary user studies, among children and adults.

  16. Experience and Expression

    ERIC Educational Resources Information Center

    Hanes, Jay Michael; Weisman, Eleanor

    2016-01-01

    Two artist-educators analyzed their creative process informed by John Dewey's concepts regarding the act of expression. The essay interweaves a description of their performance piece with a discussion of conceptual processes, including intermediality and collaboration as crucial in art making, learning, and pedagogical efficacy. Both the creation…

  17. Fostering Creative Expression.

    ERIC Educational Resources Information Center

    Hunter, Elizabeth

    1968-01-01

    Teachers can encourage youngsters to express their ideas creatively by providing help in three areas--content, language, and process. In terms of content, children often have few resources for tapping their thoughts, and may need 'pump primers' such as being told the beginning and end of a story and speculating about a variety of middles. Once…

  18. Adaptations for Written Expression.

    ERIC Educational Resources Information Center

    Lambie, Rosemary Anne

    1986-01-01

    Remedial tactics are offered for five problem areas in written expression: (1) difficulty beginning, (2) problems translating ideas to paper, (3) poor re-writing or editing, (4) lack of motivation to become involved in a writing assignment, and (5) difficulties in note-taking during class instruction. (CL)

  19. Encircling Creative Expression

    ERIC Educational Resources Information Center

    Brown, Susannah

    2007-01-01

    Artworks that are circular in nature are often referred to as mandalas. "Mandala" means center, circle, or circumference. Mandalas are created in many cultures for a variety of reasons, most of which are related to self-expression, ritual, and religion. In this article, the author describes how her students created mandalas. She also provides…

  20. Expressions of Liberty.

    ERIC Educational Resources Information Center

    Fitch, Nancy Elizabeth

    The concept of liberty has been in the forefront of the minds of African Americans ever since the beginning of slavery, and its importance continues to the present. To cope with the inability to achieve complete freedom, and with the oppressive state created by a lack of liberty, they developed ways to express their feelings about the elusiveness…

  1. Expression of Concern

    NASA Astrophysics Data System (ADS)

    Delvaux, Damien

    2016-08-01

    This is a note of a temporary expression of concern related to the publication titled, "Sapphirine and fluid inclusions in Tel Thanoun mantle xenoliths, Syria" by Ahmad Bilal, which appeared in Journal of African Earth Sciences, 116 (2016) 105-113.

  2. Expression of concern

    NASA Astrophysics Data System (ADS)

    2017-02-01

    This is an expression of concern related to the following publications: Nanostructures formed by cyclodextrin covered procainamide through supramolecular self assembly - Spectral and molecular modeling study (2015) Spectrochimica Acta - Part A: Molecular and Biomolecular Spectroscopy, 136 (PB), pp. 875-883, by Rajendiran, N., Mohandoss, T., Sankaranarayanan, R.K.

  3. Expressive Costume Portraits

    ERIC Educational Resources Information Center

    Lott, Debra

    2008-01-01

    This article describes how contemporary costumes, expressive techniques, and mixed media can take "the ordinary" out of figure studies. To pique student interest and create a meaningful figurative study, students are instructed to bring in their latest fashion accessories (hats, shawls, neck warmers, denim jackets, etc.), or shop the local thrift…

  4. Poetic expressions of vigilance.

    PubMed

    Carr, Jeanine M

    2003-11-01

    In this article, the author explores poetic transcription as an experimental form of writing. Her previous ethnographic research, in which she explored the experience of vigilance for family members who stayed at the bedside of hospitalized relatives, provided the qualitative data for the poetic compositions. In this article, she describes the background and rationale for the research, the findings, and her use of the data to transcribe them into poetic expressions of the lived experience of vigilance.

