The NS1 protein of influenza B virus binds 5’-triphosphorylated dsRNA to suppress RIG-I activation and the host antiviral response

The NS1 protein of influenza B virus binds 5’-triphosphorylated dsRNA to suppress RIG-I activation and the host antiviral response | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article The NS1 protein of influenza B virus binds 5’-triphosphorylated dsRNA to suppress RIG-I activation and the host antiviral response Gaetano Montelione, Ryan Woltz, Brandon Schweibenz, Li Chung Ma, and 11 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3874434/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted You are reading this latest preprint version Abstract Influenza A and B viruses overcome the host antiviral response to cause a contagious and often severe human respiratory disease. Here, integrative structural biology and biochemistry studies on non-structural protein 1 of influenza B virus (NS1B) reveal a previously unrecognized viral mechanism for innate immune evasion. Conserved basic groups of its C-terminal domain (NS1B-CTD) bind 5’-triphosphorylated double-stranded RNA (5’ppp-dsRNA), the primary pathogen-associated feature that activates the host retinoic acid-inducible gene I protein (RIG-I) to initiate interferon synthesis and the cellular antiviral response. Like RIG-I, NS1B-CTD preferentially binds blunt-end 5’ppp-dsRNA. NS1B-CTD also competes with RIG-I for binding 5’ppp-dsRNA, and thus suppresses activation of RIG-I’s ATPase activity. Although the NS1B N-terminal domain also binds dsRNA, it utilizes a different binding mode and lacks 5’ppp-dsRNA end preferences. In cells infected with wild-type influenza B virus, RIG-I activation is inhibited. In contrast, RIG-I activation and the resulting phosphorylation of transcription factor IRF-3 are not inhibited in cells infected with a mutant virus encoding NS1B with a R208A substitution it its CTD that eliminates its 5’ppp-dsRNA binding activity. These results reveal a novel mechanism in which NS1B binds 5’ppp-dsRNA to inhibit the RIG-I antiviral response during influenza B virus infection, and open the door to new avenues for antiviral drug discovery. Biological sciences/Structural biology/SAXS Biological sciences/Structural biology/NMR spectroscopy/Solution-state NMR Biological sciences/Structural biology/Molecular modelling Biological sciences/Microbiology/Virology Biological sciences/Biochemistry/RNA Full Text Additional Declarations Yes there is potential Competing Interest. GTM is a founder and consultant of Nexomics Biosciences, Inc. This does not represent a conflict of interest for this study. Cite Share Download PDF Status: Under Review Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-3874434","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":268846543,"identity":"a0cc2acc-cc84-4a3b-99a5-735e6ae57f14","order_by":0,"name":"Gaetano Montelione","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAtklEQVRIiWNgGAWjYPCCGjkGBsYGBgY24pSDlB4zJlkLc2IDmE2MFt323uMPPvxiS98+u7mB4UPZYcJazM6cS2yc2SeTO+fOwQbGGeeI0XIjx7CZt4ctd4ZEYgMzbxuxWv72MKdLgLT8JVoLww/mBLAWRqK0nDljOLO34ZjhDJmDDQd7zqUToeV4j8GHH39q5CWk2x8++FFmTVgLGDC2AQkJBoYDRKoHgT8QLaNgFIyCUTAKsAIATVg/+BO/j3oAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0002-9440-3059","institution":"Rensselaer Polytechnic Institute","correspondingAuthor":true,"prefix":"","firstName":"Gaetano","middleName":"","lastName":"Montelione","suffix":""},{"id":268846544,"identity":"e5459967-ecfb-4746-9768-76f50380f230","order_by":1,"name":"Ryan Woltz","email":"","orcid":"https://orcid.org/0000-0002-0446-2478","institution":"University of California, Davis","correspondingAuthor":false,"prefix":"","firstName":"Ryan","middleName":"","lastName":"Woltz","suffix":""},{"id":268846545,"identity":"c3907c6f-cc27-4143-a3fb-04ea88892e4d","order_by":2,"name":"Brandon Schweibenz","email":"","orcid":"","institution":"Rutgers Medical School","correspondingAuthor":false,"prefix":"","firstName":"Brandon","middleName":"","lastName":"Schweibenz","suffix":""},{"id":268846546,"identity":"c9401f9b-ebdd-4385-8e12-ee3a95452526","order_by":3,"name":"Li Chung Ma","email":"","orcid":"","institution":"Rutgers, The State Univ of New Jersey","correspondingAuthor":false,"prefix":"","firstName":"Li","middleName":"Chung","lastName":"Ma","suffix":""},{"id":268846547,"identity":"54c63e91-4aec-48a0-b8ea-5427067a8302","order_by":4,"name":"Rachel John","email":"","orcid":"","institution":"Rutgers, The State Univ of New Jersey","correspondingAuthor":false,"prefix":"","firstName":"Rachel","middleName":"","lastName":"John","suffix":""},{"id":268846548,"identity":"540baf2b-4528-4376-8d6d-ca5c0720a7f8","order_by":5,"name":"Sergey Vorobiev","email":"","orcid":"","institution":"Rutgers, The State Univ of New Jersey","correspondingAuthor":false,"prefix":"","firstName":"Sergey","middleName":"","lastName":"Vorobiev","suffix":""},{"id":268846549,"identity":"24103253-25a7-468a-a952-d6f878bc4548","order_by":6,"name":"Mihai