Mintázatfelismerő receptorok

Az előadások a következő témára: "Mintázatfelismerő receptorok"— Előadás másolata:

1 Mintázatfelismerő receptorok
Pathogen associated molecular pattern (PAMP) Pattern recognititon receptors: Toll-like receptors (TLR) Mannóz receptor, C-típusu lektinek scavenger receptor, N-formil metionin receptor Korlátozott diverzitás, molekuláris mintázat , csíravonalban kódolt, nem- klonális megoszlás, saját és idegen megkülönböztetése Stresszelt vagy sérült sejtek felismerése

2 A mintázatfelismerő molekulák elhelyezkedése a sejten belül
Nucleotide-binding domain, Leucine-Rich repeat containing protein , NOD like receptor (NLR), Retinoic acid inducible gene, (RIG-I), CARD: caspase activation and recruitment domain,

3 TLR Evolúcionárisan konzervált, Mikrobák felismerése
Mikrobák elleni védelem TLR 1-11 TIR domén Természetes immunitás

4 Emlősök TLR-ei Pathogen Associated Molecular Pattern

5 Toll-szerű receptorok Toll-Like Receptors (TLR)-
a gazdaszervezet védelmében Figure 1 | TIR domain in host-defence pathways.   The Toll/interelukin-1 (IL-1) receptor (TIR) domain is a protein-interaction module found in transmembrane and cytoplasmic proteins involved in animal and plant immunity. RPP5, N and L6 are prototypic examples of intracellular plant-disease-resistance proteins that contain an amino-terminal TIR domain as well as a nucleotide (ATP or GTP)-binding domain and leucine-rich repeat (LRR) domains. Drosophila has two types of protein with TIR domains: Tolls and MyD88. At least one out of nine Tolls in Drosophila, as well as MyD88, are involved in host defence. Toll is activated by a proteolytically processed form of the Spätzle protein. The cleavage of Spätzle is triggered by an unknown pattern-recognition molecule responsive for fungal and Gram-positive bacterial pathogens (see text for details). Mammals have at least four types of proteins with TIR domains: members of the Toll-like receptor (TLR) and IL-1 receptor (IL-1R) families, MyD88 and TIRAP (TIR domain-containing adaptor protein). In TLRs and IL-1Rs, the TIR domain is carboxy-terminal to LRRs and immunoglobulin domains, respectively. Both mammalian and Drosophila MyD88 contain carboxy-terminal TIR domains and amino-terminal death domains and function as adaptor proteins. TIRAP is another adaptor protein that does not have a Drosophila homologue. TIR has a carboxy-terminal TIR domain, but lacks a death domain. The amino-terminal region of TIRAP does not share similarity with any known protein. PAMP, pathogen-associated molecular pattern. Toll/interelukin-1 (IL-1) receptor (TIR) domén: Fehérje kölcsönhatásért felelős modul, lehet transzmembrán és citoplazmatikus fehérjékben Növényi és állati immunválaszban szerepel PAMP: pathogen-associated molecular pattern TIRAP: TIR domain-containing adaptor protein.

6 Emlősökben 4 féle TIR domént tartalmazó fehérje van:
A TLR család tagjai IL-1 receptor család tagjai MyD88 TIRAP (TIR domént tartalmazó adapter fehérje)

7 (Myeloid Differentiation primary-response protein 88)
MyD88-adaptor család (Myeloid Differentiation primary-response protein 88) Box 1 | MyD88-adaptor family  The figure shows the domain structure of the five known members of the MyD88 (myeloid differentiation primary-response protein 88) -adaptor family: MyD88, TIRAP (Toll/interleukin-1 receptor (TIR)-domain-containing adaptor protein; also known as MyD88-adaptor-like protein, MAL), TRIF (TIR-domain-containing adaptor protein inducing interferon- ), TRAM (TRIF-related adaptor molecule) and SARM (sterile - and armadillo-motif-containing protein)15. Evidence from genetic and biochemical studies has linked signalling from Toll-like receptor 7 (TLR7), TLR8 and TLR9 to a pathway that requires only MyD88. By contrast, signalling from TLR2, after the formation of a heterodimer with either TLR1 or TLR6, is linked to a pathway that requires a complex of MyD88 and a second adaptor, TIRAP. The activation of interferon-regulatory factor 3 (IRF3) and the subsequent induction of interferon- , which are elicited by signalling from TLR3 or TLR4, are MyD88-independent and involve a third adaptor, TRIF. A fourth adaptor, TRAM, is also involved in MyD88-independent signalling through TLR4 (Ref. 74–77), and a fifth adaptor, SARM, has been described, but its role in TLR signalling is unclear at present. At the most proximal step after ligand binding, these proteins of the MyD88-adaptor family provide specificity for the outcome of TLR signalling that is initiated following the engagement of different types of ligands by different TLRs. DD, death domain; SAM, sterile -motif. TIRAP : Toll/interleukin-1 receptor (TIR)-domain-containing adaptor protein TRAM : TRIF-related adaptor molecule TRIF : TIR-domain-containing adaptor protein inducing interferon g SARM : sterile- and armadillo-motif-containing protein A különféle TLR-ligandum kötődést követő első lépésben a MyD család következő tagjai biztosítják a specifikus jeltovábbítást : TLR7, TLR8, TLR9: MyD88 TLR2-TLR1 vagy TLR6 heterodimer: MyD88+TIRAP TLR3, TLR4: TRIF TLR4: TRAM