  5. Nucleotide Sequence of the Hantaan Virus S RNA Segment and Expression of Encoded Proteins

    DTIC Science & Technology

    1987-11-03

    stomatitis virus and influenza virus) has been shown to be important in generating cytotoxic T cells in response to viral infection (Townsend et al...termination-polyadenylation signal common to other negative-strand RNA viruses (Sendai virus: 5’-TAAGAAAA and vesicular stomatitis virus: TATGAAAA...codon also occurs in the two retroviruses, murine leukemia virus and feline leukemia virus, to produce a precursor polyprotein larger than normal 191

  6. Expression Profiling of Cell Lines Expressing Regulated NP2 Transcripts

    DTIC Science & Technology

    2004-09-01

    EGF in the presence or absence of exogenous HRS . The results will provide a framework fo r the interpretation of future gene expression studies in...e studies require further verification. Small sam- ple size, tissue heterogeneity, and inter-indivi- dual variations among human patients may result ... studies we proposed using gene expression profiling to determine change s in gene expression as a function of expression of the neurofibromatosis-2 (NF2

  7. Decoding Children's Expressions of Affect.

    ERIC Educational Resources Information Center

    Feinman, Joel A.; Feldman, Robert S.

    1982-01-01

    Mothers' ability to decode their children's nonverbal expressions of four affects (happiness, sadness, fear, and anger) was contrasted with the decoding ability of a matched group of nonmothers. Results indicate that mothers were accurately able to decode expressions of happiness but had relative difficulty with decoding expressions of sadness,…

  8. Expression of the v-mos gene alters a Mr 55,000 protein during acute infection by Moloney murine sarcoma virus.

    PubMed Central

    Singh, B; Sparrow, J T; Hedge, A M; Arlinghaus, R B

    1986-01-01

    Infection of the rat myoblast cell line, L6E9, with Moloney murine sarcoma virus (Mo-MuSV) clone 124, altered a cellular protein of Mr 55,000 (P55) within 2 days of infection. The alteration of P55 was observed as a reduction in its steady-state level in cell extracts. The reduction of P55 correlated with the appearance of p37mos in infected cells. Except for P55 and one other protein, no change was detected in the total protein pattern of infected cells compared to uninfected cells, as judged by either immunoblots of one-dimensional NaDodSO4 gels or direct two-dimensional gel analysis. P55 levels were unchanged when L6E9 cells were infected with Moloney murine leukemia virus or several different transforming retroviruses. To determine the specificity of this v-mos-induced effect on P55, L6E9 cells were acutely infected with a temperature-sensitive variant (ts110) of Mo-MuSV. When these cells were shifted from 39 degrees C to 33 degrees C, which activates the gag-mos gene product, the P55 level dropped by greater than 50% within 2-3 hr. Conversely, with a shift in temperature from 33 degrees C to 39 degrees C, the cells' P55 level returned to normal within 5 hr, starting at 30 min after shift. These results clearly show that v-mos expression in acutely infected L6E9 cells alters the cellular protein, P55. Images PMID:3012522

  9. Prion Strain Differences in Accumulation of PrPSc on Neurons and Glia Are Associated with Similar Expression Profiles of Neuroinflammatory Genes: Comparison of Three Prion Strains

    PubMed Central

    Carroll, James A.; Striebel, James F.; Rangel, Alejandra; Woods, Tyson; Phillips, Katie; Peterson, Karin E.; Race, Brent; Chesebro, Bruce

    2016-01-01

    Misfolding and aggregation of host proteins are important features of the pathogenesis of neurodegenerative diseases including Alzheimer’s disease, Parkinson’s disease, frontotemporal dementia and prion diseases. In all these diseases, the misfolded protein increases in amount by a mechanism involving seeded polymerization. In prion diseases, host prion protein is misfolded to form a pathogenic protease-resistant form, PrPSc, which accumulates in neurons, astroglia and microglia in the CNS. Here using dual-staining immunohistochemistry, we compared the cell specificity of PrPSc accumulation at early preclinical times post-infection using three mouse scrapie strains that differ in brain regional pathology. PrPSc from each strain had a different pattern of cell specificity. Strain 22L was mainly associated with astroglia, whereas strain ME7 was mainly associated with neurons and neuropil. In thalamus and cortex, strain RML was similar to 22L, but in substantia nigra, RML was similar to ME7. Expression of 90 genes involved in neuroinflammation was studied quantitatively using mRNA from thalamus at preclinical times. Surprisingly, despite the cellular differences in PrPSc accumulation, the pattern of upregulated genes was similar for all three strains, and the small differences observed correlated with variations in the early disease tempo. Gene upregulation correlated with activation of both astroglia and microglia detected in early disease prior to vacuolar pathology or clinical signs. Interestingly, the profile of upregulated genes in scrapie differed markedly from that seen in two acute viral CNS diseases (LaCrosse virus and BE polytropic Friend retrovirus) that had reactive gliosis at levels similar to our prion-infected mice. PMID:27046083

  10. Slug Expression during Melanoma Progression

    PubMed Central

    Shirley, Stephanie H.; Greene, Victoria R.; Duncan, Lyn M.; Torres Cabala, Carlos A.; Grimm, Elizabeth A.; Kusewitt, Donna F.

