Project N1
Analysis of Nod like receptor (NLR) mediated innate immunity in mammalian cells
Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne
Brief description in German:
Charakterisierung der Rolle von Nod-like Rezeptoren (NLRs) in der angeborenen Immunantwort
Die meisten Körperzellen sind in der Lage nach einer Infektion durch Mikroorganismen (z.B. Bakterien oder Viren) eine Entzündungsreaktion auszulösen. Unsere Forschungsgruppe analysiert eine spezielle Gruppe von sogenannten "Mustererkennungsrezeptoren", den "Nod-like receptors" (NLRs). NLRs erkennen Bestandteile von Mikroorganismen, die Körperzellen infiziert haben. Mutationen in einigen dieser NLRs sind daher mit schweren entzündlichen Erkrankungen wie z.B. Morbus Crohn assoziiert. Obwohl im Menschen bisher 23 NLR-Proteine identifiziert wurden, ist derzeit nur für einige wenige die biologische Funktion bekannt.
Wir konzentrieren uns auf die funktionelle Charakterisierung der NLR-Proteine NOD1 und NOD2, die bestimmte bakterielle Bestandteile erkennen. Zudem versuchen wir ,die Funktion von bisher nicht beschriebenen NLRs im Detail zu entschlüsseln. Hierzu benutzen wir biochemische und zellbiologische Methoden und verwenden den Erreger der Bakterienruhr Shigella flexneri als Modellsystem.
 | Signaling pathways of NOD1 and NOD2 after activation by invasive bacteria, here depicted for the example of the invasive bacterium Shigella flexneri (please klick on picture to enlarge). |
Analysis of Nod like receptor (NLR) mediated innate immunity in mammalian cells
In order to defend against invading pathogens mammals have evolved a sophisticate immune systems that can detect and eliminate invading pathogens. Briefly, it can be divided into two parts: the innate immune system and the adaptive immune system. The innate immune system relies on germ line encoded receptors expressed on a variety of different cell-types that detect pathogens. Engagement of these receptors leads to inflammatory responses that also instruct the adaptive arm by priming its specialized cells which can generate specific antibodies and T cell receptors to virtually any antigen by somatic gene rearrangements.
Recognition of pathogens by the innate immune system relies on detection of invariant molecules derived from the intruder by specialized receptors on the host cell. In mammals, sensing of these so called "pathogen-associated-molecular-pattern" (PAMP) is mediated by specialized pattern-recognition-receptors (PRRs). Different types of PRRs show distinct sub-cellular localization, allowing the host to react to extracellular, vesicular and cytosolic presented PAMPs.
One class of PRRs, the nucleotide-binding domain, leucine-rich repeat containing (NLR) protein family gained much attention as it was shown that members of this family are critically involved in mounting immune responses to bacterial peptidoglycan fragments and in controlling release of the key inflammatory cytokine IL-1beta.
Over 20 NLRs are encoded in the human genome that share distant homology to proteins that refer immunity in plants. As hallmark they share a tripartite molecular architecture with a centrally located ATPase domain, a NACHT domain that mediates oligomerization and activation of these proteins, followed by a leucine-rich repeat region (LRR) at the C-terminus.
With the some rare exceptions, the N-terminal part of the NLRs consists of a domain that adopts a death-domain fold. Most NLR members thereby have either a caspase activation and recruitment domain (CARD) or a pyrin domain (PYD) domain, connecting the respective NLR to different downstream signalling events. Well studied examples of CARD domain containing NLRs are NOD1 and NOD2 that respond to molecular subunits of peptidoglycan, the mayor component of the cell-wall of Gram-positive and -negative bacteria and mediate NF-kappaB, MAPK and caspase activation.
Examples of PYD containing NLRs comprise NLRP1 and NLRP3, which form high molecular weight platforms, so-called inflammasomes that lead to activation of caspase-1 and subsequent IL-1beta release upon encounter of DAMPs and certain PAMPs. Mutations in NOD1 and NOD2 are linked to severe inflammatory disorders, such as inflammatory bowel disease, Crohn's disease (CD), Blau-syndrom, asthma and early onset sarcoidosis.
 | Ectopically expressed NOD2 (red) localizes at the actin rich (actin=green) entry foci of S. flexneri (blue) in human HeLa cells (see Kufer et al. IAI 2006). |
However, the molecular details underlying NLR-activation, -signalling and crosstalk between NLRs and other PRRs are still poorly defined. Moreover, most human NLRs still await detailed characterization.
The research interest of our group is the characterization of regulatory mechanisms of NLRs and the elucidation of their signalling pathways, using biochemical and cell-biological approaches. A second focus lies on the detailed analysis of the sub-cellular localization and activation of NLRs using the invasive human pathogen Shigella flexneri and other pathogens as model systems. Thereby a particular focus lies on the characterization of the NLR proteins NOD1, NOD2 and NLRC5.
 | Interplay in bacterial sensing by NLRs. See Kufer and Sansonetti, 2007 for details (please klick on picture to enlarge). |
Selected recent publications (2005 - date):
Neerincx A, Rodriguez GM, Steimle V, Kufer TA. NLRC5 Controls Basal MHC Class I Gene Expression in an MHC Enhanceosome-Dependent Manner. J Immunol. 2012 [Epub ahead of print]
Bielig H., Dongre M., Zurek B., Wai SN., Kufer T.A., A role for quorum sensing in regulating innate immune responses mediatedt by Vibrio cholerae outer membrane vesicles (OMVs). Gut Microbes 2011 Sep 1;2(5)
Maekawa T., Kufer T.A., Schulze-Lefert P. NLR functions in plant and animal immune systems: so far and yet so close. Nat Immunol. 2011 Aug 18;12(9):817-26. doi: 10.1038/ni.2083.
