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Project 21
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Project 21

Veit Hornung

Induction of type I interferon by intracellular double stranded DNA

Institute for Clinical Chemistry and Pharmacology, University of Bonn

Publications

Brief description in German:
Das angeborene Immunsystem hat vielfältige Strategien entwickelt, um fremde Nukleinsäuren zu detektieren. Wir konnten vor kurzem einen neuartigen intrazellulären doppelstängigen (ds) DNA-Erkennungsmechanismus aufdecken, der indirekt durch die transkriptionelle Aktivität von RNA Polymerase III stimuliert wird. Intrazelluläre dsDNA wird von RNA Polymerase III in 5’ Triphosphat-RNA transkribiert, die wiederum RIG-I aktiviert. In diesem Projekt möchten wir diesen neuen molekularen Mechanismus charakterisieren und die Rolle dieses Detektions-systems in der Zell-autonomen, antiviralen Immunantwort gegen DNA Viren entschlüsseln.

Summary
The innate immune system has evolved versatile ways to detect foreign nucleic acids. We have recently discovered a novel intracellular double stranded (ds) DNA sensing pathway that works indirectly via the formation of an endogenous RNA intermediate.

RNA polymerase III transcribes artificially in-troduced intracellular dsDNA in a promoter-independent fashion and generates a 5’ triphosphate RNA molecule that is detected by the cytosolic RNA helicase RIG-I. We will further characterize the molecular mechanisms of this pathway and determine its role in the cell-autonomous, antiviral immune response against DNA viruses.

Several classes of germ-line encoded pattern recognition receptors (PRRs) have been identified, which sense self and non-self nucleic acids and trigger potent innate immune responses. These PRRs include membrane bound toll-like receptors (TLRs) and cytoplasmic RNA helicases, in-cluding RIG-I and MDA5.

While these known PRRs largely explain the innate immune response towards RNA viruses, little is known about mechanisms that sense foreign DNA within the cyto-sol. We have recently characterized the novel inflammasome receptor AIM2 (absent in mela-noma 2) that detects viral DNA in the cytosol. While activation of AIM2 triggers caspase-1 activation and cell death upon dsDNA recognition, type I interferon (IFN) induction is not attenu-ated in the absence AIM2. Therefore, additional sensing mechanisms must exist that allow our immune system to detect foreign DNA in the cytosol leading to type I IFN induction and in-flammatory cytokine production.

In the course of our studies on the novel receptor protein AIM2, we have made the surprising observation that certain types of dsDNA (e.g. poly(dA:dT)) triggered type I IFN production is exclusively mediated via RIG-I in the human system. This is due to the formation of an endogenous RNA ligand that in turn activates RIG-I.

We have furthermore shown that RNA polymerase III (RNA Pol III) is the critical enzyme that converts poly(dA:dT) into 5’ triphosphate dsRNA ligand, namely poly(rA:rU). In the case of poly(dA:dT), RNA Pol III-mediated transcription is independent of classical RNA Pol III promoter sites. Under physiological conditions, this pathway is important in sensing Epstein-Barr virus encoded small RNAs, which are transcribed by RNA Pol III and then trigger RIG-I activation.

We want to further elucidate the molecular mechanisms of this pathway and further substantiate its role in the cell-autonomous innate immune response to DNA viruses. We will explore the sequence requirements for RNA Pol III-mediated promoter-independent transcription of dsDNA and furthermore study the localization of RNA Pol III tran-scription of exogenous DNA. Furthermore, we will use HSV1 as a model pathogen to dissect the mechanisms of this pathway in the innate defense against DNA viruses.

Altogether we expect that these studies may significantly enhance our understanding of antiviral immunity and also potentially open new avenues for therapeutic approaches.

 

List of relevant publications
Ablasser, A., Bauernfeind, F., Hartmann, G., Latz, E., Fitzgerald, K.A.*, and Hornung, V.* 2009. RIG-I-dependent sensing of poly(dA:dT) through the induction of an RNA polymerase III-transcribed RNA intermediate. Nat Immunol 10: 1065-1072.

Schlee, M., Roth, A., Hornung, V., Hagmann, C.A., Wimmenauer, V., Barchet, W., Coch, C., Janke, M., Mihailovic, A., Wardle, G., Juranek, S., Kato, H., Kawai, T., Poeck, H., Fitzgerald, K.A., Takeuchi, O., Akira, S., Tuschl, T., Latz, E., Ludwig, J., and Hartmann, G. 2009. 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.

Lichtnekert, J., Vielhauer, V., Zecher, D., Kulkarni, O.P., Clauss, S., Segerer, S., Hornung, V., Mayadas, T.N., Beutler, B., Akira, S., and Anders, H.J. 2009. Trif is not required for immune complex glomerulonephritis: dying cells activate mesangial cells via Tlr2/Myd88 rather than Tlr3/Trif. Am J Physiol Renal Physiol 296: F867-874.

Hornung, V., Ablasser, A., Charrel-Dennis, M., Bauernfeind, F., Horvath, G., Caffrey, D.R., Latz, E., and Fitzgerald, K.A. 2009. AIM2 recognizes cytosolic dsDNA and forms a caspase-1-activating inflammasome with ASC. Nature 458: 514-518.

Besch, R., Poeck, H., Hohenauer, T., Senft, D., Hacker, G., Berking, C., Hornung, V., Endres, S., Ruzicka, T., Rothenfusser, S., and Hartmann, G. 2009. Proapoptotic signaling induced by RIG-I and MDA-5 results in type I interferon-independent apoptosis in human melanoma cells. J Clin Invest 119: 2399-2411.