    2012-01-01

    Slug (Snai2), a member of the Snail family of zinc finger transcription factors, plays a role in the epithelial-to-mesenchymal transformation (EMT) that occurs during melanocyte emigration from the neural crest. A role for Slug in the EMT-like loss of cell adhesion and increased cell motility exhibited during melanoma progression has also been proposed. Our immunohistochemical studies of melanoma arrays, however, revealed that Slug expression was actually higher in nevi than in primary or metastatic melanomas. Moreover, Slug expression in melanomas was not associated with decreased expression of E-cadherin, the canonical Slug target in EMT. Comparisons of endogenous Slug and E-cadherin expression in cultured normal human melanocytes and melanoma cell lines supported our immunohistochemical findings. Expression of exogenous Slug in melanocytes and melanoma cells in vitro, however, suppressed E-cadherin expression, enhanced N-cadherin expression, and stimulated cell migration and invasion. Interestingly, both in tumors and cultured cell lines, there was a clear relationship between expression of Slug and MITF, a transcription factor known to regulate Slug expression during development. Taken together, our findings suggest that Slug expression during melanomagenesis is highest early in the process and that persistent Slug expression is not required for melanoma progression. The precise role of Slug in melanomagenesis remains to be elucidated and may be related to its interactions with other drivers of EMT, such as Snail. PMID:22503751

  11. Expressing fear enhances sensory acquisition.

    PubMed

    Susskind, Joshua M; Lee, Daniel H; Cusi, Andrée; Feiman, Roman; Grabski, Wojtek; Anderson, Adam K

    2008-07-01

    It has been proposed that facial expression production originates in sensory regulation. Here we demonstrate that facial expressions of fear are configured to enhance sensory acquisition. A statistical model of expression appearance revealed that fear and disgust expressions have opposite shape and surface reflectance features. We hypothesized that this reflects a fundamental antagonism serving to augment versus diminish sensory exposure. In keeping with this hypothesis, when subjects posed expressions of fear, they had a subjectively larger visual field, faster eye movements during target localization and an increase in nasal volume and air velocity during inspiration. The opposite pattern was found for disgust. Fear may therefore work to enhance perception, whereas disgust dampens it. These convergent results provide support for the Darwinian hypothesis that facial expressions are not arbitrary configurations for social communication, but rather, expressions may have originated in altering the sensory interface with the physical world.

  12. Maximally Expressive Modeling

    NASA Technical Reports Server (NTRS)

    Jaap, John; Davis, Elizabeth; Richardson, Lea

    2004-01-01

    Planning and scheduling systems organize tasks into a timeline or schedule. Tasks are logically grouped into containers called models. Models are a collection of related tasks, along with their dependencies and requirements, that when met will produce the desired result. One challenging domain for a planning and scheduling system is the operation of on-board experiments for the International Space Station. In these experiments, the equipment used is among the most complex hardware ever developed; the information sought is at the cutting edge of scientific endeavor; and the procedures are intricate and exacting. Scheduling is made more difficult by a scarcity of station resources. The models to be fed into the scheduler must describe both the complexity of the experiments and procedures (to ensure a valid schedule) and the flexibilities of the procedures and the equipment (to effectively utilize available resources). Clearly, scheduling International Space Station experiment operations calls for a maximally expressive modeling schema.

