{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T05:22:53Z","timestamp":1788931373063,"version":"build-2803163510"},"reference-count":38,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2020,10,18]],"date-time":"2020-10-18T00:00:00Z","timestamp":1602979200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000005","name":"U.S. Department of Defense","doi-asserted-by":"publisher","award":["PRMRP PR160117"],"award-info":[{"award-number":["PRMRP PR160117"]}],"id":[{"id":"10.13039\/100000005","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000738","name":"U.S. Department of Veterans Affairs","doi-asserted-by":"publisher","award":["I01BX003863"],"award-info":[{"award-number":["I01BX003863"]}],"id":[{"id":"10.13039\/100000738","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>The Zika virus (ZIKV), like other flaviviruses, produces several species of sub-genomic RNAs (sfRNAs) during infection, corresponding to noncoding RNA fragments of different lengths that result from the exonuclease degradation of the viral 3\u2032 untranslated region (UTR). Over the course of infection, these sfRNAs accumulate in the cell as a result of an incomplete viral genome degradation of the 3\u2032 UTR by the host 5\u2032 to 3\u2032 exoribonuclease, Xrn1. The halting of Xrn1 in the 3\u2032 UTR is due to two RNA pseudoknot structures in the 3\u2032 UTR, termed exoribonuclease-resistant RNA1 and 2 (xrRNA1&amp;2). Studies with related flaviviruses have shown that sfRNAs are important for pathogenicity and inhibiting both mosquito and mammalian host defense mechanisms. However, these investigations have not included ZIKV and there is very limited data addressing how sfRNAs impact infection in a whole animal model or specific tissues. In this study, we generate a sfRNA1-deficient ZIKV (X1) by targeted mutation in the xrRNA1 3\u2032 UTR structure. We find that the X1 virus lacks the production of the largest ZIKV sfRNA species, sfRNA1. Using the X1 virus to infect adult Ifnar1\u2212\/\u2212 mice, we find that while the lack of sfRNA1 does not alter ZIKV replication in the spleen, there is a significant reduction of ZIKV genome replication in the brain and placenta compared to wild-type ZIKV infection. Despite the attenuated phenotype of the X1 ZIKV, mice develop a robust neutralizing antibody response. We conclude that the targeted disruption of xrRNA1 results in tissue-specific attenuation while still supporting robust neutralizing antibody responses. Future studies will need to investigate the tissue-specific mechanisms by which ZIKV sfRNAs influence infection and may utilize targeted xrRNA mutations to develop novel attenuated flavivirus vaccine approaches.<\/jats:p>","DOI":"10.3390\/v12101177","type":"journal-article","created":{"date-parts":[[2020,10,18]],"date-time":"2020-10-18T21:26:06Z","timestamp":1603056366000},"page":"1177","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":10,"title":["Disruption of Zika Virus xrRNA1-Dependent sfRNA1 Production Results in Tissue-Specific Attenuated Viral Replication"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-7339-4675","authenticated-orcid":false,"given":"Hadrian","family":"Sparks","sequence":"first","affiliation":[{"name":"Department of Immunology &amp; Microbiology, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Brendan","family":"Monogue","sequence":"additional","affiliation":[{"name":"Department of Immunology &amp; Microbiology, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benjamin","family":"Akiyama","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Genetics and 4RNA BioScience Initiative, University of Colorado Denver School of Medicine, Aurora, CO 80045, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jeffrey","family":"Kieft","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Genetics and 4RNA BioScience Initiative, University of Colorado Denver School of Medicine, Aurora, CO 80045, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9305-2683","authenticated-orcid":false,"given":"J. David","family":"Beckham","sequence":"additional","affiliation":[{"name":"Department of Immunology &amp; Microbiology, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, USA"},{"name":"Department of Medicine, Division of Infectious Diseases, University of Colorado School of Medicine, Aurora, CO 80045, USA"},{"name":"Rocky Mountain Regional VA Medical Center, Aurora, CO 80045, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,10,18]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1148","DOI":"10.1126\/science.aah3963","article-title":"Zika virus produces noncoding RNAs using a multi-pseudoknot structure that confounds a cellular exonuclease","volume":"354","author":"Akiyama","year":"2016","journal-title":"Science"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"579","DOI":"10.1016\/j.chom.2008.10.007","article-title":"A highly structured, nuclease-resistant, noncoding RNA produced by flaviviruses is required for pathogenicity","volume":"4","author":"Pijlman","year":"2008","journal-title":"Cell Host Microbe"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"307","DOI":"10.1126\/science.1250897","article-title":"The structural basis of pathogenic subgenomic flavivirus RNA (sfRNA) production","volume":"344","author":"Chapman","year":"2014","journal-title":"Science"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1016\/j.vetmic.2013.04.026","article-title":"Japanese