{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,8,7]],"date-time":"2026-08-07T16:48:58Z","timestamp":1786121338884,"version":"build-2736575974"},"posted":{"date-parts":[[2026,7,3]]},"group-title":"Immunology","reference-count":68,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,7,3]],"date-time":"2026-07-03T00:00:00Z","timestamp":1783036800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/publicdomain\/zero\/1.0\/"}],"funder":[{"award":["HHSN261200800001E"],"award-info":[{"award-number":["HHSN261200800001E"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,8,5]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                <jats:p>Interleukin-27 (IL-27) is an anti-HIV cytokine that induces 14 novel microRNAs (miRNAs) in primary CD4(+) T cells. We previously reported that transfection of two of these miRNA mimics, miRTC10 and miRTC14 into human primary macrophages, differentially induced interferon (IFN)-\u03b12, -\u03b18, -\u03b113, and -\u03bb1 expression. However, the mechanism underlying this activation remains unclear. Here, we demonstrate that miRTC14 does not directly target canonical IFN-regulatory genes but instead engages with pyruvate carboxylase (PC) and laboratory of genetics and physiology-2 (LGP2\/DHX58) as direct binding partner proteins. Functional analysis revealed that miRTC14 transfection induces IFN expression by more than 100-fold (p&lt;0.001), whereas PC or LGP2 depletion by siRNAs markedly attenuated this response (50\u2013100-fold reduction, p&lt;0.01). Reconstitution of PC and LGP2 in deficient HEK293 cells restored miRTC14-driven IFN induction. Notably, IFN activation depended on sequence features at the duplex termini. However, PC\u2013miRTC14\u2013LGP2 axis activates TBK1-dependent phosphorylation of IRF3\/7, similarly to canonical RNA sensors (RIG-I\/MDA5), but this process induced differential IFN subtype. These findings establish PC as a miRNA-binding protein and define a previously unrecognized RNA-sensing mechanism, linking metabolic enzymes to RNA sequence-dependent innate immunity.<\/jats:p>\n                <jats:sec>\n                  <jats:title>GRAPHICAL ABSTRACT<\/jats:title>\n                  <jats:fig id=\"ufig1\" position=\"float\" orientation=\"portrait\" fig-type=\"figure\">\n                    <jats:label>Graphical Abstract:<\/jats:label>\n                    <jats:caption>\n                      <jats:title>microRNA sequence\u2013dependent activation of innate immunity by pyruvate carboxylase (PC).<\/jats:title>\n                      <jats:p>\n                        PC\u2013miRTC14\u2013LGP2 axis induces TBK1 phosphorylation via an as-yet-unidentified adapter(s). The activated TBK1 (p-TBK1) subsequently phosphorylates IRF3 and\/or IRF7 leading to selective IFN subtypes (IFN-\u03b12, IFN-\u03b18, IFN-\u03b113, and IFN-\u03bb1). (Created in BioRender. Kariyawasam, U. (2027)\n                        <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" ext-link-type=\"uri\" xlink:href=\"https:\/\/BioRender.com\/u82s04p\">https:\/\/BioRender.com\/u82s04p<\/jats:ext-link>\n                        )\n                      <\/jats:p>\n                    <\/jats:caption>\n                    <jats:graphic xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" xlink:href=\"735367v2_ufig1\" position=\"float\" orientation=\"portrait\"\/>\n                  <\/jats:fig>\n                <\/jats:sec>","DOI":"10.64898\/2026.06.29.735367","type":"posted-content","created":{"date-parts":[[2026,7,4]],"date-time":"2026-07-04T03:31:25Z","timestamp":1783135885000},"source":"Crossref","is-referenced-by-count":0,"title":["Beyond Metabolism: Pyruvate Carboxylase Acts as a Sequence-Selective Small RNA Sensor Orchestrating Antiviral Responses"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-2226-9399","authenticated-orcid":false,"given":"Udeshika","family":"Kariyawasam","sequence":"first","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Suranjana","family":"Goswami","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ming","family":"Hao","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rosana","family":"Wiscovitch-Russo","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Qian","family":"Chen","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jun","family":"Yang","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ju","family":"Qu","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mayra","family":"Marquez","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hongyan","family":"Sui","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Weizhong","family":"Chang","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-0910-5297","authenticated-orcid":false,"given":"Tomozumi","family":"Imamichi","sequence":"additional","affiliation":[{"name":"Laboratory