Dengue virus (DENV) is one of the most important arthropod-borne pathogens that cause life-threatening diseases in humans. However, no vaccine or specific antiviral is available for dengue. As seen in other RNA viruses, the innate immune system plays a key role in controlling DENV infection and disease outcome. Although the interferon (IFN) response, which is central to host protective immunity, has been reported to limit DENV replication, the molecular details of how DENV infection is modulated by IFN treatment are elusive. In this study, by employing a gain-of-function screen using a type I IFN-treated cell-derived cDNA library, we identified a previously uncharacterized gene, C19orf66, as an IFN-stimulated gene (ISG) that inhibits DENV replication, which we named Repressor of yield of DENV (RyDEN). Overexpression and gene knockdown experiments revealed that expression of RyDEN confers resistance to all serotypes of DENV in human cells. RyDEN expression also limited the replication of hepatitis C virus, Kunjin virus, Chikungunya virus, herpes simplex virus type 1, and human adenovirus. Importantly, RyDEN was considered to be a crucial effector molecule in the IFN-mediated anti-DENV response. When affinity purification-mass spectrometry analysis was performed, RyDEN was revealed to form a complex with cellular mRNA-binding proteins, poly(A)-binding protein cytoplasmic 1 (PABPC1), and La motif-related protein 1 (LARP1). Interestingly, PABPC1 and LARP1 were found to be positive modulators of DENV replication. Since RyDEN influenced intracellular events on DENV replication and, suppression of protein synthesis from DENV-based reporter construct RNA was also observed in RyDEN-expressing cells, our data suggest that RyDEN is likely to interfere with the translation of DENV via interaction with viral RNA and cellular mRNA-binding proteins, resulting in the inhibition of virus replication in infected cells.
Lindenbach BD, Rice CM (2003) Molecular biology of flaviviruses. Adv Virus Res 59: 23–61. pmid:14696326 doi: 10.1016/s0065-3527(03)59002-9
[3]
Fischl W, Bartenschlager R (2011) Exploitation of cellular pathways by Dengue virus. Curr Opin Microbiol 14: 470–475. doi: 10.1016/j.mib.2011.07.012. pmid:21798792
[4]
Fernandez-Garcia MD, Mazzon M, Jacobs M, Amara A (2009) Pathogenesis of flavivirus infections: using and abusing the host cell. Cell Host Microbe 5: 318–328. doi: 10.1016/j.chom.2009.04.001. pmid:19380111
[5]
Morrison J, Aguirre S, Fernandez-Sesma A (2012) Innate immunity evasion by Dengue virus. Viruses 4: 397–413. doi: 10.3390/v4030397. pmid:22590678
[6]
Nasirudeen AM, Wong HH, Thien P, Xu S, Lam KP, et al. (2011) RIG-I, MDA5 and TLR3 synergistically play an important role in restriction of dengue virus infection. PLoS Negl Trop Dis 5: e926. doi: 10.1371/journal.pntd.0000926. pmid:21245912
[7]
Tsai YT, Chang SY, Lee CN, Kao CL (2009) Human TLR3 recognizes dengue virus and modulates viral replication in vitro. Cell Microbiol 11: 604–615. doi: 10.1111/j.1462-5822.2008.01277.x. pmid:19134117
[8]
