DOCSLIB.ORG

High Transdominant Revm10 Protein Levels Are Required to Inhibit HIV-1 Replication in Cell Lines and Primary T Cells: Implication for Gene Therapy of AIDS

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
High Transdominant Revm10 Protein Levels Are Required to Inhibit HIV-1 Replication in Cell Lines and Primary T Cells: Implication for Gene Therapy of AIDS Gene Therapy (1997) 4, 128–139 1997 Stockton Press All rights reserved 0969-7128/97 $12.00 High transdominant RevM10 protein levels are required to inhibit HIV-1 replication in cell lines and primary T cells: implication for gene therapy of AIDS I Plavec1, M Agarwal1,KEHo2, M Pineda1, J Auten1, J Baker1, H Matsuzaki3, S Escaich4, M Bonyhadi1 and E Bo¨ hnlein1 1Progenesys Program, SyStemix Inc, 3155 Porter Drive, Palo Alto, CA 94304, USA Expression of antiviral genes in CD4+ T cells has been pro- uniformly higher than from internal promoters (eg CMV, posed as a strategy for gene therapy of AIDS. Over the PGK). Analysis of selected vectors in acutely and chron- past years, we and others have developed retroviral vec- ically HIV-infected cell lines suggested that threshold levels tors encoding the RevM10 protein, a dominant-negative of RevM10 expression are required to achieve inhibition of mutant of the HIV-1 Rev trans-activator protein. We could HIV replication. LTR-driven RevM10 expression also demonstrate gene transfer and inhibition of HIV-1 repli- yielded high steady-state protein levels in activated primary cation in cultured T cell lines and primary T cells. However, T cells resulting in inhibition of HIV replication, and there little is known about the levels of the antiviral protein was no apparent difference between the MoMLV, MPSV required to achieve a therapeutic effect, particularly in pri- and MESV-LTR vectors. However, RevM10 expression mary cells. In this report, we compare different vector was down-regulated in resting primary cells and conse- designs with regard to expression of the antiviral gene to quently anti-HIV efficacy was significantly reduced. Taken develop an optimal vector for clinical applications. Our together, the data suggest that relatively high steady-state results demonstrate that intracellular steady-state RevM10 levels of RevM10 protein are required to achieve inhibition protein levels expressed from the Moloney murine leuke- of HIV replication and that the MPSV- and MESV-derived mia virus (MoMLV), myeloproliferative sarcoma virus retroviral vectors show no advantage over the MoMLV- (MPSV) or mouse embryonic stem cell virus (MESV) pro- based vectors for expression of anti-HIV genes in human moters located in the long terminal repeat (LTR) were T cells. Keywords: gene therapy; HIV-1; retroviral vectors; transgene expression; T cells Introduction of Rev function.9 Furthermore, a number of studies have demonstrated that RevM10 can inhibit viral replication in Human immunodeficiency virus type-1 (HIV-1) encodes HIV-infected T cell lines and primary T cells.10–15 several regulatory genes including Rev (reviewed in Refs Intracellular expression of genes which can suppress 1–3). Through binding to a structured RNA sequence, HIV-1 replication has been proposed as an alternative to termed the Rev-responsive element (RRE), Rev promotes standard therapies for the treatment of AIDS.16,17 A large 4,5 the export of RRE-containing mRNAs from the nucleus number of antiviral genes including RNA decoys, anti- leading to cytoplasmic accumulation of incompletely sense RNA, transdominant mutants (including RevM10), spliced HIV-1 transcripts. First, these transcripts serve as ribozymes and intracellular antibodies (reviewed in Ref. templates for translation of viral proteins gag, pol and env. 16) have been shown to inhibit effectively HIV-1 in vitro. Second, the nonspliced genomic mRNA is needed for The target cell population for delivery of anti-HIV genes infectious particle formation. HIV-1 Rev function requires could be peripheral blood mononuclear cells,18,19 or alter- 6–8 interaction with one or more cellular host proteins. natively hematopoietic stem cells.16 