www.<strong>in</strong>vivogen.com/<strong>in</strong>nate-immunity 51 INNATE IMMUNITY 3
INNATE IMMUNITY 3 Nod-Like Receptors NOD-Like Receptors (NLRs, also known as CATERPILLERs) constitute a recently identified family of <strong>in</strong>tracellular pattern recognition receptors (PRRs), which conta<strong>in</strong>s more than 20 members <strong>in</strong> mammals. Although the ligands and functions of many of these receptors are not known, their primary role is to recognize cytoplasmic pathogen-associated molecular patterns (PAMPs) and/or endogenous danger signals also called damageassociated molecular patterns (DAMPs), <strong>in</strong>duc<strong>in</strong>g immune responses. NLRs are characterized by a tripartite-doma<strong>in</strong> organization with a conserved nucleotide b<strong>in</strong>d<strong>in</strong>g oligomerization doma<strong>in</strong> (NOD) and leuc<strong>in</strong>e-rich repeats (LRRs). The general doma<strong>in</strong> structure consists of C-term<strong>in</strong>al LRRs <strong>in</strong>volved <strong>in</strong> microbial sens<strong>in</strong>g, a centrally located NOD doma<strong>in</strong> and an N-term<strong>in</strong>al effector region compris<strong>in</strong>g a prote<strong>in</strong>-prote<strong>in</strong> <strong>in</strong>teraction doma<strong>in</strong> such as the CARD, Pyr<strong>in</strong> or BIR doma<strong>in</strong>. NLRs have been grouped <strong>in</strong>to several subfamilies on the basis of their effector doma<strong>in</strong>s: NODs, NALPs, CIITA, IPAF, and NAIPs. NODs and IPAF conta<strong>in</strong> CARD effector doma<strong>in</strong>s, whereas NALPs and NAIPs conta<strong>in</strong> pyr<strong>in</strong> (PYD) effector doma<strong>in</strong>s and three BIR doma<strong>in</strong>s, respectively. NOD1 and NOD2 The first mammalian NLRs reported to sense <strong>in</strong>tracellular microbial PAMPs are NOD1 (CARD4) and NOD2 (CARD15), which conta<strong>in</strong> one and two N-term<strong>in</strong>al CARD doma<strong>in</strong>s, respectively. They recognize peptidoglycan (PGN), an essential constituent of the bacterial cell wall. NOD1 and NOD2 detect specific motifs with<strong>in</strong> the PGN. NOD1 senses the D-g-glutamyl-meso- DAP dipeptide (iE-DAP) which is found <strong>in</strong> PGN of all Gram-negative and certa<strong>in</strong> Gram-positive bacteria 1,2 whereas NOD2 recognizes the muramyl dipeptide (MDP) structure found <strong>in</strong> almost all bacteria 3 . Thus NOD2 acts as a general sensor of PGN and NOD1 is <strong>in</strong>volved <strong>in</strong> the recognition of a specific subset of bacteria. Both iE-DAP and MDP must be delivered <strong>in</strong>tracellularly either by bacteria that <strong>in</strong>vade the cell or through other cellular uptake mechanisms to be detected by NOD1 and NOD2 respectively. NOD1 and NOD2 signal via the ser<strong>in</strong>e/threon<strong>in</strong>e RIP2 (RICK,CARDIAK) k<strong>in</strong>ase through CARD-CARD homophilic <strong>in</strong>teractions 4 . Once activated, RIP2 mediates ubiquit<strong>in</strong>ation of NEMO/IKKg lead<strong>in</strong>g to the activation of NF-kB and the production of <strong>in</strong>flammatory cytok<strong>in</strong>es such as TNF-a and IL-6 5 . In addition to the NF-kB pathway, NOD1 and NOD2 stimulation <strong>in</strong>duces the activation of MAPKs 6 . Recent work has implicated CARD9 <strong>in</strong> the selective control of NOD2-dependent p38 and JNK signal<strong>in</strong>g 7 . The physiological importance of NOD1 and NOD2 <strong>in</strong> immune responses is evident from the l<strong>in</strong>kage of their mutations with <strong>in</strong>flammatory diseases <strong>in</strong> humans. Genetic variation <strong>in</strong> NOD2 is associated with Crohn’s disease, one of the major forms of <strong>in</strong>flammatory bowel diseases 8 . Several NOD1 polymorphisms are l<strong>in</strong>ked to the development of atopic eczema and asthma 9 . Schematic structure of Lys-PGN (found <strong>in</strong> Gram-positive bacteria) and DAP-PGN (found <strong>in</strong> Gram-negative bacteria) 52 www.