  13. Maximally Expressive Task Modeling

    NASA Technical Reports Server (NTRS)

    Japp, John; Davis, Elizabeth; Maxwell, Theresa G. (Technical Monitor)

    2002-01-01

    Planning and scheduling systems organize "tasks" into a timeline or schedule. The tasks are defined within the scheduling system in logical containers called models. The dictionary might define a model of this type as "a system of things and relations satisfying a set of rules that, when applied to the things and relations, produce certainty about the tasks that are being modeled." One challenging domain for a planning and scheduling system is the operation of on-board experiment activities for the Space Station. The equipment used in these experiments is some of the most complex hardware ever developed by mankind, the information sought by these experiments is at the cutting edge of scientific endeavor, and the procedures for executing the experiments are intricate and exacting. Scheduling is made more difficult by a scarcity of space station resources. The models to be fed into the scheduler must describe both the complexity of the experiments and procedures (to ensure a valid schedule) and the flexibilities of the procedures and the equipment (to effectively utilize available resources). Clearly, scheduling space station experiment operations calls for a "maximally expressive" modeling schema. Modeling even the simplest of activities cannot be automated; no sensor can be attached to a piece of equipment that can discern how to use that piece of equipment; no camera can quantify how to operate a piece of equipment. Modeling is a human enterprise-both an art and a science. The modeling schema should allow the models to flow from the keyboard of the user as easily as works of literature flowed from the pen of Shakespeare. The Ground Systems Department at the Marshall Space Flight Center has embarked on an effort to develop a new scheduling engine that is highlighted by a maximally expressive modeling schema. This schema, presented in this paper, is a synergy of technological advances and domain-specific innovations.

  14. A Comparison of Artificial Subtle Expressions with Human-like Expressions on Expressing Confidence Level

    NASA Astrophysics Data System (ADS)

    Komatsu, Takanori; Kobayashi, Kazuki; Yamada, Seiji; Funakoshi, Kotaro; Nakano, Mikio

    Expressing the confidence level of a system's suggestions by using speech sounds is an important cue to users of the system for perceiving how likely it is for the suggestions to be correct. We assume that expressing confidence levels by using human-like expressions would cause users to have a poorer impression of the systems than if artificial subtle expressions (ASEs) were used when the quality of the presented information does not match the expressed confidence level. We confirmed that this assumption was correct by conducting a psychological experiment.

  15. Understanding tissue expression evolution: from expression phylogeny to phylogenetic network.

    PubMed

    Gu, Xun

    2016-03-01

    Our understanding of tissue expression evolution in multi-cellular model organisms has been considerably advanced with the help of high-throughput technologies from EST, microarray to RNA-seq. Yet, many controversies remained unsolved, ranging from the evolutionary patterns of tissue expressions to expression phylogenetic analysis. Moreover, despite numerous reports published, it is desirable to have a general framework for study of tissue expression evolution. In this article, we first provide an up-to-date and concise review for the study of tissue expression evolution in multi-cellular organisms. While the expression phylogeny of the same tissues sampled from closely or intermediately related species largely reflects the species phylogeny, we demonstrate that phylogenetic network approach may shed some lights for our understanding of the developmental similarity and evolutionary relatedness during the multi-tissue evolution.

  16. Individual and Maturational Differences in Infant Expressivity.

    ERIC Educational Resources Information Center

    Field, Tiffany

    1989-01-01

    Reports that, even though young infants can discriminate among different facial expressions, there are individual differences in infants' expressivity and ability to produce and discriminate facial expressions. (PCB)

  17. Automated Learning of Temporal Expressions.

    PubMed

    Redd, Douglas; Shaoa, YiJun; Yang, Jing; Divita, Guy; Zeng-Treitler, Qing

    2015-01-01

    Clinical notes contain important temporal information that are critical for making clinical diagnosis and treatment as well as for retrospective analyses. Manually created regular expressions are commonly used for the extraction of temporal information; however, this can be a time consuming and brittle approach. We describe a novel algorithm for automatic learning of regular expressions in recognizing temporal expressions. Five classes of temporal expressions are identified. Keywords specific to those classes are used to retrieve snippets of text representing the same keywords in context. Those snippets are used for Regular Expression Discovery Extraction (REDEx). These learned regular expressions are then evaluated using 10-fold cross validation. Precision and recall are very high, above 0.95 for most classes.

  18. Differential Expression Analysis for Pathways

    PubMed Central

    Haynes, Winston A.; Higdon, Roger; Stanberry, Larissa; Collins, Dwayne; Kolker, Eugene

    2013-01-01

    Life science technologies generate a deluge of data that hold the keys to unlocking the secrets of important biological functions and disease mechanisms. We present DEAP, Differential Expression Analysis for Pathways, which capitalizes on information about biological pathways to identify important regulator