Retroviral vectors are RevM10, a mutant of Rev retains the ability to bind to presently the most widely used system to deliver genes RRE but it does not transactivate, possibly because of its into those cells.20 Irrespective of the target cell popu- 8,9 inability to interact with host factor(s). When co- lation, the success of gene therapy will depend on both expressed with wild-type Rev protein in transient trans- efficient gene transfer and persistent high-level gene fection assays, RevM10 acts as a transdominant inhibitor expression. Initially, efforts in developing retroviral vec- tors were mainly focused on improving virus titers to increase gene transfer and only recently more work is Correspondence: E Bo¨hnlein 2 being invested in optimizing vectors for efficient trans- Present addresses: Genentech Inc, 460 Point San Bruno Boulevard, South gene expression (reviewed in Refs 21–23). A wide variety San Francisco, California 94080-4990, USA; 3Affymetrix, 3380 Central Expressway, Santa Clara, California 95051, USA; 4Rhoˆne-Poulenc Rorer, of vector constructions have been described including 94403 Vitry-sur-Seine, France different promoters (retroviral LTRs, internal promoters), Received 15 August 1996; accepted 29 October 1996 orientation of the gene of interest (forward, reverse), self- RevM10 expression in cell lines and primary T cells I Plavec et al 129 inactivating LTR vectors and other LTR modifications, driven transcript (Figure 2, lower left panel) resulting in and insertion of an expression cassette into the LTR RevM10 protein levels approximately 10-fold higher than (double-copy vectors) (reviewed in Refs 21, 23). Despite internal promoter vectors (Figure 3). No apparent differ- all these studies, the optimal vector design for the ence between the MoMLV-LTR and MPSV-LTR vectors expression of a particular gene needs to be determined could be detected in CEMSS cells and the different empirically and the final result may depend on the com- primer binding sites also had no measurable impact. To bination of both gene and vector sequences.24–26 further improve expression of the LTR-RevM10 vectors, As part of our efforts to develop clinical protocols for the tk-Neo sequence was replaced by the SV40-Puro tran- gene therapy of AIDS we have tested a number of retro- scription unit (LMSPL vector; Figure 1). This modifi- viral vector designs to optimally express the transdomi- cation resulted in a two- to three-fold increase in RevM10 nant RevM10 protein. In particular, we carefully ana- protein levels in CEMSS cells (compare LMTNL with lyzed vector expression in primary T cells at various LMSPL, Figure 3). stages of activation. We also evaluated the anti-HIV In all vectors (Figure 1; numbers 4–8), the packaging potency of selected vectors in chronically and acutely signal is encoded by a short retroviral sequence13,36 and HIV-infected cell lines as well as in primary T cells. The the RevM10 protein is synthesized from an unspliced results suggest that the extent of inhibition of HIV-1 repli- full-length messenger RNA. The presence of an intron in cation is a function of the relative expression of RevM10 a transcription unit can improve the expression of some protein in those cells, supporting the hypothesis that cDNAs.37 To test this possibility, we constructed RevM10 RevM10 acts as an intracellular competitive inhibitor.9 vectors based on the LXSN38 (construct LMSPL-INT1; Implications of these findings for the development of Figure 1, No. 9) and MFG design39 (construct LMSPL- effective retrovirus vector-based anti-HIV gene therapies INT2; Figure 1, No. 10) in which protein expression relies are discussed. on the efficient splicing of the primary transcript.39,40 The ratio of spliced to unspliced transcripts was higher for the LMSPL-INT2 vector compared to the LMSPL-INT1 Results (Figure 2, lower right panel). This finding was not sur- prising since the splicing in the MFG vector had been Retroviral vector LTR-driven expression yields high optimized for protein expression.39 However, both vec- steady-state levels of RevM10 protein in transduced