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Project 20
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Project 20

Gunther Hartmann, Martin Schlee

Activation modes of RIG-I

Institute for Clinical Chemistry and Pharmacology, University of Bonn

Publications

Brief description in German:
Der Typ-I-IFN induzierende Immunrezeptor RIG-I detektiert virale RNA im Zytosol und hat eine zentrale Bedeutung für die antivirale Immunantwort gegen die meisten RNA-Viren in Vertebraten. Nach derzeitigen Erkenntnissen werden zwei Arten von Liganden durch RIG-I erkannt: (I) Kurze doppelsträngige (ds) RNA mit 5’Triphosphat am terminalen Basenpaar oder (II) sehr lange dsRNA ohne terminale Modifikationen. Durch Zellen mit RIG-I-Mutanten, die nur Typ II-Liganden erkennen, soll die biologische Relevanz von Typ II-Liganden bei viralen und bakteriellen Infektionen analysiert werden. Desweiteren soll in zellulären K.O.-Modellen von RIG-I-Kofaktoren oder Kompetitoren deren Einfluss auf die RNA-Erkennung durch RIG-I analysiert werden.

Summary
The type I IFN inducing innate immune receptor RIG-I detects and is essential for the initiation of immune responses against RNA viruses in vertebrates. In the last funding period we characterized the interaction of RIG-I with its ligand (5’triphosphorylated double-stranded RNA; ppp-dsRNA) by introduction of RNA backbone modifications, crystallization and structure guided mutational analysis of RIG-I.

In this way, we identified the interacting amino acids in the RIG-I C-terminal domain (CTD) responsible for ligand requirements (5’triphosphate, blunt end, dsRNA, as defined by us previously). Furthermore, we observed that RNA of intracellular bacteria activates RIG-I during infection.

According to the accepted literature RIG-I appears to possess two different activation modes: Recognition of short ppp-dsRNA (class I) and non-modified long double stranded RNA (class II). By exploiting our knowledge of the critical RIG-I-ligand interaction in the RIG-I CTD for ppp-dsRNA (class I) recognition and using a new knock-out technology (CRISPR), we aim at developing a cellular knock-out/knock-in system able to determine the RNA species (class I or II RIG-I ligands) critically involved in RIG-I stimulation during infection by viruses or bacteria.

Additionally, we want to determine the impact of RIG-I CTD binding co-activators and cytosolic RNA receptors, which might act as competitors, on RIG-I activation and ligand requirements. Using a cellular FRET based RIG-I co-activator interaction assay we aim at identifying viral proteins targeting the RIG-I/co-activator interaction.

List of publications resulting from the project

Schuberth-Wagner C, Ludwig J, Bruder AK, Herzner AM, Zillinger T, Goldeck M, Schmidt T, Schmid-Burgk JL, Kerber R, Wolter S, Stümpel JP, Roth A, Bartok E, Drosten C, Coch C, Hornung V, Barchet W, Kümmerer BM, Hartmann G, Schlee M. A Conserved Histidine in the RNA Sensor RIG-I Controls Immune Tolerance to N1-2'O-Methylated Self RNA. Immunity 43(1):41-51 (2015)

Herzner AM, Hagmann CA, Goldeck M, Wolter S, Kübler K, Wittmann S, Gramberg T, Andreeva L, Hopfner KP, Mertens C, Zillinger T, Jin T, Xiao TS, Bartok E, Coch C, Ackermann D, Hornung V, Ludwig J, Barchet W, Hartmann G, Schlee M. Sequence-specific activation of the DNA sensor cGAS by Y-form DNA structures as found in primary HIV-1 cDNA. Nat Immunol.16(10):1025-33 (2015)

Goubau D, Schlee M*, Deddouche S, Pruijssers AJ, Zillinger T,  Goldeck M,  Schuberth C, Van der Veen AG, Fujimura T, Rehwinkel J, Iskarpatyoti JA, Barchet W, Ludwig J,  Dermody TS, Hartmann G &  Reis e Sousa C. Antiviral immunity via RIG-I-mediated recognition of RNA bearing 5'-diphosphates Nature. 514 (7522), 372-5, *shared first author (2014)

