Toll-like Receptor Signaling During Infection and Inflammation Prof. Luke O Neill

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1 Toll-like Receptor Signalling 1 School of Biochemistry and Immunology Trinity College Dublin Ireland Inflammatory diseases: the facts 2 Overall incidence: 25% Major pathologies: Infectious diseases Rheumatoid arthritis Osteoarthritis Asthma Multiple sclerosis Crohn s disease Ulcerative Colitis Several types of anti-inflammatory drugs Newer drugs: Enbrel, Remicade, Rituxin, Tosilimumab New findings: Innate Immunity A starting point: a Russian, Mechnikov Nobel Prize for Medicine 1908 shared with Ehrlich for work on immunity Acceptance speech, December 11 th 1908: I will enlarge on the resistance of the body to disease and show you the practical value of pure research Animal organisms are very complex so I chose lower animals with very simple organisation to observe immunity Research should concentrate on the conditions capable of strengthening the phagocytic reaction 3 1

2 An Irishman and inflammation: Almroth Wright, founding father of innate immunity Firm advocate of stimulating the phagocytes Discoverer of opsonisation Wikipedia: Wright proposed that logic should be part of medical training but his idea was never adopted 4 Up to 1990s+: focus on the Ag Inflammation Inflammatory mediators and cytokines: e.g., IL1, TNF, IL6, IL12, IL8, etc. Macrophage Ag 5 Pathogens MHC/Ag presentation Co-stimulation Adaptive immunity: T and B cells 6 2

3 Our understanding of immunological recognition by clonally distributed receptors is approaching an asymptote Our knowledge of T and B cells mechanisms is approaching an asymptote Why do we use adjuvants? To be quite honest the answer is not known IL1 is required for priming and clonal expansion of T cells. One role for adjuvants is to induce IL1 The system predominantly discriminates infectious non-self from non-infectious self by means of a system of pattern recognition receptors..a new immunology that integrates innate and induced immunology Providing explanations for...induced second signals, the myriad of T cell surface molecules and A role for receptors that recognise patterns of microbial structures. This will require a rediscovery of microbiology by immunologists 7 Approaching the asymptote? evolution and revolution in immunology Cold Spring Harbour Symposia on Quantitative Biology 1989, 54: parts to the pathogen Inflammation Inflammatory mediators and cytokines: e.g., IL1, TNF, IL6, IL12, IL8, etc. Pattern recognition receptors Dendritic cell 8 PAMPs Ag Pathogens MHC/Ag presentation Co-stimulation Adaptive immunity: T and B cells Pentraxins Innate immunity 2009: 7 families of PRRs Phosphocholines Bacterial cell walls (e.g., LPS) Flagellin Fungal cell wall (e.g., Zymosan) LRR Toll-like receptors Bacterial surface Fungal surface (mannose) Fungal cell wall (beta-glucan) CRD MBL TIR ITAM Dectins Microbial genomes (dsrna, ssrna, dsdna) Endosome Toll-like receptors Bacterial cell wall fragments Bacterial toxins Uric acid Flagellin Viruses Potassium efflux ROS Lysosome disruption NOD-like receptors LRR NACHT CARD - PYRIN 9 Viral RNA RIG-I-like receptors Helicase CARD HIN200 family DNA HIN200 Pyrin 3

4 AIM2 Vaccinia PYRIN NALP3 Innate immunity to viruses 2010 Influenza Adenovirus MVA, EMCV PYRIN RIG-I Rabies VSV CARDs MAVS EBV, HSV Adenovirus RNA Pol III MDA5 Vaccinia EMCV MVA CARDs NOD2 RSV PYRIN CARD Innate immunity to viruses 2010 HSV, others? ASC CARD PYRIN TRAF3 K63 Uq NEMO STING IKKe TBK1 DDX3?? DAI DNARs? CMV Caspase 1 NF-kB IRF3/7 10 IL1β Pro-IL1β IFNβ 11 The sea urchin: 222 TLRs : NF-kappaBand p38 as key IL1 signals 12 4

5 13 Sims JE and Dower S, IL-1R family 1994 Eur Cyt Network 5, Toll! 14 Wild-type larvae Toll mutant Amazing Wild Weird Great Brilliant Cool Bodacious Superb Corky Mad Wow Go h-iontach The mouldy fly Toll-deficient Drosophila 15 Jules Hoffman and colleagues,