of Human Retrovirology and Immunoinformatics, Frederick National Laboratory for Cancer Research, Frederick, MD, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026080621402269000_2026.06.29.735367v2.1","doi-asserted-by":"publisher","DOI":"10.1016\/S1357-2725(97)00147-7"},{"key":"2026080621402269000_2026.06.29.735367v2.2","doi-asserted-by":"publisher","DOI":"10.1042\/bj3400001"},{"key":"2026080621402269000_2026.06.29.735367v2.3","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbamcr.2010.03.018"},{"key":"2026080621402269000_2026.06.29.735367v2.4","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1186\/s40170-021-00256-7","article-title":"Pyruvate carboxylase and cancer progression","volume":"9","year":"2021","journal-title":"Cancer Metab"},{"key":"2026080621402269000_2026.06.29.735367v2.5","doi-asserted-by":"crossref","unstructured":"Stacpoole, P.W . (2017) Therapeutic Targeting of the Pyruvate Dehydrogenase Complex\/Pyruvate Dehydrogenase Kinase (PDC\/PDK) Axis in Cancer. J Natl Cancer Inst, 109.","DOI":"10.1093\/jnci\/djx071"},{"key":"2026080621402269000_2026.06.29.735367v2.6","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2021.08.031"},{"key":"2026080621402269000_2026.06.29.735367v2.7","doi-asserted-by":"publisher","DOI":"10.1111\/1753-0407.70050"},{"key":"2026080621402269000_2026.06.29.735367v2.8","doi-asserted-by":"crossref","first-page":"22002","DOI":"10.1038\/srep22002","article-title":"Pyruvate Carboxylase Activates the RIG-I-like Receptor-Mediated Antiviral Immune Response by Targeting the MAVS signalosome","volume":"6","year":"2016","journal-title":"Scientific Reports"},{"key":"2026080621402269000_2026.06.29.735367v2.9","doi-asserted-by":"publisher","DOI":"10.1007\/s00109-018-1622-0"},{"key":"2026080621402269000_2026.06.29.735367v2.10","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1002124"},{"key":"2026080621402269000_2026.06.29.735367v2.11","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-immunol-032713-120156"},{"key":"2026080621402269000_2026.06.29.735367v2.12","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.0912986107"},{"key":"2026080621402269000_2026.06.29.735367v2.13","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-immunol-042617-053309"},{"key":"2026080621402269000_2026.06.29.735367v2.14","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1921861117"},{"key":"2026080621402269000_2026.06.29.735367v2.15","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkp059"},{"key":"2026080621402269000_2026.06.29.735367v2.16","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M109.007179"},{"key":"2026080621402269000_2026.06.29.735367v2.17","doi-asserted-by":"crossref","unstructured":"O\u2019Brien, J. , Hayder, H. , Zayed, Y. and Peng, C . (2018) Overview of MicroRNA Biogenesis, Mechanisms of Actions, and Circulation. Frontiers in Endocrinology, Volume 9 - 2018.","DOI":"10.3389\/fendo.2018.00402"},{"key":"2026080621402269000_2026.06.29.735367v2.18","doi-asserted-by":"crossref","first-page":"e1627","DOI":"10.1002\/wrna.1627","article-title":"microRNA strand selection: Unwinding the rules","volume":"12","year":"2021","journal-title":"Wiley Interdiscip Rev RNA"},{"key":"2026080621402269000_2026.06.29.735367v2.19","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2009.01.002"},{"key":"2026080621402269000_2026.06.29.735367v2.20","doi-asserted-by":"publisher","DOI":"10.1093\/jb\/mvq113"},{"key":"2026080621402269000_2026.06.29.735367v2.21","doi-asserted-by":"publisher","DOI":"10.1158\/0008-5472.CAN-10-2010"},{"key":"2026080621402269000_2026.06.29.735367v2.22","doi-asserted-by":"publisher","DOI":"10.1038\/nrd.2016.246"},{"key":"2026080621402269000_2026.06.29.735367v2.23","doi-asserted-by":"crossref","first-page":"e10213","DOI":"10.1002\/btm2.10213","article-title":"Cytosolic delivery of nucleic acids: The case of ionizable lipid nanoparticles","volume":"6","year":"2021","journal-title":"Bioeng Transl