encephalitis virus non-coding RNA inhibits activation of interferon by blocking nuclear translocation of interferon regulatory factor 3","volume":"166","author":"Chang","year":"2013","journal-title":"Vet. Microbiol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"e01892","DOI":"10.7554\/eLife.01892","article-title":"RNA structures that resist degradation by Xrn1 produce a pathogenic Dengue virus RNA","volume":"3","author":"Chapman","year":"2014","journal-title":"Elife"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"10145","DOI":"10.1128\/JVI.00930-16","article-title":"Noncoding Subgenomic Flavivirus RNA Is Processed by the Mosquito RNA Interference Machinery and Determines West Nile Virus Transmission by Culex pipiens Mosquitoes","volume":"90","author":"Goertz","year":"2016","journal-title":"J. Virol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"322","DOI":"10.1016\/j.virol.2015.08.009","article-title":"Flavivirus sfRNA suppresses antiviral RNA interference in cultured cells and mosquitoes and directly interacts with the RNAi machinery","volume":"485","author":"Moon","year":"2015","journal-title":"Virology"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"13486","DOI":"10.1128\/JVI.01104-12","article-title":"Noncoding flavivirus RNA displays RNA interference suppressor activity in insect and Mammalian cells","volume":"86","author":"Schnettler","year":"2012","journal-title":"J. Virol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"9436","DOI":"10.1093\/nar\/gku657","article-title":"Induction and suppression of tick cell antiviral RNAi responses by tick-borne flaviviruses","volume":"42","author":"Schnettler","year":"2014","journal-title":"Nucleic Acids Res."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"5708","DOI":"10.1128\/JVI.00207-12","article-title":"West Nile virus noncoding subgenomic RNA contributes to viral evasion of the type I interferon-mediated antiviral response","volume":"86","author":"Schuessler","year":"2012","journal-title":"J. Virol."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Pallares, H.M., Costa Navarro, G.S., Villordo, S.M., Merwaiss, F., de Borba, L., Gonzalez Lopez Ledesma, M.M., Ojeda, D.S., Henrion-Lacritick, A., Morales, M.A., and Fabri, C. (2020). Zika Virus Subgenomic Flavivirus RNA Generation Requires Cooperativity between Duplicated RNA Structures That Are Essential for Productive Infection in Human Cells. J. Virol., 94.","DOI":"10.1128\/JVI.00343-20"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1016\/j.virol.2013.09.016","article-title":"Dengue virus subgenomic RNA induces apoptosis through the Bcl-2-mediated PI3k\/Akt signaling pathway","volume":"448","author":"Liu","year":"2014","journal-title":"Virology"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Weger-Lucarelli, J., Duggal, N.K., Bullard-Feibelman, K., Veselinovic, M., Romo, H., Nguyen, C., Ruckert, C., Brault, A.C., Bowen, R.A., and Stenglein, M. (2017). Development and Characterization of Recombinant Virus Generated from a New World Zika Virus Infectious Clone. J. Virol., 91.","DOI":"10.1128\/JVI.00172-17"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"15D-3","DOI":"10.1002\/9780471729259.mc15d03s31","article-title":"Propagation, quantification, detection, and storage of West Nile virus","volume":"31","author":"Brien","year":"2013","journal-title":"Curr. Protoc. Microbiol."},{"key":"ref_15","first-page":"471","article-title":"Superovulation strategies for 6 commonly used mouse strains","volume":"50","author":"Luo","year":"2011","journal-title":"J. Am. Assoc. Lab. Anim. Sci."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"pdb-prot5439","DOI":"10.1101\/pdb.prot5439","article-title":"Purification of RNA using TRIzol (TRI reagent)","volume":"2010","author":"Rio","year":"2010","journal-title":"Cold Spring Harb. Protoc."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"720","DOI":"10.1016\/j.chom.2016.03.010","article-title":"A Mouse Model of Zika Virus Pathogenesis","volume":"19","author":"Lazear","year":"2016","journal-title":"Cell Host Microbe"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"672","DOI":"10.1016\/j.chom.2018.04.003","article-title":"An Immunocompetent Mouse Model of Zika Virus Infection","volume":"23","author":"Gorman","year":"2018","journal-title":"Cell Host Microbe"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"2029","DOI":"10.1261\/rna.034330.112","article-title":"A noncoding RNA produced by arthropod-borne flaviviruses inhibits the cellular exoribonuclease XRN1 and alters host mRNA stability","volume":"18","author":"Moon","year":"2012","journal-title":"RNA"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"217","DOI":"10.1126\/science.aab3369","article-title":"Dengue subgenomic RNA binds TRIM25 to inhibit interferon expression for epidemiological fitness","volume":"350","author":"Manokaran","year":"2015","journal-title":"Science"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Donald, C.L., Brennan, B., Cumberworth, S.L., Rezelj, V.V., Clark, J.J., Cordeiro, M.T., Freitas de Oliveira Franca, R., Pena, L.J., Wilkie, G.S., and Da Silva Filipe, A. (2016). Full Genome Sequence and sfRNA Interferon Antagonist Activity of Zika Virus from Recife, Brazil. PLoS Negl. Trop. Dis., 10.","DOI":"10.1371\/journal.pntd.0005048"},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Bidet, K., Dadlani, D., and