Solotchi","email":"","orcid":"","institution":"Rutgers Medical School","correspondingAuthor":false,"prefix":"","firstName":"Mihai","middleName":"","lastName":"Solotchi","suffix":""},{"id":268846550,"identity":"8ac1472b-a770-4df1-af99-ed4e17b34337","order_by":7,"name":"Smita Patel","email":"","orcid":"https://orcid.org/0000-0002-2523-4933","institution":"Rutgers Medical School","correspondingAuthor":false,"prefix":"","firstName":"Smita","middleName":"","lastName":"Patel","suffix":""},{"id":268846551,"identity":"89547c86-fc98-411e-98a4-e0e5484a4635","order_by":8,"name":"Susan Tsutakawa","email":"","orcid":"https://orcid.org/0000-0002-4918-4571","institution":"Lawrence Berkeley National Laboratory","correspondingAuthor":false,"prefix":"","firstName":"Susan","middleName":"","lastName":"Tsutakawa","suffix":""},{"id":268846552,"identity":"1c32fc9b-d556-446b-b8bd-dc36bd6c71fd","order_by":9,"name":"Greg Hura","email":"","orcid":"","institution":"Lawrence Berkeley National Laboratory","correspondingAuthor":false,"prefix":"","firstName":"Greg","middleName":"","lastName":"Hura","suffix":""},{"id":268846553,"identity":"22ba1e63-5dd3-483d-a1b9-1a46686230f7","order_by":10,"name":"Chen Zhao","email":"","orcid":"","institution":"University of Texas at Austin","correspondingAuthor":false,"prefix":"","firstName":"Chen","middleName":"","lastName":"Zhao","suffix":""},{"id":268846554,"identity":"1d38f4a2-81b7-4c18-a693-142a3ba35dee","order_by":11,"name":"Robert Krug","email":"","orcid":"","institution":"University of Texas at Austin","correspondingAuthor":false,"prefix":"","firstName":"Robert","middleName":"","lastName":"Krug","suffix":""},{"id":268846555,"identity":"1ff7aa71-30cf-426b-b0be-9795b1aec3c7","order_by":12,"name":"Ben Shurina","email":"","orcid":"","institution":"Rensselaer Polytechnic Institute","correspondingAuthor":false,"prefix":"","firstName":"Ben","middleName":"","lastName":"Shurina","suffix":""},{"id":268846556,"identity":"cdd625ee-2a05-495f-b912-9371debea785","order_by":13,"name":"G.V.T. 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Here, integrative structural biology and biochemistry studies on non-structural protein 1 of influenza B virus (NS1B) reveal a previously unrecognized viral mechanism for innate immune evasion. Conserved basic groups of its C-terminal domain (NS1B-CTD) bind 5’-triphosphorylated double-stranded RNA (5’ppp-dsRNA), the primary pathogen-associated feature that activates the host retinoic acid-inducible gene I protein (RIG-I) to initiate interferon synthesis and the cellular antiviral response. Like RIG-I, NS1B-CTD preferentially binds blunt-end 5’ppp-dsRNA. NS1B-CTD also competes with RIG-I for binding 5’ppp-dsRNA, and thus suppresses activation of RIG-I’s ATPase activity. Although the NS1B N-terminal domain also binds dsRNA, it utilizes a different binding mode and lacks 5’ppp-dsRNA end preferences. In cells infected with wild-type influenza B virus, RIG-I activation is inhibited. In contrast, RIG-I activation and the resulting phosphorylation of transcription factor IRF-3 are not inhibited in cells infected with a mutant virus encoding NS1B with a R208A substitution it its CTD that eliminates its 5’ppp-dsRNA binding activity. These results reveal a novel mechanism in which NS1B binds 5’ppp-dsRNA to inhibit the RIG-I antiviral response during influenza B virus infection, and open the door to new avenues for antiviral drug discovery.","manuscriptTitle":"The NS1 protein of influenza B virus binds 5’-triphosphorylated dsRNA to suppress RIG-I activation and the host antiviral response","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-02-01 01:32:09","doi":"10.21203/rs.3.rs-3874434/v1","editorialEvents":[],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"nature-communications","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"NCOMMS","sideBox":"Learn more about [Nature Communications](http://www.nature.com/ncomms/)","snPcode":"","submissionUrl":"https://mts-ncomms.nature.com/","title":"Nature Communications","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature Communications","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"b042494b-c9ae-4b6a-9a44-e827e203cc8c","owner":[],"postedDate":"February 1st, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[{"id":28325187,"name":"Biological sciences/Structural biology/SAXS"},{"id":28325188,"name":"Biological sciences/Structural biology/NMR spectroscopy/Solution-state NMR"},{"id":28325189,"name":"Biological sciences/Structural biology/Molecular modelling"},{"id":28325190,"name":"Biological sciences/Microbiology/Virology"},{"id":28325191,"name":"Biological sciences/Biochemistry/RNA"}],"tags":[],"updatedAt":"2024-02-01T01:32:09+00:00","versionOfRecord":[],"versionCreatedAt":"2024-02-01 01:32:09","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-3874434","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-3874434","identity":"rs-3874434","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}