title: mRNA is an endogenous ligand for Toll-like receptor 3.
abstract: Toll-like receptors (TLRs) are the basic signaling receptors of the innate immune system. They are activated by molecules associated with pathogens or injured host cells and tissue. TLR3 has been shown to respond to double stranded (ds) RNA, a replication intermediary for many viruses. Here we present evidence that heterologous RNA released from or associated with necrotic cells or generated by in vitro transcription also stimulates TLR3 and induces immune activation. To assess RNA-mediated TLR3 activation, human embryonic kidney 293 cells stably expressing TLR3 and containing a nuclear factor-kappaB-dependent luciferase reporter were generated. Exposing these cells to in vitro transcribed RNA resulted in a TLR3-dependent induction of luciferase activity and interleukin-8 secretion. Treatment with in vitro transcribed mRNA activated nuclear factor-kappaB via TLR3 through a process that was dose-dependent and involved tyrosine phosphorylation. Furthermore, in vitro transcribed natural or 2'-fluoro-substituted mRNA induced the expression of TLR3, interferon regulatory factor-1, tumor necrosis factor-alpha, and interleukin-1 receptor-associated kinase-M mRNA in human dendritic cells (DCs). DCs responded to mRNA treatment by expressing activation markers, and this maturation was inhibited by antagonistic TLR3-specific antibody. Endogenous RNA released from or associated with necrotic cells also stimulated DCs, leading to interferon-alpha secretion, which could be abolished by pretreatment of necrotic cells with RNase. These results demonstrate that RNA, likely through secondary structure, is a potent host-derived activator of TLR3. This finding has potential physiologic relevance because RNA escaping from damaged tissue or contained within endocytosed cells could serve as an endogenous ligand for TLR3 that induces or otherwise modulates immune responses.
authors.affiliation: Division of Neurosurgery and Infectious Diseases, University of Pennsylvania, 3610 Hamilton Walk, Philadelphia, PA 19104, USA. kariko@mail.med.upenn.edu
authors.forename: Katalin
authors.initials: K
authors.lastname: Karikó
authors.forename: Houping
authors.initials: H
authors.lastname: Ni
authors.forename: John
authors.initials: J
authors.lastname: Capodici
authors.forename: Marc
authors.initials: M
authors.lastname: Lamphier
authors.forename: Drew
authors.initials: D
authors.lastname: Weissman
mesh_terms: D015152:Blotting, Northern; D002460:Cell Line; D003713:Dendritic Cells / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D053592:Interleukin-1 Receptor-Associated Kinases; D016209:Interleukin-8 / Q000378:metabolism; D008024:Ligands; D008156:Luciferases / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009336:Necrosis; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D012313:RNA / Q000737:chemistry; D012330:RNA, Double-Stranded / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000378:metabolism; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology; D051196:Toll-Like Receptor 3; D051193:Toll-Like Receptors; D014158:Transcription, Genetic; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry; D015854:Up-Regulation
chemical_list: D004791:Enzyme Inhibitors; D016209:Interleukin-8; D008024:Ligands; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; C495344:TLR3 protein, human; D051196:Toll-Like Receptor 3; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; D014443:Tyrosine; D012313:RNA; D019833:Genistein; D008156:Luciferases; D011494:Protein Kinases; D053592:Interleukin-1 Receptor-Associated Kinases; D019311:Staurosporine