8 A TLR-ek ligandum specificitása
Figure 3 | Ligand specificities of TLRs.   Toll-like receptors (TLRs) recognize a variety of pathogen-associated molecular patterns (PAMPs). Recognition of lipopolysaccharide (LPS) by TLR4 is aided by two accessory proteins: CD14 and MD-2. TLR2 recognizes a broad range of structurally unrelated ligands and functions in combination with several (but not all) other TLRs, including TLR1 and TLR6. TLR3 is involved in recognition of double-stranded (dsRNA). TLR5 is specific for bacterial flagellin, whereas TLR9 is a receptor for unmethylated CpG motifs, which are abundant in bacterial DNA. G+, Gram-positive; G–, Gram negative; GPI, glycophosphoinositol; RSV, respiratory syncytial virus. Toll-like receptors (TLRs) recognize a variety of pathogen-associated molecular patterns (PAMPs).

9 1,2,4,5,6,8 1,2,6,3,8, , ,2,6,3,9 1,2,6,37,9,5 TLR expression A dendritikus sejtek egyes populációi nem átfedő típusú TLR-t expresszálnak: Különböző TLR-ek a sejtmembránon és a lizoszóma kompartmentben eltérő receptorokat fejeznek ki. Myeloid DCs : baktérium, gomba, vírus patogének felismerésére felszíni receptorok, Válasz: IL-2, TNF, IL-6 termelés Monocyta: nincs TLR3, érés közben expresszió nő Plazmocitoid DC: TLR7, TLR9, vírusokra Type I IFN termeléssel válaszol Másodlagos nyirokszervek: vér eredetű DC átfedő, de nem azonos TLR

10 Toll signalling pathways
Figure 4 | Toll signalling pathways.   The Toll-like receptor (TLR) and interleukin-1 receptor (IL-1R)-family members share several signalling components, including the adaptor MyD88, Toll-interacting protein (TOLLIP), the protein kinase IRAK (IL-1R-associated kinase) and TRAF6 (TNF receptor-associated factor 6). TRAF6 can activate nuclear factor- B (NF- B) through TAK1 (TGF- -activated kinase), and JNK (c-Jun N-terminal kinase) and p38 MAP kinases through MKK6 (mitogen-activated protein kinase kinase 6). TLR4 signals through another adaptor in addition to MyD88–TIRAP (Toll/interelukin-1 (IL-1) receptor domain-containing adaptor protein), which activates MyD88-independent signalling downstream of TLR4. The protein kinase PKR functions downstream of TIRAP, but its importance in this pathway has not yet been established. TOLLIP (Toll-interacting protein ) IRAK (IL-1R-associated kinase) TRAF6 (TNF receptor-associated factor 6) TAK1 (TGF-b activated kinase), TIRAP (Toll/interelukin-1 (IL-1) receptor domain-containing adaptor protein) MKK6 (mitogen-activated protein kinase kinase 6).

11 Role of TLRs in the control of adaptive immunity
TOLL-LIKE RECEPTORS AND INNATE IMMUNITY Role of TLRs in the control of adaptive immunity Figure 5 | Role of TLRs in the control of adaptive immunity.   TLRs sense the presence of infection through recognition of PAMPs (pathogen-associated molecular patterns). Recognition of PAMPs by Toll-like receptors (TLRs) expressed on antigen-presenting cells (APC), such as dendritic cells, upregulates cell-surface expression of co-stimulatory (CD80 and CD86) molecules and major histocompatibility complex class II (MHC II) molecules. TLRs also induce expression of cytokines, such as interleukin-12 (IL)-12, and chemokines and their receptors, and trigger many other events associated with dendritic cell maturation. Induction of CD80/86 on APCs by TLRs leads to the activation of T cells specific for pathogens that trigger TLR signalling. IL-12 induced by TLRs also contributes to the differentiation of activated T cells into T helper (TH)1 effector cells. It is not yet known whether TLRs have any role in the induction of TH2 responses. IFN- ; interferon- ; PRR, pattern-recognition receptor. ? TLR PAMP felismerése az MHCII és CD80/86 kostimulátor molekulák overexpresszióját váltja ki, citokin, kemokin és ezek receptorainak expresszióját indukálja.