tors yielded similar steady-state intracellular RevM10 T cells protein levels and these were not significantly higher We started our study by testing vectors in which RevM10 when compared to the intronless LMSPL vector (Figure expression is driven by an internal promoter using the 2). Thus, inclusion of an intron did not further improve immediate–early promoter of the human cytomegalo- RevM10 protein expression. An unexpected short Rev- virus27 (LNCML vector; Figure 1) and the mouse phos- specific transcript was detected in the LMSPL RNA phoglycerate kinase gene promoter28 (LNMPL and (Figure 2). This RNA is a splice product between the LPMPL vectors; Figure 1). Human CEMSS T lymphoid splice donor in the retroviral packaging sequence and a cells were transduced with amphotropic retroviral vec- splice acceptor located in the RevM10 coding sequence tors harvested from PA317-based producer cells and and does not encode functional RevM10 protein (data not selected either with G418 or puromycin to yield homo- shown). As expected, a 10-fold higher virus titer was genous populations of transduced cells. RevM10 protein obtained with the LMSPL-INT1 vector because of the and RNA expression were analyzed by Rev-specific extended packaging sequence38 (Figure 1). immunofluorescence29 and Northern blotting, respect- ively. Regardless of the internal promoter used, low RevM10 protein threshold level is required for inhibition RevM10 RNA expression levels were observed in trans- of HIV-1 replication in chronically infected T cell lines duced cells (Figure 2) which resulted in intracellular We have used an assay system based on chronically HIV- RevM10 protein levels barely detectable above back- infected A3.01/ProHygro T cells to compare the anti-HIV ground (Figure 3). Since it has been reported that the neo- efficacy of the different vectors expressing various levels mycin phosphotransferase gene (Neo) gene sequence can of the RevM10 protein.13 The A3.01/ProHygro cells were act as a transcriptional silencer,30 we attempted to transduced by co-cultivation with the PA317-derived improve RevM10 expression by replacing the Neo- retroviral vector producer cells followed by selection in resistance gene with the puromycin-resistance gene G418 and hygromycin.
Load more

Recommended publications
  • The Zinc-Finger Antiviral Protein Recruits the RNA Processing Exosome to Degrade the Target Mrna
    The zinc-finger antiviral protein recruits the RNA processing exosome to degrade the target mRNA Xuemin Guo, Jing Ma, Jing Sun, and Guangxia Gao* Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China Edited by John M. Coffin, Tufts University School of Medicine, Boston, MA, and approved November 3, 2006 (received for review August 14, 2006) Zinc-finger antiviral protein (ZAP) is a host antiviral factor that putative RNA helicase (Kiaa0052), and a protein that is specifically specifically inhibits the replication of Moloney murine leukemia phosphorylated in the M phase of the cell cycle (Mpp6) (36, 38). virus (MLV) and Sindbis virus (SIN) by preventing accumulation of The RNase-PH domain subunits and the S1/KH RNA-binding the viral mRNA in the cytoplasm. In previous studies, we demon- domain subunits are considered to be the core components of the strated that ZAP directly binds to its specific target mRNAs. In this exosome, whereas Kiaa0052 and Mpp6 are considered to be article, we provide evidence indicating that ZAP recruits the RNA accessory factors (36, 38). Yeast PM/Scl-100 is found only in the processing exosome to degrade the target RNA. ZAP comigrated nuclear exosome (36, 46). with the exosome in sucrose or glycerol velocity gradient centrif- The structure of the exosome is not yet determined. Based on the ugation. Immunoprecipitation of ZAP coprecipitated the exosome results of mammalian-two-hybrid and yeast-two-hybrid experi- components. In vitro pull-down assays indicated that ZAP directly ments (38, 56–59), the six RNase-PH domain-containing subunits interacted with the exosome component hRrp46p and that the are thought to assemble into a doughnut-shaped ring.