Med"},{"key":"2026080621402269000_2026.06.29.735367v2.24","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkt844"},{"key":"2026080621402269000_2026.06.29.735367v2.25","doi-asserted-by":"publisher","DOI":"10.1097\/qai.0000000000002565"},{"key":"2026080621402269000_2026.06.29.735367v2.26","doi-asserted-by":"crossref","unstructured":"Imamichi, T. , Goswami, S. , Hu, X. , Laverdure, S. , Yang, J. , Qiu, J. , Chen, Q. , Sherman, B.T. and Chang, W . (2021) MicroRNA Profiles in Monocyte-Derived Macrophages Generated by Interleukin-27 and Human Serum: Identification of a Novel HIV-Inhibiting and Autophagy-Inducing MicroRNA. Int J Mol Sci, 22.","DOI":"10.3390\/ijms22031290"},{"key":"2026080621402269000_2026.06.29.735367v2.27","doi-asserted-by":"publisher","DOI":"10.1084\/jem.20120572"},{"key":"2026080621402269000_2026.06.29.735367v2.28","doi-asserted-by":"crossref","first-page":"1550699","DOI":"10.3389\/fimmu.2025.1550699","article-title":"Interleukin-27-polarized HIV-resistant M2 macrophages are a novel subtype of macrophages that express distinct antiviral gene profiles in individual cells: implication for the antiviral effect via different mechanisms in the individual cell-dependent manner","volume":"16","year":"2025","journal-title":"Front Immunol"},{"key":"2026080621402269000_2026.06.29.735367v2.29","doi-asserted-by":"publisher","DOI":"10.1089\/aid.1996.12.191"},{"key":"2026080621402269000_2026.06.29.735367v2.30","doi-asserted-by":"publisher","DOI":"10.1006\/meth.1997.0487"},{"key":"2026080621402269000_2026.06.29.735367v2.31","doi-asserted-by":"crossref","unstructured":"Sui, H. , Chen, Q. , Yang, J. , Srirattanapirom, S. and Imamichi, T . (2022) Manganese enhances DNA- or RNA-mediated innate immune response by inducing phosphorylation of TANK-binding kinase 1. iScience, 25.","DOI":"10.2139\/ssrn.4063315"},{"key":"2026080621402269000_2026.06.29.735367v2.32","unstructured":"Green, M.R. and Sambrook, J . (2012) Molecular cloning. A Laboratory Manual 4th, 448."},{"key":"2026080621402269000_2026.06.29.735367v2.33","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gku1189"},{"key":"2026080621402269000_2026.06.29.735367v2.34","doi-asserted-by":"crossref","first-page":"100890","DOI":"10.1016\/j.mex.2020.100890","article-title":"A pull-down assay using DNA\/RNA-conjugated beads with a customized competition strategy: An effective approach to identify DNA\/RNA binding proteins","volume":"7","year":"2020","journal-title":"MethodsX"},{"key":"2026080621402269000_2026.06.29.735367v2.35","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkz285"},{"key":"2026080621402269000_2026.06.29.735367v2.36","unstructured":"Chen, W . Teaching Bio-Layer Interferometry: Protein\u2013Protein Versus Protein\u2013Small Molecule Interaction Analysis on an Octet R8. Biochemistry and Molecular Biology Education, n\/a."},{"key":"2026080621402269000_2026.06.29.735367v2.37","doi-asserted-by":"publisher","DOI":"10.1038\/S41586-024-07487-W"},{"key":"2026080621402269000_2026.06.29.735367v2.38","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btz184"},{"key":"2026080621402269000_2026.06.29.735367v2.39","doi-asserted-by":"crossref","first-page":"W241","DOI":"10.1093\/nar\/gkx236","article-title":"mCSM\u2013NA: predicting the effects of mutations on protein\u2013nucleic acids interactions","volume":"45","year":"2017","journal-title":"Nucleic Acids Res"},{"key":"2026080621402269000_2026.06.29.735367v2.40","doi-asserted-by":"crossref","first-page":"5560","DOI":"10.3390\/ijms21155560","article-title":"PremPRI: predicting the effects of missense mutations on protein\u2013RNA interactions","volume":"21","year":"2020","journal-title":"International journal of molecular sciences"},{"key":"2026080621402269000_2026.06.29.735367v2.41","doi-asserted-by":"publisher","DOI":"10.1016\/j.jbc.2024.105711"},{"key":"2026080621402269000_2026.06.29.735367v2.42","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M900818200"},{"key":"2026080621402269000_2026.06.29.735367v2.43","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gky427"},{"key":"2026080621402269000_2026.06.29.735367v2.44","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-0716-3441-7_5"},{"key":"2026080621402269000_2026.06.29.735367v2.45","doi-asserted-by":"crossref","first-page":"bbag109","DOI":"10.1093\/bib\/bbag109","article-title":"Differentiation of RNA-protein docking structures through molecular dynamics simulation and machine learning methods","volume":"27","year":"2026","journal-title":"Brief. Bioinform"},{"key":"2026080621402269000_2026.06.29.735367v2.46","doi-asserted-by":"publisher","DOI":"10.1002\/prot.22570"},{"key":"2026080621402269000_2026.06.29.735367v2.47","first-page":"116","article-title":"IL-27-inducible