12 A TLR9 jelpálya modellje B sejtben
A proposed model for TLR9 signaling in B cells. Environmental CpG-containing DNAs (mammalian or non-mammalian) are directly endocytosed or internalized after binding to anti-DNA BCRs. TLR9, normally generated and found in the endoplasmic reticulum, encounters CpG DNAs in lysosomal/late endosomal compartments, where it initiates a signaling cascade via MyD88–IRAK–TRAF6–TAK1, similar to TLR4 (Figure 1 but without TIRAP. TAK1 activates NIK, which activates the IKK complex, which in turn phosphorylates the IkB proteins that normally sequester NF-kB proteins in the cytoplasm. TAK1 also activates the JNK and p38 MAPK pathways, leading to the activation of AP-1 complexes. Both the NF-kB and AP-1 proteins enter the nucleus where they activate target genes involved in B cell activation, proliferation, and immunoglobulin (Ig) production. In anergic B cells, such as anti-DNA B cells in non-autoimmune hosts, BCR signaling is dissociated from activation such that BCR signaling alone results in suboptimal NF-kB activation, perhaps related to tonic ERK activation. TLR9 ligation can bypass and/or co-stimulate this mechanism, leading to autoreactive B cell activation. Abbreviations: AP-1, activating protein; BCR, B cell receptor; IkB, inhibitor of NF-kB; IKK, IkB kinase; IRAK, IL-1R-associated kinase; NIK, NF-kB-inducing kinase; TAK, TGF-b-activated kinase; TRAF, TNF-receptor-associated factor; TIRAP, TIR-domain-containing adaptor protein. A TLR9 jelpálya modellje B sejtben In anergic B cells, such as anti-DNA B cells in non-autoimmune hosts, BCR signaling is dissociated from activation such that BCR signaling alone results in suboptimal NF-kB activation, perhaps related to tonic ERK activation. TLR9 ligation can bypass and/or co-stimulate this mechanism, leading to autoreactive B cell activation.

13 TLR szerepe a tolerancia áttörésében
Combined stimulation through the TLR and an autoreactive BCR leads to activation of B cells in multiple ways Autoreactive B cells can then proliferate, or up-regulate co-stimulatory molecules turning them into more efficient APCs. They also secrete cytokines and can differentiate into plasma cells that produce large amounts of immunoglobulins (Ig). In addition, expression of activation induced cytidine deaminase (AID) is directly induced by TLR signals. This results in class switching to higher affinity immunoglobulins, and –if autoreactivity is retained–, increasingly pathogenic antibodies are generated. TLR szerepe a tolerancia áttörésében

14 TLR structure and signalling
leucine rich repeats, LRR TIR domén UBC13 (ubiquitin-conjugating enzyme 13) and UEV1A (ubiquitin-conjugating enzyme E2 variant 1). This leads to ubiquitylation of TRAF6, which induces the activation of TAK1. Figure 1 | TLR structure and signalling.   a | Toll-like receptors (TLRs) and interleukin-1 receptors (IL-1Rs) have a conserved cytoplasmic domain, that is known as the Toll/IL-1R (TIR) domain. The TIR domain is characterized by the presence of three highly homologous regions (known as boxes 1, 2 and 3). Despite the similarity of the cytoplasmic domains of these molecules, their extracellular regions differ markedly: TLRs have tandem repeats of leucine-rich regions (known as leucine rich repeats, LRR), whereas IL-1Rs have three immunoglobulin (Ig)-like domains. b | Stimulation of TLRs triggers the association of MyD88 (myeloid differentiation primary-response protein 88), which in turn recruits IRAK4 (IL-1R-associated kinase 4), thereby allowing the association of IRAK1. IRAK4 then induces the phosphorylation of IRAK1. TRAF6 (tumour-necrosis-factor-receptor-associated factor 6) is also recruited to the receptor complex, by associating with phosphorylated IRAK1. Phosphorylated IRAK1 and TRAF6 then dissociate from the receptor and form a complex with TAK1 (transforming-growth-factor- -activated kinase), TAB1 (TAK1-binding protein 1) and TAB2 at the plasma membrane (not shown), which induces the phosphorylation of TAB2 and TAK1. IRAK1 is degraded at the plasma membrane, and the remaining complex (consisting of TRAF6, TAK1, TAB1 and TAB2) translocates to the cytosol, where it associates with the ubiquitin ligases UBC13 (ubiquitin-conjugating enzyme 13) and UEV1A (ubiquitin-conjugating enzyme E2 variant 1). This leads to the ubiquitylation of TRAF6, which induces the activation of TAK1. TAK1, in turn, phosphorylates both mitogen-activated protein (MAP) kinases and the IKK complex (inhibitor of nuclear factor- B (I B)-kinase complex), which consists of IKK- , IKK- and IKK- (also known as IKK1, IKK2 and nuclear factor- B (NF- B) essential modulator, NEMO, respectively). The IKK complex then phosphorylates I B, which leads to its ubiquitylation and subsequent degradation. This allows NF- B to translocate to the nucleus and induce the expression of its target genes. TAK1 (transforming-growth-factor-b-activated kinase), TAB1 (TAK1-binding protein 1)