    [Show full text]
  • Extracellular Localization of Pokeweed Antiviral Protein MICHAEL P
    Proc. Natl. Acad. Sci. USA Vol. 83, pp. 5053-5056, July 1986 Biochemistry Extracellular localization of pokeweed antiviral protein MICHAEL P. READY*, DENNIS T. BROWNt, AND JON D. ROBERTUS** *Clayton Foundation Biochemical Institute, Department of Chemistry, and tCell Research Institute and Department of Microbiology, University of Texas, Austin, TX 78712 Communicated by Esmond E. Snell, March 24, 1986 ABSTRACT Pokeweed antiviral protein is an enzyme of molecule may well be inactive until it is processed and Mr 29,000 known to inactivate a wide variety of eukaryotic packaged in the seed. ribosomes. We have used electron microscopy to show that the However, the case is not so clear for proteins such as the antibody specific for the protein is bound within the cell wall pokeweed enzyme, which are not cytotoxic to animals. In matrix of leaf mesophyll cells from Phytolacca americana. Any addition, reports have suggested that pokeweed antiviral penetration or breakage of the cell wall and membrane could protein does not inhibit protein synthesis on pokeweed allow the enzyme to enter the cytoplasm, where it is likely to ribosomes (19, 20). If this were true, it would mean that inhibit protein synthesis in the damaged cell. We speculate that pokeweed could not shut down its own ribosomes if they pokeweed antiviral protein is a defensive agent whose principal were usurped by an invading virus. Recently we speculated 'function is probably antiviral. (5) that this state of affairs is unlikely; a protein that makes up as much as 0.5% of the plant's soluble protein and that Many higher plants contain proteins that enzymatically attacks ribosomes with a Kcat of 400 mol/mol per min must inhibit protein synthesis on eukaryotic ribosomes.
    [Show full text]
  • Interferon-Inducible Antiviral Effectors
    REVIEWS Interferon-inducible antiviral effectors Anthony J. Sadler and Bryan R. G. Williams Abstract | Since the discovery of interferons (IFNs), considerable progress has been made in describing the nature of the cytokines themselves, the signalling components that direct the cell response and their antiviral activities. Gene targeting studies have distinguished four main effector pathways of the IFN-mediated antiviral response: the Mx GTPase pathway, the 2′,5′-oligoadenylate-synthetase-directed ribonuclease L pathway, the protein kinase R pathway and the ISG15 ubiquitin-like pathway. As discussed in this Review, these effector pathways individually block viral transcription, degrade viral RNA, inhibit translation and modify protein function to control all steps of viral replication. Ongoing research continues to expose additional activities for these effector proteins and has revealed unanticipated functions of the antiviral response. Pattern-recognition Interferon (IFN) was discovered more than 50 years ago in components of the IFNR signalling pathway (STAT1 receptors as an agent that inhibited the replication of influenza (signal transducer and activator of transcription 1), TYK2 (PRRs). Host receptors that can virus1. The IFN family of cytokines is now recognized as (tyrosine kinase 2) or UNC93B) die of viral disease, with sense pathogen-associated a key component of the innate immune response and the the defect in IFNAR (rather than IFNGR) signalling molecular patterns and initiate 6–9 signalling cascades that lead to first line of defence against viral infection. Accordingly, having the more significant role . an innate immune response. IFNs are currently used therapeutically, with the most The binding of type I IFNs to the IFNAR initiates a These can be membrane bound noteworthy example being the treatment of hepatitis C signalling cascade, which leads to the induction of more (such as Toll-like receptors) or virus (HCV) infection, and they are also used against than 300 IFN-stimulated genes (ISGs)10.
