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13225–13230 (1995).

ACKNOWLEDGMENTS

We thank all of the members of the Sabatini and Sawyerslaboratories for critical readings of this manuscript and helpfuldiscussions. We also thank N. D. Novikov, M. A. Domser, K. C. Wood,

D. J. Spencer, T. R. Peterson, C. Thoreen, S. Kinkel, M. J. Evans,R. Bose, P. Watson, O. S. Andersen, S. C. Blanchard, T. E. McGraw,N. Rosen, and A. Smogorzewska for helpful discussions; S. Silverfor assistance with the RNAi screens; V. Boyko and Y. Romin of theMSKCC Molecular Cytology Core Facility for assistance withconfocal microscopy; A. Kazlauskas for helpful discussions aboutPDGFR biology; and the late A. Hall for helpful input about GTPasebiology. Funds for the RNAi screen were provided by the StarrCancer Consortium Award I2-A117. D.B.W. was supported by theDepartment of Defense Prostate Cancer Research Program via theOffice of the Congressionally Directed Medical Research ProgramsPredoctoral Prostate Cancer Training Award PC094483, as well asby NIH Medical Scientist Training Program grant GM07739. R.Z.was supported by the Glenn Foundation for Medical Research, theNIH Director’s New Innovator Award 1DP2CA195761-01, and thePew-Stewart Scholarship for Cancer Research. This work was

partially supported by NIH grants CA103866 and AI47389 (toD.M.S.) and CA155169 and CA092629 (to C.L.S.). Data from theRNAi screen described in this manuscript can be found in thesupplementary materials.

SUPPLEMENTARY MATERIALS

www.sciencemag.org/content/350/6257/211/suppl/DC1Materials and MethodsFigs. S1 to S15Tables S1 to S7References (38–55)

14 December 2014; accepted 24 August 2015Published online 3 September 201510.1126/science.aaa4903

VIROLOGY

Dengue subgenomic RNA bindsTRIM25 to inhibit interferonexpression for epidemiological fitnessGayathri Manokaran,1,2,3 Esteban Finol,1,3,4 Chunling Wang,5 Jayantha Gunaratne,3,6

Justin Bahl,7 Eugenia Z. Ong,1 Hwee Cheng Tan,1 October M. Sessions,1 Alex M. Ward,1

Duane J. Gubler,1 Eva Harris,5 Mariano A. Garcia-Blanco,1,8,9 Eng Eong Ooi1,3,10,11*

The global spread of dengue virus (DENV) infections has increased viral genetic diversity,some of which appears associated with greater epidemic potential. The mechanismsgoverning viral fitness in epidemiological settings, however, remain poorly defined. Weidentified a determinant of fitness in a foreign dominant (PR-2B) DENV serotype 2 (DENV-2)clade, which emerged during the 1994 epidemic in Puerto Rico and replaced an endemic (PR-1)DENV-2 clade.The PR-2B DENV-2 produced increased levels of subgenomic flavivirus RNA(sfRNA) relative to genomic RNA during replication. PR-2B sfRNA showed sequence-dependent binding to and prevention of tripartite motif 25 (TRIM25) deubiquitylation, which iscritical for sustained and amplified retinoic acid–inducible gene 1 (RIG-I)–induced type Iinterferon expression. Our findings demonstrate a distinctive viral RNA–host proteininteraction to evade the innate immune response for increased epidemiological fitness.

Dengue outbreaks emerge when DENVs areintroduced into populationswith lowherdimmunity.However, several outbreakshaveensued when new DENV strains emergedand displaced endemic strains, albeit of the

same serotype and genotype (1–6), suggesting thatgenetic variation altered viral fitness in epidemi-ological settings.

To gain insights into how DENV genome var-iation affects its epidemiological phenotype, weexamined aDENV-2 clade replacement event thatcoincided with an epidemic of severe dengue inPuerto Rico in 1994 (1, 7, 8). Phylogenetic analysisof the complete coding sequences of DENV-2 iso-lated from Puerto Rico and neighboring countriesshowed that threedistinct cladeswithin theAsian/American genotype circulated in 1994 (fig. S1), con-cordant with previous findings (1). Clade PR-1 wasprevalent from 1986 to 1995. Clade PR-2 containedviruses in two subclades, both first isolated in 1994:PR-2A persisted at low levels until 1998, whereasPR-2B spread and replaced PR-1 to become thedominant DENV-2 in Puerto Rico from 1995 to2007. Immune escape is an unlikely explanationfor PR-2B emergence, as both PR-1 and PR-2Bbelong to the Asian/American genotype. Fur-thermore, prM and E amino acid sequenceanalysis showed weak bootstrap values (fig. S2),indicating low levels of antigenic differencesbetween the clades. Instead, PR-2B emergenceand subsequent replacement of PR-1 virusescould be due to differences in epidemiologicalfitness.