novel microRNA, hsa-mir-7705, predominantly elicits IFN-\u03b1 from human monocyte-derived macrophages in an RNA sequence and structure dependent manner: Mo-P11-12","volume":"100","year":"2017","journal-title":"Cytokine"},{"key":"2026080621402269000_2026.06.29.735367v2.48","doi-asserted-by":"publisher","DOI":"10.1099\/0022-1317-79-7-1647"},{"key":"2026080621402269000_2026.06.29.735367v2.49","doi-asserted-by":"publisher","DOI":"10.1038\/298859a0"},{"key":"2026080621402269000_2026.06.29.735367v2.50","doi-asserted-by":"publisher","DOI":"10.1016\/0022-2836(85)90401-2"},{"key":"2026080621402269000_2026.06.29.735367v2.51","doi-asserted-by":"publisher","DOI":"10.1038\/s41580-021-00430-1"},{"key":"2026080621402269000_2026.06.29.735367v2.52","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.80.10.5059-5064.2006"},{"key":"2026080621402269000_2026.06.29.735367v2.53","doi-asserted-by":"crossref","unstructured":"Peterson, S.M. , Thompson, J.A. , Ufkin, M.L. , Sathyanarayana, P. , Liaw, L. and Congdon, C.B . (2014) Common features of microRNA target prediction tools. Frontiers in Genetics, Volume 5 - 2014.","DOI":"10.3389\/fgene.2014.00023"},{"key":"2026080621402269000_2026.06.29.735367v2.54","doi-asserted-by":"crossref","unstructured":"Sherman, B.T. , Panzade, G. , Dotrang, T. , Hao, M. , Xu, L. , Li, X. , Baseler, M.W. , Lane, H.C. , Imamichi, T. and Chang, W . (2026) DAVID: a web server for functional annotation and functional enrichment analysis of gene lists (2025 update). Nucleic Acids Research, gkag470.","DOI":"10.1093\/nar\/gkag470"},{"key":"2026080621402269000_2026.06.29.735367v2.55","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btr661"},{"key":"2026080621402269000_2026.06.29.735367v2.56","doi-asserted-by":"publisher","DOI":"10.1016\/j.molimm.2019.08.007"},{"key":"2026080621402269000_2026.06.29.735367v2.57","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-biophys-083012-130404"},{"key":"2026080621402269000_2026.06.29.735367v2.58","doi-asserted-by":"publisher","DOI":"10.1172\/JCI11948"},{"key":"2026080621402269000_2026.06.29.735367v2.59","doi-asserted-by":"crossref","first-page":"894","DOI":"10.1007\/s000180050482","article-title":"Cathepsin A\/protective protein: an unusual lysosomal multifunctional protein","volume":"56","year":"1999","journal-title":"Cellular and Molecular Life Sciences CMLS"},{"key":"2026080621402269000_2026.06.29.735367v2.60","doi-asserted-by":"crossref","first-page":"1081","DOI":"10.1007\/s12551-021-00902-w","article-title":"DICER: structure, function, and regulation","volume":"13","year":"2021","journal-title":"Biophysical reviews"},{"key":"2026080621402269000_2026.06.29.735367v2.61","doi-asserted-by":"publisher","DOI":"10.1111\/j.1742-4658.2008.06411.x"},{"key":"2026080621402269000_2026.06.29.735367v2.62","doi-asserted-by":"publisher","DOI":"10.1038\/nature03868"},{"key":"2026080621402269000_2026.06.29.735367v2.63","doi-asserted-by":"publisher","DOI":"10.1042\/BCJ20160759"},{"key":"2026080621402269000_2026.06.29.735367v2.64","doi-asserted-by":"publisher","DOI":"10.15252\/embj.201797479"},{"key":"2026080621402269000_2026.06.29.735367v2.65","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.00640-14"},{"key":"2026080621402269000_2026.06.29.735367v2.66","doi-asserted-by":"crossref","unstructured":"Liu, R. , Niu, Y. , Qi, Y. , Li, H. , Yang, G. and Shan, S . (2022) Transcriptome analysis identifies LGP2 as an MDA5-mediated signaling activator following spring viremia of carp virus infection in common carp (Cyprinus carpio L.). Frontiers in Immunology, Volume 13 - 2022.","DOI":"10.3389\/fimmu.2022.1019872"},{"key":"2026080621402269000_2026.06.29.735367v2.67","doi-asserted-by":"publisher","DOI":"10.1016\/j.cyto.2015.02.010"},{"key":"2026080621402269000_2026.06.29.735367v2.68","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0064202"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.64898\/2026.06.29.735367","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,8,7]],"date-time":"2026-08-07T04:41:04Z","timestamp":1786077664000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.64898\/2026.06.29.735367"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,3]]},"references-count":68,"URL":"https:\/\/doi.org\/10.64898\/2026.06.29.735367","relation":{},"subject":[],"published":{"date-parts":[[2026,7,3]]},"subtype":"preprint"}}