    [Show full text]
  • Since January 2020 Elsevier Has Created a COVID-19 Resource Centre with Free Information in English and Mandarin on the Novel Coronavirus COVID- 19
    Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID- 19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active. Review To translate, or not to translate: viral and host mRNA regulation by interferon-stimulated genes Melody M.H. Li, Margaret R. MacDonald, and Charles M. Rice Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY, USA 0 0 Type I interferon (IFN) is one of the first lines of cellular and 2 -5 -oligoadenylate synthetase (OAS)/RNAseL func- defense against viral pathogens. As a result of IFN sig- tion to block translation to limit virus replication (Box 1). naling, a wide array of IFN-stimulated gene (ISG) pro- This review focuses on the more recently described ISGs ducts is upregulated to target different stages of the viral that regulate host or viral translation, localize to transla- life cycle. We review recent findings implicating a subset tionally silent granules, and interfere with miRNA-medi- of ISGs in translational regulation of viral and host ated silencing of host transcripts.
    [Show full text]
  • The Antiviral Protein Viperin Enhances the Host Interferon
    bioRxiv preprint doi: https://doi.org/10.1101/493098; this version posted December 11, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The Antiviral Protein Viperin Enhances The Host Interferon Response Following STING Activation Keaton Crosse1, Ebony Monson1, Monique Smith1, Yeu-Yang Tseng2, Kylie Van der Hoek3, Peter Revill4, David Tscharke2, Michael Beard3, Karla Helbig1 1 Department of Physiology, Anatomy and Microbiology, La Trobe University, Bundoora, VIC, Australia 2 John Curtin School of Medical Research, The Australian National University, Canberra, ACT, Australia 3 School of Biological Sciences, The University of Adelaide, Adelaide, SA, Australia 4 Victorian Infectious Diseases Reference Laboratory, Royal Melbourne Hospital, Peter Doherty Institute for Infection and Immunity, Melbourne, VIC, Australia Correspondence to be addressed to: Dr Karla Helbig Department of Physiology, Anatomy and Microbiology, La Trobe University 1 Kingsbury Drive, Bundoora, Vic, 3083 Email: [email protected] Key words: viperin, interferon stimulated genes, interferon, innate immunity, virus, STING, TBK1, dsDNA 1 bioRxiv preprint doi: https://doi.org/10.1101/493098; this version posted December 11, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Abstract Viperin is an interferon-inducible protein that is critical for eliciting an effective immune response against many diverse viral pathogens. As such, viperin has been implicated in interactions with many functionally unrelated host and viral proteins, making it increasingly difficult to determine a unifying mechanism of viperin’s antiviral activity.
    [Show full text]
  • Expression of Pokeweed Antiviral Protein in Transgenic Plants
    Plant Physiol. (1997) 114: 1113-1 121 Expression of Pokeweed Antiviral Protein in Transgenic Plants lnduces Virus Resistance in Grafted Wild-Type Plants lndependently of Salicylic Acid Accumulation and Pathogenesis-Related Protein Synthesis’ Sergey Smirnov, Vladimir Shulaev, and Nilgun E. Tumer* Center for Agricultura1 Molecular Biology and Department of Plant Pathology, Rutgers University, P.O. Box 231, New Brunswick, New Jersey 08903-0231 pokeweed (Phytolacca americana) plants. These proteins are Pokeweed antiviral protein (PAP), a 29-kD protein isolated from similar in molecular mass (29, 30, and 29.5 kD, respec- Phytolacca americana, inhibits translation by catalytically removing tively) but are expressed at different developmental stages a specific adenine residue from the large rRNA of the 60s subunit of and in different tissues of pokeweed (Wyatt and Shepherd, eukaryotic ribosomes. Transgenic tobacco (Nicofiana fabacum) 1969; Irvin et al., 1980; Barbieri et al., 1982). PAP depuri- plants expressing PAP or a variant (PAP-V) were shown to be resis- nates ribosomes from pokeweed and other plants (Bonness tant to a broad spectrum of plant viruses. Expression of PAP-v in et al., 19941, as well as mammalian, yeast, and bacterial transgenic plants induces synthesis of pathogenesis-related proteins ribosomes. In addition, PAI’ effectively inhibits infection and a very weak (~2-fold)increase in salicylic acid levels. Using by a number of different plant (Wyatt and Shepherd, 1969; reciproca1 grafting experiments, we demonstrate here that trans- Tomlinson et al., 1974; Chen et al., 1992) and animal viruses genic tobacco rootstocks expressing PAP-v induce resistance to (Tomlinson et al., 1974; Ussery et al., 1977), including hu- tobacco mosaic virus infection in both N.