<strong>in</strong>vivogen.com/<strong>in</strong>nate-immunity NALPs The NALP subfamily consists of 14 members that are characterized by the presence of PYD effector doma<strong>in</strong>s. Although the functions of many of the NALPs are largely unknown, several NALPs play a key role <strong>in</strong> the regulation of caspase-1 by form<strong>in</strong>g a mutiprote<strong>in</strong> complex known as the ‘<strong>in</strong>flammasome’. Caspase-1 participates <strong>in</strong> the process<strong>in</strong>g and subsequent release of pro<strong>in</strong>flammatory cytok<strong>in</strong>es, such as IL-1b and IL-18. At least two types of NALP <strong>in</strong>flammasomes have been identified: the NALP1 <strong>in</strong>flammasome compris<strong>in</strong>g NALP1 (NLRP1, CARD7), ASC, Caspase-1 and Caspase-5, and the NALP3 <strong>in</strong>flammasome conta<strong>in</strong><strong>in</strong>g NALP3 (NLRP3, cryopyr<strong>in</strong>, CIAS1), ASC, Card<strong>in</strong>al and Caspase-1 10 . NALP1 and NALP3 recruit through their PYD doma<strong>in</strong> the adaptor prote<strong>in</strong> ASC which <strong>in</strong> turn <strong>in</strong>teracts with Caspase-1 via a CARD-CARD <strong>in</strong>teraction. NALP1 also recruits Caspase-5 via its additional CARD effector doma<strong>in</strong> at the C term<strong>in</strong>us, whereas NALP3, lack<strong>in</strong>g such a CARD, <strong>in</strong>teracts with the CARDconta<strong>in</strong><strong>in</strong>g adaptor Card<strong>in</strong>al to recruit additional Caspase-1. Two molecules have been recently described to activate NALP1: MDP and the anthrax tox<strong>in</strong>. In vitro reconstitution experiments revealed that NALP1 oligomerization is a two step mechanism requir<strong>in</strong>g MDP and ATP 11 . In vivo studies with NALP1 knock-out mice are necessary to confirm that MDP is a true NALP1 ligand. A mur<strong>in</strong>e variant of NALP1 (NALP1b) was shown to respond to the anthrax tox<strong>in</strong> suggest<strong>in</strong>g an engagement of the NALP1 <strong>in</strong>flammasome <strong>in</strong> the immune response to Bacillus anthracis <strong>in</strong>fection 12 . NALP3 mediates caspase-1 activation <strong>in</strong> response to a wide variety of stimuli: whole bacteria (Listeria monocytogenes, Staphylococcus aureus), bacterial RNA, synthetic pur<strong>in</strong>e-like compounds (R848, R837), uric acid crystals, extracellular ATP and pore-form<strong>in</strong>g tox<strong>in</strong>s (nigeric<strong>in</strong>, maitotox<strong>in</strong>) 13- 15 . Activation of Caspase-1 <strong>in</strong>duced by NALP3 appears to be TLR<strong>in</strong>dependent, whereas secretion of mature IL-1b seems to require two stimuli <strong>in</strong>volv<strong>in</strong>g the TLRs and NALP3. The first stimulus, a TLR ligand such as LPS, triggers the generation of pro-IL-1b, while the second, a stimulus such as ATP, <strong>in</strong>duces oligomerization and <strong>in</strong>flammasome assembly 16-17 . In addition, studies suggest that NALP3 does not sense ATP directly but rather <strong>in</strong>tracellular potassium depletion result<strong>in</strong>g from ATP signal<strong>in</strong>g. Bacterial tox<strong>in</strong>s, such as nigeric<strong>in</strong> and maitotox<strong>in</strong> that cause a change <strong>in</strong> <strong>in</strong>tracellular ion composition activate the NALP3 <strong>in</strong>flammasome. Thus, NALP3 appears to serve as an activator of the <strong>in</strong>flammasome <strong>in</strong> response to specific tox<strong>in</strong>s, endogenous danger signals released by damaged cells or tissues (uric acid crystal, elevated ATP), or microbial pathogens. Mutations <strong>in</strong> NALP3 are the cause for several human diseases such as familial cold auto<strong>in</strong>flammatory syndrome and Muckle-Wells syndrome 18 .