Loo YM, Fornek J, Crochet N, Bajwa G, Perwitasari O, et al. (2008) Distinct RIG-I and MDA5 signaling by RNA viruses in innate immunity. J Virol 82: 335–345. pmid:17942531 doi: 10.1128/jvi.01080-07
[9]
Ashour J, Laurent-Rolle M, Shi PY, Garcia-Sastre A (2009) NS5 of dengue virus mediates STAT2 binding and degradation. J Virol 83: 5408–5418. doi: 10.1128/JVI.02188-08. pmid:19279106
[10]
Ho LJ, Hung LF, Weng CY, Wu WL, Chou P, et al. (2005) Dengue virus type 2 antagonizes IFN-alpha but not IFN-gamma antiviral effect via down-regulating Tyk2-STAT signaling in the human dendritic cell. J Immunol 174: 8163–8172. pmid:15944325 doi: 10.4049/jimmunol.174.12.8163
[11]
Jones M, Davidson A, Hibbert L, Gruenwald P, Schlaak J, et al. (2005) Dengue virus inhibits alpha interferon signaling by reducing STAT2 expression. J Virol 79: 5414–5420. pmid:15827155 doi: 10.1128/jvi.79.9.5414-5420.2005
[12]
Mazzon M, Jones M, Davidson A, Chain B, Jacobs M (2009) Dengue virus NS5 inhibits interferon-alpha signaling by blocking signal transducer and activator of transcription 2 phosphorylation. J Infect Dis 200: 1261–1270. doi: 10.1086/605847. pmid:19754307
[13]
Munoz-Jordan JL, Laurent-Rolle M, Ashour J, Martinez-Sobrido L, Ashok M, et al. (2005) Inhibition of alpha/beta interferon signaling by the NS4B protein of flaviviruses. J Virol 79: 8004–8013. pmid:15956546 doi: 10.1128/jvi.79.13.8004-8013.2005
[14]
Munoz-Jordan JL, Sanchez-Burgos GG, Laurent-Rolle M, Garcia-Sastre A (2003) Inhibition of interferon signaling by dengue virus. Proc Natl Acad Sci USA 100: 14333–14338. pmid:14612562 doi: 10.1073/pnas.2335168100
[15]
Diamond MS, Roberts TG, Edgil D, Lu B, Ernst J, et al. (2000) Modulation of Dengue virus infection in human cells by alpha, beta, and gamma interferons. J Virol 74: 4957–4966. pmid:10799569 doi: 10.1128/jvi.74.11.4957-4966.2000
[16]
Johnson AJ, Roehrig JT (1999) New mouse model for dengue virus vaccine testing. J Virol 73: 783–786. pmid:9847388
[17]
Perry ST, Buck MD, Lada SM, Schindler C, Shresta S (2011) STAT2 mediates innate immunity to Dengue virus in the absence of STAT1 via the type I interferon receptor. PLoS Pathog 7: e1001297. doi: 10.1371/journal.ppat.1001297. pmid:21379341
[18]
Ashour J, Morrison J, Laurent-Rolle M, Belicha-Villanueva A, Plumlee CR, et al. (2010) Mouse STAT2 restricts early dengue virus replication. Cell Host Microbe 8: 410–421. doi: 10.1016/j.chom.2010.10.007. pmid:21075352
[19]
Kurane I, Innis BL, Nimmannitya S, Nisalak A, Meager A, et al. (1993) High levels of interferon alpha in the sera of children with dengue virus infection. Am J Trop Med Hyg 48: 222–229. pmid:8447527 doi: 10.1172/jci115457
[20]
Kurane I, Innis BL, Nimmannitya S, Nisalak A, Meager A, et al. (1991) Activation of T lymphocytes in dengue virus infections. High levels of soluble interleukin 2 receptor, soluble CD4, soluble CD8, interleukin 2, and interferon-gamma in sera of children with dengue. J Clin Invest 88: 1473–1480. pmid:1939640 doi: 10.1172/jci115457
[21]
Libraty DH, Young PR, Pickering D, Endy TP, Kalayanarooj S, et al. (2002) High circulating levels of the dengue virus nonstructural protein NS1 early in dengue illness correlate with the development of dengue hemorrhagic fever. J Infect Dis 186: 1165–1168. pmid:12355369 doi: 10.1086/343813