    [Show full text]
  • CG Dinucleotide Suppression Enables Antiviral Defence Targeting Non-Self RNA Matthew A
    LETTER doi:10.1038/nature24039 CG dinucleotide suppression enables antiviral defence targeting non-self RNA Matthew A. Takata1, Daniel Gonçalves-Carneiro1, Trinity M. Zang1,2, Steven J. Soll1,2, Ashley York1, Daniel Blanco-Melo1 & Paul D. Bieniasz1,2 Vertebrate genomes exhibit marked CG suppression—that is, lower (Fig. 1f). Our synonymous mutagenesis coincidentally increased than expected numbers of 5′-CG-3′ dinucleotides1. This feature is the CG dinucleotide content in mutant segments to a level similar likely to be due to C-to-T mutations that have accumulated over to that of random sequence (Fig. 1f). We generated derivatives of hundreds of millions of years, driven by CG-specific DNA methyl mutant L, termed LCG and LOTH, respectively, containing only transferases and spontaneous methyl-cytosine deamination. Many mutations that generated new CG dinucleotides (37 of 145 original RNA viruses of vertebrates that are not substrates for DNA methyl mutations) or the 108 other mutations (Supplementary Data 1). transferases mimic the CG suppression of their hosts2–4. This We also generated mutants that maximized the CG or, as a further 4–6 property of viral genomes is unexplained . Here we show, using control, GC dinucleotide content in the same segment (LCG-HI and synonymous mutagenesis, that CG suppression is essential for LGC-HI) (Extended Data Table 1). These proviral plasmids each yielded HIV-1 replication. The deleterious effect of CG dinucleotides on similar levels of infectious virus following transfection of 293T cells HIV-1 replication was cumulative, associated with cytoplasmic RNA depletion, and was exerted by CG dinucleotides in both translated a GFP and non-translated exonic RNA sequences.
    [Show full text]
  • Prokaryotic Viperins Produce Diverse Antiviral Molecules
    Article Prokaryotic viperins produce diverse antiviral molecules https://doi.org/10.1038/s41586-020-2762-2 Aude Bernheim1, Adi Millman1, Gal Ofir1, Gilad Meitav1, Carmel Avraham1, Helena Shomar2, Masha M. Rosenberg2, Nir Tal2, Sarah Melamed1, Gil Amitai1 & Rotem Sorek1 ✉ Received: 25 February 2020 Accepted: 27 August 2020 Viperin is an interferon-induced cellular protein that is conserved in animals1. It Published online: xx xx xxxx has previously been shown to inhibit the replication of multiple viruses by producing Check for updates the ribonucleotide 3′-deoxy-3′,4′-didehydro (ddh)-cytidine triphosphate (ddhCTP), which acts as a chain terminator for viral RNA polymerase2. Here we show that eukaryotic viperin originated from a clade of bacterial and archaeal proteins that protect against phage infection. Prokaryotic viperins produce a set of modifed ribonucleotides that include ddhCTP, ddh-guanosine triphosphate (ddhGTP) and ddh-uridine triphosphate (ddhUTP). We further show that prokaryotic viperins protect against T7 phage infection by inhibiting viral polymerase-dependent transcription, suggesting that it has an antiviral mechanism of action similar to that of animal viperin. Our results reveal a class of potential natural antiviral compounds produced by bacterial immune systems. Viperin is an antiviral protein that becomes highly expressed in cells genes involved in defence systems is a strong predictor that the genes in stimulated by interferons1. In humans, this protein has broad antiviral the cluster have a role in phage resistance8,9. We denoted the genes in the activity against DNA and RNA viruses, including human cytomegalovi- defensive cluster pVips (for prokaryotic viperin homologues). As pVips rus, West Nile virus, dengue virus, hepatitis C virus and HIV1,3.