6 IL-1 receptor family The Toll / IL-1R superfamily in humans (a) Immunoglobulin domain sub-group: IL-1RI-like TIR domain Ig domain Leucine-rich repeat domain Death domain IL-1RI IL-1RAcP IL-18R IL-18RAcP ST2 IL-1Rrp2 IL-1RAPL TIGGIR1 SIGGIR (b) Leucine rich-repeat sub-group: TLRs TLR1 TLR2 TLR3 TLR4 TLR5 TLR6 TLR7 TLR8 TLR9 TLR10 (c) Adapter sub-group MyD88 Mal Trif Tram SARM 17 NOD-like receptors, RIGI-like receptors, C-type lectin receptors IL-1 IL-18 IL-33 (with AcP) IL-1F6,8 and 9 (with AcP) Orphans Partners TLR2 for blp blps, Zymosan dsrna LPS, F protein, hsp60, FN frags Flagellin Partners TLR2 for MALP-2 Imiquimod Imiquimod CpG DNA Pan-adapter Adapter for TLR2 and TLR4 Adapter for TLR3 Adapter for TLR4 Inhibitor of Trif? Toll-like receptors: how do we know they are important? Genetic variation and disease Therapeutic intervention in disease models 18 6

7 Mal S180L and disease Mal is a signalling adapter used by TLR2 and TLR4 Mal S180L heterozygotes have half the risk of malaria (3 populations), TB (3 out of 5 populations), pneumococcal pneumonia, Chagas disease and severe sepsis Mal S180L heterozygotes have a 3 fold decreased risk of SLE 19 When did Mal L180 arise in humans? 20 World distribution of MAL S180L in human populations Abbreviations: Sp, Spain; IRL, Ireland; UK, United Kingdom; Ge, Germany; Ro, Romania; Gr, Greece; Tu, Turkey; Ga, Gambia; Ma, Mali; Ta,Tanzania; Ke, Kenya; Ti, Trio Indians of Suriname; Isr, Israel; In, India; Ch, China; Pa, Papua New Guinea, Sp Irl UK Ge Ro Gr Tu SS SL LL Ga Ma Ta Ke Ti 21 Isr In Ch Pa 7

8 Worldwide human relationships inferred from genome-wide patterns of variation JZ Li et al., Science, 319, Superimposing Mal variants: S/L S/S L/L Native America Asia India Europe 22 Africa Middle East The leucine mutation probably occurred 8,000 years ago in the Middle East S/L S/S S/S S/L 80,000 yr S/L S/S S/S 23 Malaria, IPD, TB, Sepsis, SLE TLR2 TLR2 TLR2 MAL S180 MAL S180 MAL S180 MAL L180 MAL L180 MAL L180 Strong signal Modulated signal No signal Hyper-inflammation (TNF + IL-1) Increased risk of pathology/disease 24 Inflammation Lower risk of disease Less inflammation: Infection/Disease/Death 8

9 Columbus in the Americas, 1492: a family that split 67,000 years ago reunited Mal S/S Mal S/L 25 Death from infections How do TLRs work? 26 Toll-like receptors Microbial products Products of inflamed tissues 10 TLRs MyD88 Mal Trif Tram Modulation of immune and inflammatory genes Immune and inflammatory effector mechanisms 27 9

10 TLR4: ignition followed by acceleration Ignition LPS TLR4 NF-κB Pro-inflammatory gene expression IL1/TNF Acceleration Receptor for LPS from gram negative bacteria kills 600,000 people per year in Europe (septic shock 50% mortality) Most powerful immune response stimulator (half of immunology!) Implicated in many inflammatory pathologies, for example 28 TLRs in disease: allergy House dust mite allergen (Der p2) induces asthma via TLR4 triggering of airway structural cells; Hammad H et al., Nat Med (2009) 15, TLR4-MAPK-NF-κB interaction network 30 10

11 Golgi Recycling if empty of ligand TLR4 subcellular signalling LPS Plasma membrane PIP2 Mal MyD88 NF-κB No PIP2 Myr Tram Early endosome EEA1 Late endosome Rab7 E.R. Myr Tram Trif Myr Tram TAG Myr Tram TAG 31 IRF3 OFF Degraded TLR9 subcellular signalling DNA Plasma membrane DNA phagosome E.R. Unc93b Golgi endolysosome DNA MyD88 TLR9 cleaved (active) 32 Ewald SE et al., (2008) Nature 456, NF-κB 33 Summary Innate immunity involves at least 7 families of receptors for microbial products TLRs are the founder family and are involved in recognition of most pathogens NLRs, RLRs and CLRs are also very important, with respective roles in Summary anti-bacterial, anti-viral and anti-fungal responses Complex signalling pathways are activated by each family, leading to induction of host defense effector mechanisms There is great therapeutic potential in the targeting of these receptors for immune and inflammatory diseases 11

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