15 TLR4 signalling: MyD88-függő és független
Nature Reviews Immunology 4; (2004); doi: /nri1391 TOLL-LIKE RECEPTOR SIGNALLING TLR4 signalling: MyD88-függő és független útvonalak interferon regulatory factor (IRF3) Figure 2 | TLR4 signalling: MyD88-dependent and -independent pathways.   Stimulation of Toll-like receptor 4 (TLR4) facilitates the activation of two pathways: the MyD88 (myeloid differentiation primary-response protein 88)-dependent and MyD88-independent pathways. The MyD88-dependent pathway involves the early phase of nuclear factor- B (NF- B) activation, which leads to the production of inflammatory cytokines. The MyD88-independent pathway activates interferon (IFN)-regulatory factor (IRF3) and involves the late phase of NF- B activation, both of which lead to the production of IFN- and the expression of IFN-inducible genes.

16 TLR és BCR által indukált jelek összegződése bakteriális fertőzés alatt
During an infection, naïve mature B cells integrate diverse stimuli and then embark upon an appropriate differentiation program. A microbe can directly induce activation signals by engaging the specific BCR as well as by engaging germline-encoded TLRs. TLR ligands located on the surface of a microbe are sensed by the extracellular TLRs, whereas nucleic acid TLR ligands located inside a microorganisms are sensed by endosomal TLRs. TLR signaling is initially relayed through homotypic TIR-TIR interactions (blue spheres) with TIR adapters, and eventually results in the activation of pathways such as canonical NF-κB. TNF family receptors interact with corresponding surface DCs and TH cells, as well as with secreted ligands (BAFF, APRIL). CD40 induced both canonical as well as non-canonical NF-κB whereas, BAFFR, TACI, BCMA induce only non-canonical NF-κB. Both TLRs and TNFRs may induce AID, an essential enzyme for SHM and CSR. Cytokine receptors are activated by the corresponding cytokines, and are particularly important for inducing isotype-specific germline transcription during CSR. TLR-mediated polyclonal responses contribute to the production of natural antibodies during the early stages of infection, as well as to periodic re-activation of memory and plasma cells to maintain serological memory. Antigen-specific responses are typically T cell dependent and develop in germinal centers (GCs) several days after infection, though T-independent, dual TLR- and BCR-mediated antigen-specific responses may produce germline-endoded IgM and class-switched antibodies that help contain pathogens until the T-dependent response peaks. B cells activated by initial TLR and BCR engagement are primed to enter the GC reaction and undergo antibody affinity maturation, providing an emerging paradigm regarding signal integration in B cells. Deficiencies in TLR signaling compromise antibody responses, whereas inappropriate TLR-induced activation of autoreactive B cell results in autoantibody production. During an infection, naïve mature B cells integrate diverse stimuli and then embark upon an appropriate differentiation program. A microbe can directly induce activation signals by engaging the specific BCR as well as by engaging germline-encoded TLRs. TLR ligands located on the surface of a microbe are sensed by the extracellular TLRs, whereas nucleic acid TLR ligands located inside a microorganisms are sensed by endosomal TLRs. TLR signaling is initially relayed through homotypic TIR-TIR interactions (blue spheres) with TIR adapters, and eventually results in the activation of pathways such as canonical NF-κB. TNF family receptors interact with corresponding surface DCs and TH cells, as well as with secreted ligands (BAFF, APRIL). CD40 induced both canonical as well as non-canonical NF-κB whereas, BAFFR, TACI, BCMA induce only non-canonical NF-κB. Both TLRs and TNFRs may induce AID, an essential enzyme for SHM and CSR. Cytokine receptors are activated by the corresponding cytokines, and are particularly important for inducing isotype-specific germline transcription during CSR.

17 TRIF-dependent induction of IFNg
Figure 4 | TRIF-dependent induction of IFN- .   The amino-terminal region of TRIF (Toll/interleukin-1-receptor (TIR)-domain-containing adaptor protein inducing interferon (IFN)- ) interacts with both TRAF6 (tumour-necrosis-factor-receptor-associated factor 6) and TBK1 (TRAF-family-member-associated nuclear factor- B (NF- B) activator (TANK)-binding kinase 1). TRIF-dependent activation of TBK1 leads to the phosphorylation of IRF3 (IFN-regulatory factor 3), and TRAF6 mediates NF- B activation. RIP1 (receptor-interacting protein 1) mediates the NF- B activation that is induced through the carboxy-terminal region of TRIF. Activation of both NF- B and IRF3 contributes to the activation of the IFN- gene. I B, inhibitor of NF- B; TLR, Toll-like receptor. TRIF: Toll/interleukin-1-receptor (TIR)-domain-containing adaptor protein inducing interferon (IFN)- RIP1 (receptor-interacting protein 1) TBK1 (TRAF-family-member-associated nuclear factor- B (NF- B) activator (TANK)-binding kinase 1). IRF3 (IFN-regulatory factor 3),