    [Show full text]
  • Discovery of Another Anti-HIV Protein in the Search for the CD8+ Cell Anti-HIV Factor
    COMMENTARY COMMENTARY Discovery of another anti-HIV protein in the search for the CD8+ cell anti-HIV Factor Jay A. Levy1 purified cytokines, no human proteins linked Department of Medicine, University of California, San Francisco, CA 94143 to CD8+ cells have been found with the characteristics of CAF (11, 16). In PNAS, the research group led by Ian de + Nearly three decades ago, soon after the to the infected CD4 cells inhibited virus Belle reports the surprising finding that a recognition of AIDS and its causative agent, replication without affecting the viability of nuclear protein, TOE1 (target of EGR1, early + HIV, the first immune response against the the CD4 cells (1). growth response 1), could be CAF (17). TOE1 AIDS virus was identified: an unexpected This lack of cell killing was further sub- is found primarily in nucleoli and Cajal bod- + + anti-HIV activity of CD8 cells that did not stantiated when the CD8 cells were sepa- ies and is involved in a variety of intracellular + involve cell killing (1). This immune response rated by a filter from the HIV-infected CD4 events, including deadenylase activity and + differed from the classic CD8 cell antiviral cells. Virus replication was inhibited, and a spliceosome assembly (18, 19). TOE1 was ini- activity in which cytotoxic T lymphocytes kill soluble protease-sensitive factor, now known tially cloned by the de Belle laboratory and + + virus-infected cells (2). The CD8 cell non- as the CD8 cell antiviral factor (CAF), ap- found to bind to the p53 tumor suppressor – cytotoxic anti-HIV response (CNAR) became peared responsible (6 8).
    [Show full text]
  • The Short Form of the Zinc Finger Antiviral Protein Inhibits Influenza A
    VIRUS-CELL INTERACTIONS crossm The Short Form of the Zinc Finger Antiviral Protein Inhibits Influenza A Downloaded from Virus Protein Expression and Is Antagonized by the Virus-Encoded NS1 Qiannan Tang,a,b Xinlu Wang,a Guangxia Gao,a,b CAS Key Laboratory of Infection and Immunity, Institute of Biophysics, Chinese Academy of Sciences, Beijing, http://jvi.asm.org/ Chinaa; University of Chinese Academy of Sciences, Beijing, Chinab ABSTRACT Zinc finger antiviral protein (ZAP) is a host factor that specifically inhibits the replication of certain viruses. There are two ZAP isoforms arising from alternative Received 30 September 2016 Accepted 27 splicing, which differ only at the C termini. It was recently reported that the long October 2016 Accepted manuscript posted online 2 isoform (ZAPL) promotes proteasomal degradation of influenza A virus (IAV) proteins November 2016 PA and PB2 through the C-terminal poly(ADP-ribose) polymerase (PARP) domain, Citation Tang Q, Wang X, Gao G. 2017. The which is missing in the short form (ZAPS), and that this antiviral activity is antago- short form of the zinc finger antiviral protein on April 20, 2017 by NATIONAL SCIENCE LIBRARY, CAS nized by the viral protein PB1. Here, we report that ZAP inhibits IAV protein expres- inhibits influenza A virus protein expression and is antagonized by the virus-encoded NS1. sion in a PARP domain-independent manner. Overexpression of ZAPS inhibited the J Virol 91:e01909-16. https://doi.org/10.1128/ expression of PA, PB2, and neuraminidase (NA), and downregulation of the endoge- JVI.01909-16. nous ZAPS enhanced their expression.