To identify the genomic variations that segre-gated theseDENVs into different clades andmod-ulated epidemiological fitness, we applied theShimodaira-Hasegawa (SH) test (9) that comparesthe topology of region-specific with full-genometrees. This approach identifiedNS1, NS3,NS5, andthe 3′ untranslated region (3′UTR) sequence var-iations as potentially critical in segregatingDENVsinto PR-1 and PR-2B (figs. S3 to S5).NS1, NS3, and NS5 are DENV replication com-

plex components that also regulate the innateimmune response (10). Less is known about the3′UTR. During flaviviral replication, the cellular5′-to-3′ exoribonuclease digests uncapped genomesbut stalls at pseudoknot RNA structures in the3′UTR. The resultant 0.3- to 0.5-kb subgenomicflavivirus RNA (sfRNA) regulates pathogenicityin both mammalian and mosquito cells, likelythrough mechanisms and structural elementswithin its noncoding RNA that remain to be fullydefined (11–15). Recently, we showed that sfRNAcould interact with proteins to inhibit transla-tion of interferon-stimulated genes (16). Becausetwo substitutions identified in the PR-2B 3′UTRwere located where pseudoknots could form(fig. S6) (17–19), we tested for a possible role forsfRNA in modulating fitness in our viruses.To measure viral fitness in vitro, we performed

DENV infection assays in human hepatoma(HuH-7) cells. We expected PR-2B DENVs to rep-licate more efficiently than PR-1 viruses. Paradox-ically, however, more viral progeny (Fig. 1A) andgenomic RNA (gRNA) (Fig. 1B) were produced byPR-1 than PR-2B viruses at 24 hours postinfection(hpi). The difference in sfRNA levels betweenPR-1 and PR-2B viruses was smaller, although sta-tistically significant (Fig. 1C), resulting in highersfRNA:gRNA ratios in PR-2B as compared toPR-1 viruses (Fig. 1D). While the absolute valuesof the sfRNA:gRNA ratios should be interpretedwith caution, our data clearly show that theseratios are markedly higher for the epidemicPR-2B viruses.Increasedproduction of sfRNA relative to gRNA

in an epidemic clade of viruses is not specific tothe PR-2B viruses. In Nicaragua, NI-2B DENV-2emerged abruptly in 2005, displaced the endemicNI-1DENV-2, and caused increased rates of severedisease across several epidemic seasons (5). SH teston these Asian/American genotype DENV-2 iso-lates similarly identified variations in NS1, NS5,

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1Program in Emerging Infectious Diseases, Duke–NationalUniversity of Singapore Graduate Medical School, Singapore.2Defence Medical and Environmental Research Institute, DSONational Laboratories, Singapore. 3Yong Loo Lin School ofMedicine, National University of Singapore, Singapore. 4SwissTropical and Public Health Institute, Universität Basel,Switzerland. 5Division of Infectious Diseases and Vaccinology,School of Public Health, University of California, Berkeley,CA, USA. 6Institute of Molecular and Cell Biology, Singapore.7Center for Infectious Diseases, The University of TexasSchool of Public Health, Houston, TX, USA. 8Department ofBiochemistry and Molecular Biology, The University of TexasMedical Branch, Galveston, TX, USA. 9Department ofMolecular Genetics and Microbiology and Center for RNABiology, Duke University, Durham, NC, USA. 10Saw SweeHock School of Public Health, National University ofSingapore, Singapore. 11Singapore–Massachusetts Institute ofTechnology Alliance in Research and Technology InfectiousDisease Interdisciplinary Research Group, Singapore.*Corresponding author. E-mail: engeong.ooi@duke-nus.edu.sg