18 IRAK asszociációt gátolja
Negative regulation of TLR signalling IRAK asszociációt gátolja Disszociációt gátol Figure 5 | Negative regulation of TLR signalling.   Toll-like receptor (TLR)-signalling pathways are negatively regulated by several molecules that are induced by the stimulation of TLRs. IRAK-M (interleukin-1-receptor (IL-1R)-associated kinase M) inhibits the dissociation of the IRAK1–IRAK4 complex from the receptor. SOCS1 (suppressor of cytokine signalling 1) probably associates with IRAK1 and inhibits its activity. MyD88s (myeloid differentiation primary-response protein 88 short) blocks the association of IRAK4 with MyD88. The TIR (Toll/IL-1R)-domain-containing receptors SIGIRR (single immunoglobulin IL-1R-related molecule) and ST2 have also been shown to negatively modulate TLR signalling. I B, inhibitor of NF- B; IKK, I B kinase; NF- B, nuclear factor- B; TIRAP, TIR-domain-containing adaptor protein; TRAF6, tumour-necrosis-factor-receptor-associated factor 6. SOCS1 (Suppressor of cytokine signalling 1) probably associates with IRAK1 and inhibits its activity. MyD88s (myeloid differentiation primary-response protein 88 short) blocks the association of IRAK4 with MyD88. The TIR (Toll/IL-1R)-domain-containing receptors SIGIRR (single immunoglobulin IL-1R-related molecule) and ST2 have also been shown to negatively modulate TLR signalling.

19 TLRs as targets for therapy
Figure 2 | TLRs as targets for therapy.   Toll-like receptor (TLR) signalling is initiated by plasma membrane and intracellular (endosomal) TLRs following ligand recognition. TLR4 is shown as a representative membrane receptor and the endosomal TLRs (TLR7, TLR8 and TLR9) are also shown. Stimulation of TLRs triggers interaction with the adaptor molecule MyD88 (myeloid differentiation primary-response protein 88). Other adaptor molecules, TIRAP (Toll/interleukin-1 receptor (TIR)-domain-containing adaptor protein), TRIF (TIR-domain-containing adaptor molecule inducing interferon- ) and TRAM (TRIF-related adaptor molecule) are also involved in TLR4 signalling. The MyD88-dependent signalling pathway leads to the activation of nuclear factor- B (NF- B), which regulates the expression of target genes that encode pro-inflammatory mediators. Several laboratories have developed agonists of TLR4 and the endosomal TLRs that can stimulate TLR signalling and should be useful therapeutics for treating various human diseases. The key steps that could be targeted in the development of molecules to inhibit TLR signalling are also indicated. I B, inhibitor of NF- B; IKK, I B kinase; IRAK, interleukin-1-receptor-associated kinase; MPL, monophosphoryl lipid A; ODN, oligodeoxynucleotide; RIP, receptor-interacting protein; TRAF, tumour-necrosis-factor-receptor-associated factor 6.

20 Figure 1 | Mammalian TLR signalling pathways
Figure 1 | Mammalian TLR signalling pathways. A detailed knowledge of how mammalian Toll-like receptors (TLRs) signal has developed over the past 15 years. TLR5, TLR11, TLR4, and the heterodimers of TLR2–TLR1 or TLR2–TLR6 bind to their respective ligands at the cell surface, whereas TLR3, TLR7–TLR8, TLR9 and TLR13 localize to the endosomes, where they sense microbial and host-derived nucleic acids. TLR4 localizes at both the plasma membrane and the endosomes. TLR signalling is initiated by ligand-induced dimerization of receptors. Following this, the Toll–IL‑1‑resistence (TIR) domains of TLRs engage TIR domain-containing adaptor proteins (either myeloid differentiation primary-response protein 88 (MYD88) and MYD88‑adaptor-like protein (MAL), or TIR domain-containing adaptor protein inducing IFNβ (TRIF) and TRIF-related adaptor molecule (TRAM)). TLR4 moves from the plasma membrane to the endosomes in order to switch signalling from MYD88 to TRIF. Engagement of the signalling adaptor molecules stimulates downstream signalling pathways that involve interactions between IL‑1R‑associated kinases (IRAKs) and the adaptor molecules TNF receptor-associated factors (TRAFs), and that lead to the activation of the mitogen-activated protein kinases (MAPKs) JUN N-terminal kinase (JNK) and p38, and to the activation of transcription factors. Two important families of transcription factors that are activated downstream of TLR signalling are nuclear factor‑κB (NF‑κB) and the interferon-regulatory factors (IRFs), but other transcription factors, such as cyclic AMP-responsive element-binding protein (CREB) and activator protein 1 (AP1), are also important. A major consequence of TLR signalling is the induction of pro-inflammatory cytokines, and in the case of the endosomal TLRs, the induction of type I interferon (IFN). dsRNA, double-stranded RNA; IKK, inhibitor of NF-κB kinase; LPS, lipopolysaccharide; MKK, MAP kinase kinase; RIP1, receptor-interacting protein 1; rRNA, ribosomal RNA; ssRNA, single-stranded RNA; TAB, TAK1‑binding protein; TAK, TGFβ-activated kinase; TBK1, TANK-binding kinase 1.