Garcia-Blanco, M.A. (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.","DOI":"10.1371\/journal.ppat.1004242"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"2908","DOI":"10.1099\/jgv.0.000613","article-title":"Interferon gamma modulation of disease manifestation and the local antibody response to alphavirus encephalomyelitis","volume":"97","author":"Baxter","year":"2016","journal-title":"J. Gen. Virol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"705","DOI":"10.1016\/j.chom.2016.03.008","article-title":"Type III Interferons Produced by Human Placental Trophoblasts Confer Protection against Zika Virus Infection","volume":"19","author":"Bayer","year":"2016","journal-title":"Cell Host Microbe"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"303","DOI":"10.1126\/science.1059742","article-title":"Interferon-gamma-mediated site-specific clearance of alphavirus from CNS neurons","volume":"293","author":"Binder","year":"2001","journal-title":"Science"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"366","DOI":"10.1016\/j.chom.2017.08.012","article-title":"Gestational Stage and IFN-lambda Signaling Regulate ZIKV Infection In Utero","volume":"22","author":"Jagger","year":"2017","journal-title":"Cell Host Microbe"},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Baxter, V.K., and Griffin, D.E. (2020). Interferon-Gamma Modulation of the Local T Cell Response to Alphavirus Encephalomyelitis. Viruses, 12.","DOI":"10.3390\/v12010113"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Caine, E.A., Jagger, B.W., and Diamond, M.S. (2018). Animal Models of Zika Virus Infection during Pregnancy. Viruses, 10.","DOI":"10.3390\/v10110598"},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Carbaugh, D.L., Zhou, S., Sanders, W., Moorman, N.J., Swanstrom, R., and Lazear, H.M. (2020). Two genetic differences between closely-related Zika virus strains determine pathogenic outcome in mice. J. Virol., 94.","DOI":"10.1128\/JVI.00618-20"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"676","DOI":"10.1038\/s41467-017-00737-8","article-title":"A single-dose live-attenuated vaccine prevents Zika virus pregnancy transmission and testis damage","volume":"8","author":"Shan","year":"2017","journal-title":"Nat. Commun."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"763","DOI":"10.1038\/nm.4322","article-title":"A live-attenuated Zika virus vaccine candidate induces sterilizing immunity in mouse models","volume":"23","author":"Shan","year":"2017","journal-title":"Nat. Med."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"92","DOI":"10.1016\/j.ebiom.2018.08.056","article-title":"A single-dose plasmid-launched live-attenuated Zika vaccine induces protective immunity","volume":"36","author":"Zou","year":"2018","journal-title":"EBioMedicine"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"334","DOI":"10.1093\/infdis\/jit546","article-title":"Comparison of the live attenuated yellow fever vaccine 17D-204 strain to its virulent parental strain Asibi by deep sequencing","volume":"209","author":"Beck","year":"2014","journal-title":"J. Infect. Dis."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"817","DOI":"10.1016\/j.vaccine.2007.11.082","article-title":"Dengue virus type 3 vaccine candidates generated by introduction of deletions in the 3\u2019 untranslated region (3\u2019-UTR) or by exchange of the DENV-3 3\u2019-UTR with that of DENV-4","volume":"26","author":"Blaney","year":"2008","journal-title":"Vaccine"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1169","DOI":"10.1080\/15476286.2015.1094599","article-title":"New hypotheses derived from the structure of a flaviviral Xrn1-resistant RNA: Conservation, folding, and host adaptation","volume":"12","author":"Kieft","year":"2015","journal-title":"RNA Biol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"11407","DOI":"10.1128\/JVI.01159-10","article-title":"RNA structures required for production of subgenomic flavivirus RNA","volume":"84","author":"Funk","year":"2010","journal-title":"J. Virol."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Villordo, S.M., Filomatori, C.V., Sanchez-Vargas, I., Blair, C.D., and Gamarnik, A.V. (2015). Dengue virus RNA structure specialization facilitates host adaptation. PLoS Pathog., 11.","DOI":"10.1371\/journal.ppat.1004604"},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Filomatori, C.V., Carballeda, J.M., Villordo, S.M., Aguirre, S., Pallares, H.M., Maestre, A.M., Sanchez-Vargas, I., Blair, C.D., Fabri, C., and Morales, M.A. (2017). Dengue virus genomic variation associated with mosquito adaptation defines the pattern of viral non-coding RNAs and fitness in human cells. PLoS Pathog., 13.","DOI":"10.1371\/journal.ppat.1006265"}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/12\/10\/1177\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T10:23:25Z","timestamp":1760178205000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/12\/10\/1177"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,10,18]]},"references-count":38,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2020,10]]}},"alternative-id":["v12101177"],"URL":"https:\/\/doi.org\/10.3390\/v12101177","relation":{"has-preprint":[{"id-type":"doi","id":"10.1101\/2020.09.10.281949","asserted-by":"object"}]},"ISSN":["1999-4915"],"issn-type":[{"value":"1999-4915","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,10,18]]}}}