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and 3′UTR as important in segregating these vi-ruses intoNI-1 andNI-2B clades (fig. S7 and S8). Inthis case, two compensatory 3′UTRsubstitutions inNI-2B may produce energetically more stable se-condary RNA structures (fig. S9). Further analysisindicated three- to fourfold higher sfRNA:gRNAratios upon NI-2B as compared to NI-1 DENV-2infection (Fig. 1E). Collectively, these data suggestthat increased sfRNA production and possibly thesequence or structure of sfRNAmay contribute toDENV-2 fitness indistinct epidemiological settings.To understand how sfRNA production and/or

sequence could lead to greater epidemiologicalfitness, we compared the replication kinetics ofselected PR-1 and PR-2B viruses and their effecton interferon expression. Although significant dif-ferences in plaque titers and gRNA levels exist at24 hpi, PR-2B viruses grew at a faster rate thanPR-1 such that no difference in either parameterwas observed at 96 hpi (Fig. 2, A and B). In addi-tion, interferon-beta (IFN-b) expression was con-sistently lower in PR-2B– than in PR-1–infectedcells, despitediminisheddifferences inDENVgRNAlevels at later time points (Fig. 2, C to F). Thissuggests that higher sfRNA:gRNA ratios producedduring PR-2B infection suppressed IFN-b in-duction. Indeed, silencing interferon regulatoryfactor–3 (IRF-3), a transcription factor for IFN-b,increased PR-1 but not PR-2B replication (Fig. 2G).The trends observed in infection of HuH-7 cells

were recapitulated in primary monocytes, whichare target cells for DENV in humans. Antibody-enhanced infection, which is associated withgreater risk of severe dengue, produced less gRNA,greater sfRNA:gRNA ratios, and reduced IFN-b ex-pressionat24hpiwhenPR-2BinsteadofPR-1viruseswere used (Fig. 2, H to J). Likewise, at 72 hpi, dif-ferences between the plaque titers of PR-1 and

PR-2B isolates were reduced or even reversed(Fig. 2K).Given the observed association between in-

creased sfRNA:gRNA ratios and reduced IFN-bexpression, we next examined if the three nucle-otide substitutions in the 3′UTRcould functionallyaccount for the increased sfRNA:gRNA ratios. Toeliminate any contributions from NS1, NS3, andNS5, we performed mutagenesis studies on astandardized genomic backbone using a previ-ously characterized DENV-2 replicon reporter,pDENrep-FH (20). At 72 hours after electropora-tion, replicons with PR-2B residues produced lessgRNAbut higher sfRNA:gRNA ratios compared tothat bearing PR-1 residues (Fig. 3A). Likewise,luciferase signals were lower in replicons withPR-2B instead of PR-1 residues (Fig. 3B), indicat-ing that substitutions in these three nucleotidepositions can account for the observed differencesin PR-1 and PR-2B sfRNA:gRNA ratios.To investigate whether sfRNA plays a func-

tional role inmodulating IFN-b expression, we co-electroporated in vitro–transcribed sfRNA fromPR-1 or PR-2B or a size-matchedRNA control withpDENrep-FH into HuH-7 cells. As compared tonuclear factor kB (NF-kB) expression, where nodifference was seen, IFN-b expression was signif-icantly reduced with PR-2B but not PR-1 sfRNA,at 96 hours after electroporation (Fig. 3C). Corre-spondingly, pDENrep-FH replication was signifi-cantly increased when co-electroporated with PR-2BsfRNA compared to either PR-1 sfRNA or con-trol RNA (Fig. 3D). Furthermore, sfRNA but notsize-matched control RNA inhibited polyinosinic-polycytidylic acid (polyIC)–induced IFN-b ex-pression, with a greater difference observed forPR-2B than PR-1 sfRNA (fig. S10 and Fig. 3E).Collectively, these data indicate that both the

amount and sequence of sfRNA are importantin attenuating type I interferon expression forincreased DENV replication.The sequence specificity observed suggests that

IFN-b suppression is likely to be due to specificsfRNA-protein interactions (21). Using stable iso-tope labeling by amino acids in cell culture coupledwith quantitative mass spectrometry (SILAC-qMS),wedetected 198proteins thatwere enriched(at least 1.5-fold change) with DENV 3′UTR com-pared to controlRNA (table S1 and fig. S11, A andB)(22). Notably, tripartite motif containing 25(TRIM25) and mitochondrial antiviral signaling(MAVS) proteins (fig. S11C), which are knownretinoic acid–inducible gene 1 (RIG-I) signalingintermediates for type I interferon expression(23–25), were significantly enriched with PR-2Bcompared to PR-1 3′UTR.To validate our SILAC-qMS findings, we immu-

noprecipitated TRIM25 and MAVS separatelyfrom DENV-2–infected cells and quantified theprotein-bound sfRNA and gRNA. Significantlyhigher sfRNA than gRNA levels were detectedwith TRIM25 immunoprecipitation as comparedto isotype immunoglobulin G (IgG) control, withthe difference being more pronounced whenPR-2BDENVwasused (Fig. 4A). Conversely, neithersfRNAnor gRNAwas enrichedwithMAVS immu-noprecipitation (fig. S12). In addition, incubatingPR-2B sfRNA with TRIM25 yielded more RNA-protein complex than PR-1 sfRNA asmeasured onan electrophoretic mobility shift assay (EMSA)(Fig. 4B), indicating sequence-specific differencesin TRIM25 binding.Besides being an RNA-binding protein (26),