[22]
Clyde K, Kyle JL, Harris E (2006) Recent advances in deciphering viral and host determinants of dengue virus replication and pathogenesis. J Virol 80: 11418–11431. pmid:16928749 doi: 10.1128/jvi.01257-06
[23]
Schneider WM, Chevillotte MD, Rice CM (2014) Interferon-stimulated genes: a complex web of host defenses. Annu Rev Immunol 32: 513–545. doi: 10.1146/annurev-immunol-032713-120231. pmid:24555472
[24]
Schoggins JW, Wilson SJ, Panis M, Murphy MY, Jones CT, et al. (2011) A diverse range of gene products are effectors of the type I interferon antiviral response. Nature 472: 481–485. doi: 10.1038/nature09907. pmid:21478870
[25]
Schoggins JW, MacDuff DA, Imanaka N, Gainey MD, Shrestha B, et al. (2014) Pan-viral specificity of IFN-induced genes reveals new roles for cGAS in innate immunity. Nature 505: 691–695. doi: 10.1038/nature12862. pmid:24284630
[26]
Liu SY, Sanchez DJ, Aliyari R, Lu S, Cheng G (2012) Systematic identification of type I and type II interferon-induced antiviral factors. Proc Natl Acad Sci USA 109: 4239–4244. doi: 10.1073/pnas.1114981109. pmid:22371602
[27]
Helbig KJ, Carr JM, Calvert JK, Wati S, Clarke JN, et al. (2013) Viperin is induced following dengue virus type-2 (DENV-2) infection and has anti-viral actions requiring the C-terminal end of viperin. PLoS Negl Trop Dis 7: e2178. doi: 10.1371/journal.pntd.0002178. pmid:23638199
[28]
Jiang D, Weidner JM, Qing M, Pan XB, Guo H, et al. (2010) Identification of five interferon-induced cellular proteins that inhibit west nile virus and dengue virus infections. J Virol 84: 8332–8341. doi: 10.1128/JVI.02199-09. pmid:20534863
[29]
Pan XB, Han JC, Cong X, Wei L (2012) BST2/tetherin inhibits dengue virus release from human hepatoma cells. PLoS One 7: e51033. doi: 10.1371/journal.pone.0051033. pmid:23236425
[30]
Hishiki T, Han Q, Arimoto K, Shimotohno K, Igarashi T, et al. (2014) Interferon-mediated ISG15 conjugation restricts dengue virus 2 replication. Biochem Biophys Res Commun 448: 95–100. doi: 10.1016/j.bbrc.2014.04.081. pmid:24769207
[31]
Brass AL, Huang IC, Benita Y, John SP, Krishnan MN, et al. (2009) The IFITM proteins mediate cellular resistance to influenza A H1N1 virus, West Nile virus, and dengue virus. Cell 139: 1243–1254. doi: 10.1016/j.cell.2009.12.017. pmid:20064371
[32]
Fink J, Gu F, Ling L, Tolfvenstam T, Olfat F, et al. (2007) Host gene expression profiling of dengue virus infection in cell lines and patients. PLoS Negl Trop Dis 1: e86. pmid:18060089 doi: 10.1371/journal.pntd.0000086
[33]
Schoggins JW, Dorner M, Feulner M, Imanaka N, Murphy MY, et al. (2012) Dengue reporter viruses reveal viral dynamics in interferon receptor-deficient mice and sensitivity to interferon effectors in vitro. Proc Natl Acad Sci USA 109: 14610–14615. doi: 10.1073/pnas.1212379109. pmid:22908290
[34]
Diamond MS, Harris E (2001) Interferon inhibits dengue virus infection by preventing translation of viral RNA through a PKR-independent mechanism. Virology 289: 297–311. pmid:11689052 doi: 10.1006/viro.2001.1114
[35]