Berger, M., Ablasser, A., Kim, S., Bekeredjian-Ding, I., Giese, T., Endres, S., Hornung, V.*, and Hartmann, G.* 2009. TLR8-driven IL-12-dependent reciprocal and synergistic activation of NK cells and monocytes by immunostimulatory RNA. J Immunother 32: 262-271.

Bauernfeind, F.G., Horvath, G., Stutz, A., Alnemri, E.S., MacDonald, K., Speert, D., Fernandes-Alnemri, T., Wu, J., Monks, B.G., Fitzgerald, K.A., Hornung, V.*, and Latz, E.* 2009. Cutting edge: NF-kappaB activating pattern recognition and cytokine receptors license NLRP3 inflammasome activation by regulating NLRP3 expression. J Immunol 183: 787-791.

Ablasser, A., Poeck, H., Anz, D., Berger, M., Schlee, M., Kim, S., Bourquin, C., Goutagny, N., Jiang, Z., Fitzgerald, K.A., Rothenfusser, S., Endres, S., Hartmann, G.*, and Hornung, V.* 2009. Selection of molecular structure and delivery of RNA oligonucleotides to activate TLR7 versus TLR8 and to induce high amounts of IL-12p70 in primary human monocytes. J Immunol 182: 6824-6833.

Poeck, H., Besch, R., Maihoefer, C., Renn, M., Tormo, D., Morskaya, S.S., 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., Hacker, G., Neuenhahn, M., Busch, D., Ruland, J., Rothenfusser, S., Prinz, M., Hornung, V., Endres, S., Tuting, T., and Hartmann, G. 2008. 5'-Triphosphate-siRNA: turning gene silencing and Rig-I activation against melanoma. Nat Med 14: 1256-1263.

Hornung, V.*, Bauernfeind, F.*, Halle, A., Samstad, E.O., Kono, H., Rock, K.L., Fitzgerald, K.A., and Latz, E. 2008. Silica crystals and aluminum salts activate the NALP3 inflammasome through phagosomal destabilization. Nat Immunol 9: 847-856.

Halle, A., Hornung, V., Petzold, G.C., Stewart, C.R., Monks, B.G., Reinheckel, T., Fitzgerald, K.A., Latz, E., Moore, K.J., and Golenbock, D.T. 2008. The NALP3 inflammasome is involved in the innate immune response to amyloid-beta. Nat Immunol 9: 857-865.

Geeraedts, F., Goutagny, N., Hornung, V., Severa, M., de Haan, A., Pool, J., Wilschut, J., Fitzgerald, K.A., and Huckriede, A. 2008. Superior immunogenicity of inactivated whole virus H5N1 influenza vaccine is primarily controlled by Toll-like receptor signalling. PLoS Pathog 4: e1000138.

Allam, R., Pawar, R.D., Kulkarni, O.P., Hornung, V., Hartmann, G., Segerer, S., Akira, S., Endres, S., and Anders, H.J. 2008. Viral 5'-triphosphate RNA and non-CpG DNA aggravate autoimmunity and lupus nephritis via distinct TLR-independent immune responses. Eur J Immunol 38: 3487-3498.

Bourquin, C., Schmidt, L., Hornung, V., Wurzenberger, C., Anz, D., Sandholzer, N., Schreiber, S., Voelkl, A., Hartmann, G., and Endres, S. 2007. Immunostimulatory RNA oligonucleotides trigger an antigen-specific cytotoxic T-cell and IgG2a response. Blood 109: 2953-2960.

Hornung, V., Ellegast, J., Kim, S., Brzozka, K., Jung, A., Kato, H., Poeck, H., Akira, S., Conzelmann, K.K., Schlee, M., Endres, S., and Hartmann, G. 2006. 5'-Triphosphate RNA is the ligand for RIG-I. Science 314: 994-997.

Schlender, J.*, Hornung, V.*, Finke, S., Gunthner-Biller, M., Marozin, S., Brzozka, K., Moghim, S., Endres, S., Hartmann, G., and Conzelmann, K.K. 2005. Inhibition of toll-like receptor 7- and 9-mediated alpha/beta interferon production in human plasmacytoid dendritic cells by respiratory syncytial virus and measles virus. J Virol 79: 5507-5515.

Kerkmann, M., Costa, L.T., Richter, C., Rothenfusser, S., Battiany, J., Hornung, V., Johnson, J., Englert, S., Ketterer, T., Heckl, W., Thalhammer, S., Endres, S., and Hartmann, G. 2005. Spontaneous formation of nucleic acid-based nanoparticles is responsible for high interferon-alpha induction by CpG-A in plasmacytoid dendritic cells. J Biol Chem 280: 8086-8093.

Hornung, V., Guenthner-Biller, M., Bourquin, C., Ablasser, A., Schlee, M., Uematsu, S., Noronha, A., Manoharan, M., Akira, S., de Fougerolles, A., Endres, S., and Hartmann, G. 2005. Sequence-specific potent induction of IFN-alpha by short interfering RNA in plasmacytoid dendritic cells through TLR7. Nat Med 11: 263-270.

Bekeredjian-Ding, I.B., Wagner, M., Hornung, V., Giese, T., Schnurr, M., Endres, S., and Hartmann, G. 2005. Plasmacytoid dendritic cells control TLR7 sensitivity of naive B cells via type I IFN. J Immunol 174: 4043-4050.

* equally contributed

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