TRIM25 is an E3 ligase that, upon deubiquityl-ation by ubiquitin-specific peptidase 15 (USP15)(27), polyubiquitylates RIG-I for sustained and

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Fig. 1. sfRNA is a critical determinant of the epi-demic potential of DENV-2. (A) Plaque titers of PR-1 andPR-2BDENV-2 viruses at 24 hpi in HuH-7 cells infected at amultiplicity of infection (MOI) of 0.1. PFU, plaque-formingunits. (B) PR-1 and PR-2B gRNA in infected HuH-7 cells(MOI = 0.1), 24 hpi. GAPDH, glyceraldehyde-3-phosphatedehydrogenase. (C) PR-1 and PR-2B sfRNA in infectedHuH-7 cells (MOI = 0.1), 24 hpi. (D) Ratio of sfRNA:gRNAin HuH-7 cells infected with PR-1 and PR-2B viruses atMOI 0.1, 24 hpi. (E) Quantification of sfRNA:gRNA ratiosinHuH-7 cells at 48 hpiwith clinical isolates fromNicaragua.Numbers below the bars refer to the names of the viralisolates. Data are expressed as mean T SD from three in-dependent experiments. **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, as determined by t test.

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Fig. 2. PR-2B DENV-2 replication is associated with reduced expres-sion of IFN-b relative to PR-1 DENV-2. (A) Plaque titers after infection ofHuH-7 cells with selected viruses from each PR clade at 24 and 96 hpi. (B)DENV-2 gRNA levels in infected HuH-7 cells after infection (MOI = 0.5) withselected viruses from each clade at various time points postinfection. (C to F)Expression levels of IFN-b in infected HuH-7 cells at 24 hpi (C), 48 hpi (D),72 hpi (E), and 96 hpi (F). PR-1: blue bars; PR-2B: red bars. (G) Plaque titers ofsupernatant quantified at 24 hpi with representative viruses from each clade,following knockdown of IRF-3 in HuH-7 cells by small interfering RNA (siRNA).

siC: control RNA; siIRF-3: siRNA against IRF3. (H) Quantification of DENV-2gRNA in infected primary monocytes at 24 hpi. Infection was performed withselected PR-1 and PR-2B DENV-2 viruses opsonized with enhancingconcentrations of humanized 3H5 monoclonal antibody against DENV-2 Eprotein. (I) Quantification of sfRNA:gRNA ratios in primary monocytes, 24 hpi.(J) IFN-b expression in primary monocytes at 24 hpi. (K) Plaque titers fromsupernatant of infected primary monocytes at 72 hpi. Data are expressed asmean TSD from three independent experiments. n.s.: not significant; *P≤0.05,**P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001, as determined by t test.

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amplified signal transduction (24). To examine ifsfRNA interferes with these processes, we immu-noprecipitatedTRIM25 fromDENV-2–infected celllysates and probed for ubiquitin. Results showed aband identical in size to TRIM25 in infected celllysates (Fig. 4C), although the intensity of theubiquitylatedTRIM25band is strongerwith PR-2Bcompared to PR-1 DENV (Fig. 4D). Furthermore,RIG-I immunoprecipitation fromsfRNA-transfectedcell lysates followed by TRIM25 immunoblottingshowed that sfRNA did not prevent TRIM25 frombinding RIG-I (Fig. 4E). Instead, higher levels ofubiquitylated TRIM25 were coimmunoprecipi-tated with RIG-I upon transfection of PR-2B rel-ative to PR-1 sfRNA (Fig. 4E). Consistently, silencingTRIM25 did not augment PR-2B DENV-2 repli-cation, whereas PR-1 DENV-2 replication was sig-nificantly increased (Fig. 4F). These findingscollectively indicate that PR-2B sfRNA bindsTRIM25 more efficiently to interfere with its de-ubiquitylation, thus preventing amplified and sus-tained RIG-I signaling for type I IFN induction.Our report provides a mechanistic explanation