Kawano Y, Yoshida T, Hieda K, Aoki J, Miyoshi H, et al. (2004) A lentiviral cDNA library employing lambda recombination used to clone an inhibitor of human immunodeficiency virus type 1-induced cell death. J Virol 78: 11352–11359. pmid:15452256 doi: 10.1128/jvi.78.20.11352-11359.2004
[36]
Low JG, Ooi EE, Tolfvenstam T, Leo YS, Hibberd ML, et al. (2006) Early Dengue infection and outcome study (EDEN)—study design and preliminary findings. Ann Acad Med Singapore 35: 783–789. pmid:17160194
[37]
Krishna SS, Majumdar I, Grishin NV (2003) Structural classification of zinc fingers: survey and summary. Nucleic Acids Res 31: 532–550. pmid:12527760 doi: 10.1093/nar/gkg161
[38]
Ishikawa H, Barber GN (2011) The STING pathway and regulation of innate immune signaling in response to DNA pathogens. Cell Mol Life Sci 68: 1157–1165. doi: 10.1007/s00018-010-0605-2. pmid:21161320
[39]
Bidet K, Dadlani D, Garcia-Blanco MA (2014) G3BP1, G3BP2 and CAPRIN1 are required for translation of interferon stimulated mRNAs and are targeted by a dengue virus non-coding RNA. PLoS Pathog 10: e1004242. doi: 10.1371/journal.ppat.1004242. pmid:24992036
[40]
Le Sommer C, Barrows NJ, Bradrick SS, Pearson JL, Garcia-Blanco MA (2012) G protein-coupled receptor kinase 2 promotes flaviviridae entry and replication. PLoS Negl Trop Dis 6: e1820. doi: 10.1371/journal.pntd.0001820. pmid:23029581
[41]
Low JS, Wu KX, Chen KC, Ng MM, Chu JJ (2011) Narasin, a novel antiviral compound that blocks dengue virus protein expression. Antivir Ther 16: 1203–1218. doi: 10.3851/IMP1884. pmid:22155902
[42]
Ng CY, Gu F, Phong WY, Chen YL, Lim SP, et al. (2007) Construction and characterization of a stable subgenomic dengue virus type 2 replicon system for antiviral compound and siRNA testing. Antiviral Res 76: 222–231. pmid:17662475 doi: 10.1016/j.antiviral.2007.06.007
[43]
Diamond MS, Zachariah M, Harris E (2002) Mycophenolic acid inhibits dengue virus infection by preventing replication of viral RNA. Virology 304: 211–221. pmid:12504563 doi: 10.1006/viro.2002.1685
[44]
Burckstummer T, Bennett KL, Preradovic A, Schutze G, Hantschel O, et al. (2006) An efficient tandem affinity purification procedure for interaction proteomics in mammalian cells. Nat Methods 3: 1013–1019. pmid:17060908 doi: 10.1038/nmeth968
[45]
Gorgoni B, Gray NK (2004) The roles of cytoplasmic poly(A)-binding proteins in regulating gene expression: a developmental perspective. Brief Funct Genomic Proteomic 3: 125–141. pmid:15355595 doi: 10.1093/bfgp/3.2.125
[46]
Brook M, Smith JW, Gray NK (2009) The DAZL and PABP families: RNA-binding proteins with interrelated roles in translational control in oocytes. Reproduction 137: 595–617. doi: 10.1530/REP-08-0524. pmid:19225045
[47]
Mangus DA, Evans MC, Jacobson A (2003) Poly(A)-binding proteins: multifunctional scaffolds for the post-transcriptional control of gene expression. Genome Biol 4: 223. pmid:12844354
[48]
Kuhn U, Wahle E (2004) Structure and function of poly(A) binding proteins. Biochim Biophys Acta 1678: 67–84. pmid:15157733 doi: 10.1016/j.bbaexp.2004.03.008
[49]
Polacek C, Friebe P, Harris E (2009) Poly(A)-binding protein binds to the non-polyadenylated 3' untranslated region of dengue virus and modulates translation efficiency. J Gen Virol 90: 687–692. doi: 10.1099/vir.0.007021-0. pmid:19218215