21 Toll-Like Receptors LAM (Lipoarabinomannan), BLP (Bacterial Lipoprotein), and PGN (Peptidoglycans). Bacteria LBP Bacteria LAM LPS Yeast PGN Zymosan LTA BLP Flagellin LBP LPS Viral dSRNA DNA CpG Myco- Bacterial 19KDa Protein MALP2 Uropathogenic Bacteria MD2 T L R 1 T L R 2 T L R 2 T L R 6 ST2 T L R 4 T L R 4 T L R 5 CD14 SIGIRR T L R 1 T L R 1 T L R 1 2 CD14 T L R 1 3 TIRAP TIRAP MyD88 TIRAP MyD88 Rac TRAM MyD88 PI3K RIP2 Rac MyD88 BTK RIP2 TRIF PI3K MyD88s NF-kB Pathway MyD88 NF-kB IRF8 TLRs (Toll-like receptors) are transmembrane proteins expressed by cells of the innate immune system, which recognize invading microbes and activate signaling pathways that launch immune and inflammatory responses to destroy the invaders. Toll receptors were first identified in Drosophila. In mammals, the TLR family includes eleven proteins (TLR1−TLR11). Recently, two new members, TLR12 and TLR13 have been discovered in mouse, but not much information is known about them. Mammalian TLRs consist of an extracellular portion containing Leucine-rich repeats, a Transmembrane region and a Cytoplasmic tail, called the TIR (Toll-IL-1R (Interleukin-1-Receptor)) homology domain. Different TLRs serve as receptors for diverse ligands, including Bacterial cell wall components, Viral double-stranded RNA and small-molecule anti-viral or immunomodulatory compounds (Ref.1). Activation of TLRs occurs after binding of the cognate ligand. Upon activation, TLRs activates two major signaling pathways. The core pathway activated by most TLRs leads to activation of the transcription factor NF-KappaB (Nuclear Factor-KappaB) and the MAPKs (Mitogen-Activated Protein Kinases) p38 and JNK (c-Jun Kinase). The second pathway is activated by TLR3 (Toll-Like Receptor-3) and TLR4 (Toll-Like Receptor-4) and leads to activation of both NF-KappaB and another transcription factor IRF3 (Interferon Regulatory Factor-3), allowing for an additional set of genes to be induced, including anti-viral genes such as Ifn- Beta (Interferon-Beta). In this way, TLRs can tailor the innate response to pathogens. TLRs that recognize nucleic acids signal from endosomes, whereas cell-surface TLRs sense lipids and proteins. Plasma membrane localized TLRs include TLR1, TLR2, TLR4, TLR5, TLR6 and TLR10. Newly discovered TLR11, TLR12 and TLR13 are also believed to be Plasma membrane localized, whereas endosomal TLRs include TLR3, TLR7, TLR8 and TLR9 (Ref.2). Mammalian TLR4 is the signal-transducing receptor activated by the bacterial LPS (Lipopolysaccharide) and LTA (Lipotechoic Acid). Both LPS and LTA first bind to the CD14 (Cluster of Differentiation-14) receptor, which then transfer them to TLR4. TLR4 homodimerizes and forms a complex with the protein MD2, and this complex is then transported to the cell surface. The LTA can bind directly to CD14, but LPS must be delivered to CD14 by LBP (LPS-Binding Protein). Cells need both MD2 and TLR4 in order to recognize LPS. TLR4 activation engages a set of MyD88 (Myeloid Differentiation Primary-Response Protein-88) adaptor family members, including MyD88, TIRAP (TIR Domain-containing Adapter Protein), TRIF, and TRAM. BTK (Bruton Agammaglobulinemia Tyrosine Kinase) also participate in TLR signaling, although their precise role has yet to be defined (Ref.3). The MyD88 adaptor protein link TLR4 to the IRAK1 (Interleukin-1 Receptor-associated Kinase-1) and IRAK4 Serine/Threonine Kinases, leading to MyD88-dependent pathway. Upon activation of TLR4 with LPS, MyD88 recruits IRAK4, thereby allowing the association of IRAK1. IRAK4 then induces the phosphorylation of IRAK1. TRAF6 (Tumour Necrosis Factor-Receptor-Associated Factor-6) is also recruited to the receptor complex, by associating with phosphorylated IRAK1. Phosphorylated IRAK1 and TRAF6 then dissociate from the receptor and form a complex with TAK1 (TGF-Beta-Activated Kinase-1), TAB1 (TAK1-Binding Protein-1) and TAB2 (TAK1-Binding Protein-2), which induces the phosphorylation of TAB2 and TAK1. TRAF6, TAK1, TAB1 and TAB2 associate with the Ubiquitin ligases UbC13 (Ubiquitin-Conjugating Enzyme-13) and UEV1A (Ubiquitin-conjugating Enzyme E2-Variant-1). This leads to the ubiquitylation of TRAF6, which