for increased DENV fitness in an epidemiological

setting. It adds to the general theme by whichFlaviviruses use their abundant noncoding RNAto bind and inactivate RNA-binding proteins crit-ical for innate immunity (21). TRIM25 is alsotargeted by influenza virus to inhibit RIG-I sig-naling, although in this instance, inhibition ismediated by NS1 protein binding to TRIM25 (23).Reducing sustained and amplified IFN expressioncould thus be important to many viruses, as IFNrenders uninfected cells resistant to viral infection.Based on our findings, we propose a model to

explain the 1994 dengue outbreak in Puerto Rico(fig. S13). The high sfRNA:gRNA ratios producedby PR-2B viruses during early phases of the in-fection constitute a “one-two punch” against hostresponse; greater levels of sfRNA inhibit TRIM25ina sequence-dependentmanner,whereas reducedgRNA results in lower stimulation of RIG-I/mela-noma differentiation–associated protein 5 (MDA5)RNA sensors. Reduced IFN responses in the earlystages of infection would ensure availability of sus-ceptible cells for viral spread in a human host toreach viremia levels sufficient for furthermosquito-borne transmission.

In conclusion, our study provides unique mo-lecular insights into the epidemiological fitness ofDENV. It also suggests that by combining epi-demiologic studies with molecular investigations,viral phylogenetic information can be informativenot onlywith respect to virus evolution but also asa predictor of its epidemic potential.

REFERENCES AND NOTES

1. S. N. Bennett et al., J. Gen. Virol. 87, 885–893 (2006).2. M. G. Guzman et al., Nat. Rev. Microbiol. 8 (suppl.), S7–S16 (2010).3. K. S. Lee et al., Emerg. Infect. Dis. 16, 847–849 (2010).4. W. B. Messer, D. J. Gubler, E. Harris, K. Sivananthan,

A. M. de Silva, Emerg. Infect. Dis. 9, 800–809 (2003).5. M. OhAinle et al., Sci. Transl. Med. 3, 114ra128 (2011).6. A. Steel, D. J. Gubler, S. N. Bennett, Virology 405, 505–512 (2010).7. J. G. Rigau-Pérez, A. V. Vorndam, G. G. Clark, Am. J. Trop. Med.

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Biol. Evol. 18, 223–234 (2001).10. J. Ye, B. Zhu, Z. F. Fu, H. Chen, S. Cao, Vaccine 31, 461–471 (2013).11. A. Schuessler et al., J. Virol. 86, 5708–5718 (2012).12. K. C. Lin, H. L. Chang, R. Y. Chang, J. Virol. 78, 5133–5138 (2004).13. G. P. Pijlman et al., Cell Host Microbe 4, 579–591 (2008).14. J. A. Roby, G. P. Pijlman, J. Wilusz, A. A. Khromykh, Viruses 6,

404–427 (2014).

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Fig. 3. sfRNA attenuates type I interferon antiviral response, resulting inincreased viral replication. (A) Quantification of gRNA levels and sfRNA:gRNA ratios in HuH-7 cells, 72 hours after electroporation with repliconsbearing nucleotide residues representative of PR-1 or PR-2B DENV-2. Barsshow the gRNA levels (left axis), and lines show sfRNA:gRNA ratios (rightaxis). (B) Luciferase reporter activity for the original replicon (pDENrep-FH)and mutated replicons at various time points after electroporation.Determination of firefly luciferase (FLuc) levels served as internal normalizationcontrols. (C) Quantification of IFN-b and NF-kB expression using quantitative

reverse transcription–polymerase chain reaction (QRT-PCR) at 96 hours afterelectroporation and (D) luciferase reporter activity following co-electroporationof pDENrep-FH with control RNA or sfRNA from PR1 or PR2B. FLuc levelsserved as internal normalization controls. (E) Quantification of IFN-b expres-sion in HuH-7 cells transfected with polyIC only, polyIC and size-matchedcontrol RNA, or sfRNA from PR-1 or PR-2B, by QRT-PCR at 24 hours aftertransfection. Data are expressed as mean T SD from three independentexperiments. n.s.: not significant; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, asdetermined by t test.

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P. J. Bredenbeek, J. Virol. 84, 11395–11406 (2010).20. K. L. Holden et al., Virology 344, 439–452 (2006).21. K. Bidet, M. A. Garcia-Blanco, Biochem. J. 462, 215–230

(2014).22. A. M. Ward et al., RNA Biol. 8, 1173–1186 (2011).23. M. U. Gack et al., Cell Host Microbe 5, 439–449 (2009).24. M. U. Gack et al., Nature 446, 916–920 (2007).