Zurek B., Bielig H., Kufer T.A. Cell-based reporter assay to analyze activation of NOD1 and NOD2. Methods Mol Biol. 2011;748:107-19
Zurek B., Proell M., Wagner R.N., Schwarzenbacher R., Kufer T.A. Mutational analysis of human NOD1 and NOD2 NACHT domains reveals different modes of activation. Innate Immunity 2012; Feb 18(1):100-11
Keller JF, Carrouel F, Staquet MJ, Kufer TA, Baudouin C, Msika P, Bleicher F, Farges JC. Expression of NOD2 is increased in inflamed human dental pulps and lipoteichoic acid-stimulated odontoblast-like cells. Innate Immun 2011;17(1):29-34. Epub 2009 Oct. 30.
Bielig H., Rompikuntal P.K., Mitesh D., Zurek B., Lindmark B., Ramstedt M., Wai S.N. and Kufer T.A. NOD-like receptor activation by outer-membrane vesicles (OMVs) from non-O1 non-O139 Vibrio cholerae is modulated by the quorum sensing regulator HapR. Infect Immun 2011 Apr;79(4):1418-27
Kufer TA, Sansonetti PJ. NLR functions beyond pathogen recognition. Nat Immunol 2011 Feb;12(2):121-8.
Bielig H, Velder J, Saiai A, Menning M, Meemboor S, Kalka-Moll W, Krönke
M, Schmalz HG, Kufer TA. Anti-inflammatory Arene-Chromium Complexes Acting as Specific Inhibitors of NOD2 Signalling. Chem Med Chem 2010 Dec 3;5(12):2065-71.
Neerincx A, Lautz K, Menning M, Kremmer E, Zigrino P, Hosel M, Buning H, Schwarzenbacher R, Kufer TA. A role for the human NLR family member NLRC5 in antiviral responses. J Biol Chem 2010 Aug 20;285(34):26223-32.
Allison CC, Kufer TA, Kremmer E, Kaparakis M, Ferrero RL. Helicobacter pylori induces MAPK phosphorylation and AP-1 activation via a NOD1-dependent mechanism. J Immunol 2009 Dec 15;183(12):8099-109.
Bielig H, Zurek B, Kutsch A, Menning M, Philpott DJ, Sansonetti PJ, Kufer TA.
A function for AAMP in Nod2-mediated NF-kappaB activation. Mol Immunol 2009 Aug;46(13):2647-54.
Wagner R.N., Proell M., Kufer T.A., Schwarzenbacher R. Evaluation of Nod-Like Receptor (NLR) Effector Domain Interactions. PLoS One 2009;4(4):e4931.
Kufer TA. Signal transduction pathways used by NLR-type innate immune receptors. Mol Biosyst 2008 May;4(5):380-6. Epub 2008 Mar 27. Review
Simhadri VR, Reiners KS, Hansen HP, Topolar D, Simhadri VL, Nohroudi K, Kufer TA, Engert A, Pogge von Strandmann E. Dendritic cells release HLA-B-associated transcript-3 positive exosomes to regulate natural killer function. PLoS ONE 2008;3(10):e3377.
Kufer TA, Sansonetti PJ. Sensing of bacteria: NOD a lonely job. Curr Opin Microbiol 2007 Feb;10(1):62-9. Epub 2006 Dec 11. Review
Kufer T.A., Kremmer E., Adam A.C., Philpott D.J., Sansonetti P.J. The pattern-recognition molecule Nod1 is localized at the plasma membrane at sites of bacterial interaction. Cellular Microbiology 2007 Oct 26
Fritz J.H.,Le Bourhis L., Fsihi H., Viala J., Sellge G., Joao Gamelas Magalhaes J.G., Kufer T.A., Ferrero R.L., Girardin S.E., Philpott D.J. Nod1-Mediated Innate Immune Recognition of Peptidoglycan Contributes to the Onset of Adaptive Immunity. Immunity 2007, Apr 10
Legrand-Poels S. Kustermans G., Bex F., Kremmer E., Kufer T.A., Piette J. Modulation of Nod2-dependent NF-{kappa}B signaling by the actin cytoskeleton.
Journal of Cell Science 2007 ; 120(7):1299-310
Kufer T.A., Kremmer E., Banks D.J.,Philpott D.J. A role for Erbin in bacterial activation of Nod2. Infection and Immunity 2006;74(6):3115-24.
Kufer TA, Fritz JH, Philpott DJ. NACHT-LRR proteins (NLRs) in bacterial infection and immunity. Trends Microbiol 2005 Aug;13(8):381-8. Review