induces the activation of TAK1 by interacting with ECSIT (Evolutionarily Conserved Signaling Intermediate in Toll Pathways). TAK1, in turn, phosphorylates both p38 Kinases and JNKs by activating MKK3 (Mitogen-Activated Protein Kinase Kinase-3), MKK6 and MKK7. p38 and JNKs then enter the nucleus and induce the expression of their target genes. TAK1 also phosphorylates the IKK complex (Inhibitor of Kappa Light Polypeptide Gene Enhancer in B-Cells Kinase), which consists of IKK-Alpha (Inhibitor of Kappa Light Polypeptide Gene Enhancer in B-Cells Kinase of Alpha), IKK-Beta (Inhibitor of Kappa Light Polypeptide Gene Enhancer in B-Cells Kinase of Beta) and IKK-Gamma (Inhibitor of Kappa Light Polypeptide Gene Enhancer in B-Cells Kinase of Gamma). The IKK complex then phosphorylates I-KappaB, which leads to its ubiquitylation and subsequent degradation. This allows NF-KappaB to translocate to the nucleus and induce the expression of its target genes. TRIF and TRAM link TLR4 to pathways that lead to TBK1 (TANK Binding Kinase-1) and IRF3 activation (i.e., the MyD88-independent pathway) (Ref.1 & 4). TLR2 is activated by bacterial LAM (Lipoarabinomannan), BLP (Bacterial Lipoprotein), and PGN (Peptidoglycans). LAM and PGN act on TLR2 through the CD14 receptor, similar to the process followed by the TLR4 with a similar downstream affect. BLP mediates both apoptosis and NF-KappaB activation through TLR2. TLR2 is also responsible for the recognition of the Yeast cell-wall particle Zymosan. Zymosan acts through the CD14 receptor to influence TLR2. TLR2 signals the production of TNF (Tumour Necrosis Factor), through NF-KappaB pathway, from the phagocytized vesicle. TLR6 associates with TLR2 and recognizes diacylated MALP2 (Mycoplasmal macrophage-Activating Lipopeptide-2 kD) along with TLR2. Like TLR4, they also signal through MyD88 and TIRAP. PI3K (Phosphatidylinositde-3 Kinase), RIP2 (Receptor-Interacting Protein-2) and Rac (Ras-Related C3 Botulinum Toxin Substrate) are also involved in TLR6-TLR2 mediated signaling. TLR1 also associates with TLR2 and recognizes the native mycobacterial 19-kDa lipoprotein along with TLR2. TLR1-TLR2 also signals through MyD88, TIRAP, PI3K, RIP2 and Rac. TLR1 and TLR6 may participate in the activation of Macrophages by Gram-positive bacteria. TLR5 is a signaling mediator of bacterial Flagellin, thus activating NF-KappaB and may play a role in resistance to Salmonella infection (Ref.5). Human TLR10 is an orphan member of the TLR family. Genomic studies indicate that TLR10 is in a locus that also contains TLR1 and TLR6, two receptors known to function as coreceptors for TLR2. TLR10 not only homodimerize but also heterodimerize with TLRs 1 and 2. It has been found to activate gene transcription through MyD88 (Ref.6). TLR9 is responsible for the recognition of CpG islands of bacterial DNA. The extracellular CpG fragment may activate TLR9, thus inducing the endocytosis of the DNA along with TLR9, or perhaps the bacteria is phagocytized and TLR9, which has separately formed on the phagosome, is activated by the CpG islands; which ever the exact method, TLR9 activates the NF-KappaB pathway from the endocytized vesicle. Recently IRF8 (Interferon Regulatory Factor-8) has been shown to be activated by TLR9 through MyD88 (Ref.5). Besides TLR9, three other TLRs found in endosome are TLR3, TLR7 and TLR8. TLR3 activates immune cells in response to double-stranded Viral RNA. The stimulation of the TLR3 triggers TRIF activation that activates IRF3 through TBK1, independent of MyD88, which lead to the secretion of Ifn-Beta. TRIF also activates RIP1 (Receptor-Interacting Protein-1) and TRAF6, which may further activate NF-KappaB pathway. Small anti-viral compounds activate immune cells via the TLR7 MyD88 dependent signaling pathway. TLR7 binds with MyD88 and activates IRAF and TRAF6. TRAF6 then activates TANK (also known as I-TRAF). TANK interacts with TBK1 and IKK-Epsilon to activate IRF3. TLR7 or TLR8 may also activate IRF7 through activation of MyD88, BTK and TRAF6, thus inducing anti-viral responses by producing Ifn-Alpha (Interferon-Alpha). Recently, Mouse TLR11 has been identified as a participant in defense against