    [Show full text]
  • Innate Immune Responses to Avian Influenza Viruses in Ducks
    veterinary sciences Review Innate Immune Responses to Avian Influenza Viruses in Ducks and Chickens Danyel Evseev 1,2 and Katharine E. Magor 1,2,* 1 Department of Biological Sciences, University of Alberta, Edmonton, AB T6G 2E9, Canada; [email protected] 2 Li Ka Shing Institute of Virology, University of Alberta, Edmonton, AB T6G 2E9, Canada * Correspondence: [email protected]; Tel.: +1-(780)-492-5498 Received: 2 November 2018; Accepted: 4 January 2019; Published: 10 January 2019 Abstract: Mallard ducks are important natural hosts of low pathogenic avian influenza (LPAI) viruses and many strains circulate in this reservoir and cause little harm. Some strains can be transmitted to other hosts, including chickens, and cause respiratory and systemic disease. Rarely, these highly pathogenic avian influenza (HPAI) viruses cause disease in mallards, while chickens are highly susceptible. The long co-evolution of mallard ducks with influenza viruses has undoubtedly fine-tuned many immunological host–pathogen interactions to confer resistance to disease, which are poorly understood. Here, we compare innate responses to different avian influenza viruses in ducks and chickens to reveal differences that point to potential mechanisms of disease resistance. Mallard ducks are permissive to LPAI replication in their intestinal tissues without overtly compromising their fitness. In contrast, the mallard response to HPAI infection reflects an immediate and robust induction of type I interferon and antiviral interferon stimulated genes, highlighting the importance of the RIG-I pathway. Ducks also appear to limit the duration of the response, particularly of pro-inflammatory cytokine expression. Chickens lack RIG-I, and some modulators of the signaling pathway and may be compromised in initiating an early interferon response, allowing more viral replication and consequent damage.
    [Show full text]
  • Revealing the Host Antiviral Protein ZAP-S As an Inhibitor of SARS-Cov-2 Programmed Ribosomal Frameshifting
    bioRxiv preprint doi: https://doi.org/10.1101/2021.05.31.445667; this version posted May 31, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Revealing the host antiviral protein ZAP-S as an inhibitor of SARS-CoV-2 programmed ribosomal frameshifting Authors Matthias M. Zimmer1,†, Anuja Kibe1,†, Ulfert Rand2, Lukas Pekarek1, Luka Cicin-Sain2, Neva Caliskan1,3,* Affiliations 1 Helmholtz Institute for RNA-based Infection Research (HIRI), Helmholtz Zentrum für Infektionsforschung (Helmholtz Centre for Infection Research), Josef-Schneider-Strasse 2, 97080, Würzburg, Germany 2 Helmholtz Zentrum für Infektionsforschung, Inhoffenstrasse 7, 38124, Braunschweig, Germany 3 Medical Faculty, Julius-Maximilians University Würzburg, 97074, Würzburg, Germany † These authors contributed equally to this work. *Corresponding author. E-Mail: [email protected] Abstract Programmed ribosomal frameshifting (PRF) is a fundamental gene expression event in many viruses including SARS-CoV-2, which allows production of essential structural and replicative enzymes from an alternative reading frame. Despite the importance of PRF for the viral life cycle, it is still largely unknown how and to what extent cellular factors alter mechanical properties of frameshifting RNA molecules and thereby impact virulence. This prompted us to comprehensively dissect the interplay between the host proteome and the SARS-CoV-2 frameshift element. Here, we reveal that zinc-finger antiviral protein (ZAP-S) is a direct and specific regulator of PRF in SARS-CoV-2 infected cells. ZAP- S overexpression strongly impairs frameshifting and viral replication. Using in vitro ensemble and single-molecule techniques, we further demonstrate that ZAP-S directly interacts with the SARS-CoV-2 RNA and ribosomes and interferes with the folding of the frameshift RNA.
    [Show full text]