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26. N. R. Choudhury et al., Cell Reports 9, 1265–1272 (2014).27. E. K. Pauli et al., Sci. Signal. 7, ra3 (2014).

ACKNOWLEDGMENTS

This work was supported by the Singapore National MedicalResearch Council; the Duke–National University of SingaporeSignature Research Program funded by the Ministry of Health,Singapore; the Institute of Molecular and Cell Biology, Agency ofScience, Technology and Research, Singapore; and grants R01GM087405 and R01 AI089526 from the U.S. National Institutes ofHealth (E.H. and M.A.G.-B., respectively). We thank K. Bidet andV. Dhanasekaran for their invaluable advice throughout this work.

We thank Y. Chandran and S. M. Cave for technical assistance andM. Krishnan for the IRF-3 knockdown protocol. We also thank theanonymous reviewers for their helpful suggestions.

SUPPLEMENTARY MATERIALS

www.sciencemag.org/content/350/6257/217/suppl/DC1Materials and MethodsFigs. S1 to S13Table S1References (28–40)

13 April 2015; accepted 22 June 2015Published online 2 July 201510.1126/science.aab3369

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Fig. 4. sfRNA binds TRIM25 to inhibit TRIM25 deubiquitylation. (A) RNA-immunoprecipitation (RIP) was performed on HuH-7 cells at 24 hpi with onevirus from each clade (MOI = 5). Infected cells were subjected to TRIM25 orIgG control pull-down followed by immunoblotting with TRIM25 to confirmefficacy of pull-down. Input: cell lysate before RIP; Sup: supernatant afterincubation of antibody-coated beads with cell lysate; Pellet: final product afterwashing. Bar graphs show fold enrichment of sfRNA and gRNA upon pull-down with TRIM25, as measured by QRT-PCR. (B) EMSA of synthetic PR-1 andPR-2B sfRNA (30 nM) incubated with increasing concentrations of TRIM25.Arrow indicates the position of the sfRNA-protein (ribonucleoprotein) complex.Amount of ribonucleoprotein complex was quantified and expressed as apercentage of total RNA signal. (C) Lysates from HuH-7 cells infected with one

virus from each clade (MOI = 5) were subjected to TRIM25 or IgG control pull-down followed by immunoblotting with TRIM25 and ubiquitin (UB). Arrowindicates ubiquitylated bands. (D) Bar chart depicts percentage of ubiquity-lated TRIM25 over total TRIM25. (E) After transfection with PR-1 or PR-2BsfRNA, HuH-7 cell lysates were subjected to RIG-I pull-down followed byimmunoblotting with RIG-I, TRIM25, and UB. (F) PR-1 and PR-2B DENV-2replication at 24 hpi in TRIM25-silenced HuH-7 cells infected with one virusfrom each clade. siC: control RNA; siTRIM25: siRNA against TRIM25. Differ-ences in gel electrophoresis conditions could have led to slight variations inmobility of prestained protein markers. Data are expressed as mean T SD fromthree independent experiments. n.s.: not significant; *P ≤ 0.05, **P ≤ 0.01,****P ≤ 0.0001, as determined by t test.

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fitnessDengue subgenomic RNA binds TRIM25 to inhibit interferon expression for epidemiological

October M. Sessions, Alex M. Ward, Duane J. Gubler, Eva Harris, Mariano A. Garcia-Blanco and Eng Eong OoiGayathri Manokaran, Esteban Finol, Chunling Wang, Jayantha Gunaratne, Justin Bahl, Eugenia Z. Ong, Hwee Cheng Tan,

originally published online July 2, 2015DOI: 10.1126/science.aab3369 (6257), 217-221.350Science 

, this issue p. 217Scienceantiviral response, and so by reducing host immunity was able to increase its own fitness.amounts of genomic viral RNA. sfRNA bound to and inhibited TRIM25, a protein important for activating the host's epidemic strain produced elevated amounts of subgenomic flavivirus RNA (sfRNA), a viral noncoding RNA, relative toexample, the emergence of a new clade of dengue virus (DENV) that caused an outbreak in Puerto Rico in 1994. The

examined oneet al.How such a takeover occurs at a molecular level, however, remains an open question. Manokaran For some pathogenic viruses, outbreaks occur when a new viral strain emerges and displaces the endemic strain.

Orchestrating a viral takeover

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