Uropathogenic bacteria. The ligands for Mouse TLR12 and TLR13 are currently unknown. Three of them are believed to signal through MyD88 (Ref.7 & 8). TLR signaling is also negatively regulated by various proteins. The cell-surface receptors ST2 (also known as T1) and SIGIRR (Single ImmunoGlobulin IL-IR-Related molecule (TIR-8)) function as inhibitory receptors, sequestering proteins from signaling complexes and preventing TLR2, TLR4 and TLR9 signaling. IRAKM (Interleukin-1 Receptor-associated Kinase-M), TollIP (Toll-Interacting Protein) and a splice variant of MyD88, known as MyD88s, probably interfere with the recruitment and activation of IRAK4 and IRAK1. Recently, Triad3A, a RING-finger E3 ligase, has been shown to promote ubiquitylation of TLR4 and TLR9, targeting these TLRs for degradation and thereby negatively regulating the intensity and duration of TLR signaling. The balance between activation and inhibition is the key determinant of signal strength of TLR pathways (Ref.9 & 10). In addition to the innate immune response, evidence implicates the involvement of the TLR family in a spectrum of systemic disorders following bacterial infections including Sepsis, Cardiac Ischemia, Peridontitis, and Cerebral palsy. The TLRs that control the onset of an acute inflammatory response are critical antecedents for the development of adaptive acquired immunity. Genetic and developmental variation in the expression of microbial pattern recognition receptors may affect the individual's predisposition to infections in childhood and may contribute to susceptibility to severe neonatal inflammatory diseases, allergies, and autoimmune diseases (Ref.11). References: 1. Beutler B. Inferences, questions and possibilities in Toll-like receptor signalling. Nature Jul 8;430(6996): PubMed ID: 2. Nishiya T, DeFranco AL. Ligand-dependent Toll-like receptor 4 (TLR4)-oligomerization is directly linked with TLR4-signaling. J. Endotoxin Res. 2004;10(4): PubMed ID: 3. Saitoh S, Akashi S, Yamada T, Tanimura N, Matsumoto F, Fukase K, Kusumoto S, Kosugi A, Miyake K. The Toll-like receptor 2 is Recruited to Macrophage Phagosomes and Discriminates between Pathogens. Nature: Dec.16, 1999: Vol.402, pp PubMed ID: 4. Yamamoto M, Akira S. TIR domain--containing adaptors regulate TLR-mediated signaling pathways. Nippon. Rinsho Dec;62(12): PubMed ID: 5. Hallman M, Ramet M, Ezekowitz RA. Toll-like receptors as sensors of pathogens. Pediatr. Res Sep; 50(3): PubMed ID: 6. Hasan U, Chaffois C, Gaillard C, Saulnier V, Merck E, Tancredi S, Guiet C, Briere F, Vlach J, Lebecque S, Trinchieri G, Bates EE. Human TLR10 is a functional receptor, expressed by B cells and plasmacytoid dendritic cells, which activates gene transcription through MyD88. J. Immunol Mar 1;174(5): PubMed ID: 7. Barton GM, Medzhitov R. Linking Toll-like receptors to IFN-alpha/beta expression. Nat. Immunol May;4(5):432-3. PubMed ID: 8. Uematsu S, Sato S, Yamamoto M, Hirotani T, Kato H, Takeshita F, Matsuda M, Coban C, Ishii KJ, Kawai T, Takeuchi O, Akira S. Interleukin-1 receptor-associated kinase-1 plays an essential role for Toll-like receptor (TLR)7- and TLR9-mediated interferon-{alpha} induction. J. Exp. Med Mar 21;201(6): PubMed ID: 9. Meylan E, Burns K, Hofmann K, Blancheteau V, Martinon F, Kelliher M, Tschopp J. RIP1 is an essential mediator of Toll-like receptor 3-induced NF-kappa B activation. Nat. Immunol May;5(5):503-7. PubMed ID: 10. Brint EK, Xu D, Liu H, Dunne A, McKenzie AN, O'Neill LA, Liew FY. ST2 is an inhibitor of interleukin 1 receptor and Toll-like receptor 4 signaling and maintains endotoxin tolerance. Nat. Immunol Apr;5(4):373-9. PubMed ID: 11. Cook DN, Pisetsky DS, Schwartz DA. Toll-like receptors in the pathogenesis of human disease. Nat. Immunol Oct;5(10):975-9. PubMed ID: MyD88s IRAKM IRAK4 ToIIIP TRAF6 MyD88 UbC13 TRAF6 IRAK1 IRAK2 UEV1A T L R 9 ENDOSOME ECSIT TAB2 Triad3 TAB1 T L R 3 TAK1 MKK7 IKKs T L R 7 T L R 8 MKK6 MKK3 MEKK3 IKKs MyD88 IkB IRF7 NF-kB TRIF BTK TRAF6 p38 MyD88 IkB RIP3 IRAK NF-kB TBK1 TRAF6 IFN-a JNK RIP IRF7 CREB IKKe NUCLEUS TANK IFN-b SLAM,CD80, CD83 IRF3 NF-kB IRF8 2009 ProteinLounge.com C C-Jun ATF2 TNF,COX2, IL-18 21 IFN-Responsive Genes