[50]
Bayfield MA, Yang R, Maraia RJ (2010) Conserved and divergent features of the structure and function of La and La-related proteins (LARPs). Biochim Biophys Acta 1799: 365–378. doi: 10.1016/j.bbagrm.2010.01.011. pmid:20138158
[51]
Ichihara K, Shimizu H, Taguchi O, Yamaguchi M, Inoue YH (2007) A Drosophila orthologue of larp protein family is required for multiple processes in male meiosis. Cell Struct Funct 32: 89–100. pmid:17951964 doi: 10.1247/csf.07027
[52]
Chauvet S, Maurel-Zaffran C, Miassod R, Jullien N, Pradel J, et al. (2000) dlarp, a new candidate Hox target in Drosophila whose orthologue in mouse is expressed at sites of epithelium/mesenchymal interactions. Dev Dyn 218: 401–413. pmid:10878606 doi: 10.1002/1097-0177(200007)218:3<401::aid-dvdy1009>3.0.co;2-6
[53]
Blagden SP, Gatt MK, Archambault V, Lada K, Ichihara K, et al. (2009) Drosophila Larp associates with poly(A)-binding protein and is required for male fertility and syncytial embryo development. Dev Biol 334: 186–197. doi: 10.1016/j.ydbio.2009.07.016. pmid:19631203
[54]
Burrows C, Abd Latip N, Lam SJ, Carpenter L, Sawicka K, et al. (2010) The RNA binding protein Larp1 regulates cell division, apoptosis and cell migration. Nucleic Acids Res 38: 5542–5553. doi: 10.1093/nar/gkq294. pmid:20430826
[55]
Aoki K, Adachi S, Homoto M, Kusano H, Koike K, et al. (2013) LARP1 specifically recognizes the 3' terminus of poly(A) mRNA. FEBS Lett 587: 2173–2178. doi: 10.1016/j.febslet.2013.05.035. pmid:23711370
[56]
Takahashi H, Takahashi C, Moreland NJ, Chang YT, Sawasaki T, et al. (2012) Establishment of a robust dengue virus NS3-NS5 binding assay for identification of protein-protein interaction inhibitors. Antiviral Res 96: 305–314. doi: 10.1016/j.antiviral.2012.09.023. pmid:23072882
[57]
Schmidt EK, Clavarino G, Ceppi M, Pierre P (2009) SUnSET, a nonradioactive method to monitor protein synthesis. Nat Methods 6: 275–277. doi: 10.1038/nmeth.1314. pmid:19305406
[58]
Groat-Carmona AM, Orozco S, Friebe P, Payne A, Kramer L, et al. (2012) A novel coding-region RNA element modulates infectious dengue virus particle production in both mammalian and mosquito cells and regulates viral replication in Aedes aegypti mosquitoes. Virology 432: 511–526. doi: 10.1016/j.virol.2012.06.028. pmid:22840606
[59]
Meertens L, Carnec X, Lecoin MP, Ramdasi R, Guivel-Benhassine F, et al. (2012) The TIM and TAM families of phosphatidylserine receptors mediate dengue virus entry. Cell Host Microbe 12: 544–557. doi: 10.1016/j.chom.2012.08.009. pmid:23084921
[60]
Yi Z, Sperzel L, Nurnberger C, Bredenbeek PJ, Lubick KJ, et al. (2011) Identification and characterization of the host protein DNAJC14 as a broadly active flavivirus replication modulator. PLoS Pathog 7: e1001255. doi: 10.1371/journal.ppat.1001255. pmid:21249176
[61]
Kranzusch PJ, Lee AS, Berger JM, Doudna JA (2013) Structure of human cGAS reveals a conserved family of second-messenger enzymes in innate immunity. Cell Rep 3: 1362–1368. doi: 10.1016/j.celrep.2013.05.008. pmid:23707061
[62]