Hagmann CA, Herzner AM, Abdullah Z,  Zillinger T, Jakobs C,  Schuberth C, Coch C, Higgins PG, Wisplinghoff W, Barchet W, Hornung V,  Hartmann G*, Schlee M. RIG-I Detects Triphosphorylated RNA of Listeria monocytogenes during Infection in Non-Immune Cells. PLoS ONE 8: e62872. doi:10.1371/ journal.pone.0062872, *shared last author (2013)

Abdullah Z, Schlee M*, Roth S, Abu Mraheil M, Barchet W, Böttcher J, Hain T, Geiger S, Hayakawa Y, Fritz JH, Civril F, Hopfner K-P,  Kurts C,  Ruland J, Hartmann G, Chakraborty T and Knolle PA. RIG-I detects infection with live Listeria by sensing secreted bacterial nucleic acids. The EMBO Journal 31, 4153 – 4164. *shared first author (2012)

Dann A , Poeck H, Croxford AL, Gaupp S, Kierdorf K, Knust M, Pfeifer D, Maihoefer C, Endres S, Kalinke U, Meuth SG, Wiendl H, Knobeloch KP, Akira S, Waisman A, Hartmann G, Prinz M. Systemic activation of RIG-I-like helicases limits Th1/Th17-mediated autoimmunity in the CNS. Nature Neuroscience, 15:98-106 (2012)

Ebert G, Poeck H, Lucifora J, Baschuk N, Esser K, Esposito I, Hartmann G, Protzer U. 5' Triphosphorylated small interfering RNAs control replication of hepatitis B virus and induce an interferon response in human liver cells and mice. Gastroenterology, 141:696-706 (2011)

Wang Y, Ludwig J, Schuberth C, Goldeck M, Schlee M, Li H, Juranek S, Sheng G, Micura R, Tuschl T°, Hartmann G°, Patel DJ°. Structural and functional insights into 5'-ppp RNA pattern recognition by the innate immune receptor RIG-I. Nature Structural & Molecular Biology, 17:781-7, °shared corresponding authors (2010)

Schlee M, Roth A, Hornung V, Hagmann CA, Wimmenauer V, Barchet W, Coch C, Janke M, Mihailovic A, Wardle G, Juranek S, Kato H, Kawai T, Poeck H, Fitzgerald KA, Takeuchi O, Akira S, Tuschl T, Latz E, Ludwig J, Hartmann G. Recognition of 5′ Triphosphate by RIG-I Helicase Requires Short Blunt Double-Stranded RNA as Contained in Panhandle of Negative-Strand Virus. Immunity. 31: 25-34. (2009)

Poeck H, Bscheider M, Gross O, Finger K, Roth S, Rebsamen M, Hannesschläger N, Schlee M, Rothenfusser S, Barchet W, Kato H, Akira S, Inoue S, Endres S, Peschel C, Hartmann G, Hornung V, Ruland J. Recognition of RNA virus by RIG-I results in activation of CARD9 and inflammasome signaling for interleukin 1 beta production. Nat Immunol. 11(1):63-9 (2010)

Poeck H, Besch R, Maihoefer C, Renn M, Tormo D, Morskaya SS, Kirschnek S, Gaffal E, Landsberg J, Hellmuth J, Schmidt A, Anz D, Bscheider M, Schwerd T, Berking C, Bourquin C, Kalinke U, Kremmer E, Kato H, Akira S, Meyers R, Häcker G, Neuenhahn M, Busch D, Ruland J, Rothenfusser S, Prinz M, Hornung V, Endres S, Tüting T, Hartmann G. 5'-Triphosphate-siRNA: turning gene silencing and Rig-I activation against melanoma. Nat Med. 14(11):1256-63. (2008)

Hornung V, Ellegast J, Kim S, Brzózka K, Jung A, Kato H, Poeck H, Akira S, Conzelmann KK, Schlee M, Endres S, Hartmann G. 5'-Triphosphate RNA is the ligand for RIG-I. Science. 314(5801):994-7(2006)

Reviews
Schlee M. Master sensors of pathogenic RNA-RIG-I like receptors. Immunobiology. 218(11):1322-35 (2013)

Van den Boorn JG, Hartmann G. Turning tumors into vaccines: co-opting the innate immune system. Immunity. 39(1):27-37 (2013)

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