McKinney C, Yu D, Mohr I (2013) A new role for the cellular PABP repressor Paip2 as an innate restriction factor capable of limiting productive cytomegalovirus replication. Genes Dev 27: 1809–1820. doi: 10.1101/gad.221341.113. pmid:23964095
[63]
Khaleghpour K, Kahvejian A, De Crescenzo G, Roy G, Svitkin YV, et al. (2001) Dual interactions of the translational repressor Paip2 with poly(A) binding protein. Mol Cell Biol 21: 5200–5213. pmid:11438674 doi: 10.1128/mcb.21.15.5200-5213.2001
[64]
Merret R, Descombin J, Juan YT, Favory JJ, Carpentier MC, et al. (2013) XRN4 and LARP1 are required for a heat-triggered mRNA decay pathway involved in plant acclimation and survival during thermal stress. Cell Rep 5: 1279–1293. doi: 10.1016/j.celrep.2013.11.019. pmid:24332370
[65]
Kozlov G, Safaee N, Rosenauer A, Gehring K (2010) Structural basis of binding of P-body-associated proteins GW182 and ataxin-2 by the Mlle domain of poly(A)-binding protein. J Biol Chem 285: 13599–13606. doi: 10.1074/jbc.M109.089540. pmid:20181956
[66]
Walsh D, Mohr I (2011) Viral subversion of the host protein synthesis machinery. Nat Rev Microbiol 9: 860–875. doi: 10.1038/nrmicro2655. pmid:22002165
[67]
Ariumi Y, Kuroki M, Kushima Y, Osugi K, Hijikata M, et al. (2011) Hepatitis C virus hijacks P-body and stress granule components around lipid droplets. J Virol 85: 6882–6892. doi: 10.1128/JVI.02418-10. pmid:21543503
[68]
Blight KJ, McKeating JA, Rice CM (2002) Highly permissive cell lines for subgenomic and genomic hepatitis C virus RNA replication. J Virol 76: 13001–13014. pmid:12438626 doi: 10.1128/jvi.76.24.13001-13014.2002
[69]
Deng L, Adachi T, Kitayama K, Bungyoku Y, Kitazawa S, et al. (2008) Hepatitis C virus infection induces apoptosis through a Bax-triggered, mitochondrion-mediated, caspase 3-dependent pathway. J Virol 82: 10375–10385. doi: 10.1128/JVI.00395-08. pmid:18768989
[70]
Tan BH, Suzuki Y, Takahashi H, Ying PH, Takahashi C, et al. (2014) Identification of RFPL3 protein as a novel E3 ubiquitin ligase modulating the integration activity of human immunodeficiency virus, type 1 preintegration complex using a microtiter plate-based assay. J Biol Chem 289: 26368–26382. doi: 10.1074/jbc.M114.561662. pmid:25107902
[71]
Callahan JD, Wu SJ, Dion-Schultz A, Mangold BE, Peruski LF, et al. (2001) Development and evaluation of serotype- and group-specific fluorogenic reverse transcriptase PCR (TaqMan) assays for dengue virus. J Clin Microbiol 39: 4119–4124. pmid:11682539 doi: 10.1128/jcm.39.11.4119-4124.2001
[72]
Richardson J, Molina-Cruz A, Salazar MI, Black Wt (2006) Quantitative analysis of dengue-2 virus RNA during the extrinsic incubation period in individual Aedes aegypti. Am J Trop Med Hyg 74: 132–141. pmid:16407358
[73]
Khromykh AA, Kenney MT, Westaway EG (1998) trans-Complementation of flavivirus RNA polymerase gene NS5 by using Kunjin virus replicon-expressing BHK cells. J Virol 72: 7270–7279. pmid:9696822
[74]
Sawasaki T, Ogasawara T, Morishita R, Endo Y (2002) A cell-free protein synthesis system for high-throughput proteomics. Proc Natl Acad Sci USA 99: 14652–14657. pmid:12409616 doi: 10.1073/pnas.232580399
