Pattern Recognition Receptors in Health and Diseases PDF

Signal Transduction and Targeted Therapy www.nature.com/sigtrans REVIEW ARTICLE OPEN Pattern recognition receptors in health and diseases...

Signal Transduction and Targeted Therapy www.nature.com/sigtrans REVIEW ARTICLE OPEN Pattern recognition receptors in health and diseases Danyang Li1,2 and Minghua Wu1,2 Pattern recognition receptors (PRRs) are a class of receptors that can directly recognize the specific molecular structures on the surface of pathogens, apoptotic host cells, and damaged senescent cells. PRRs bridge nonspecific immunity and specific immunity. Through the recognition and binding of ligands, PRRs can produce nonspecific anti-infection, antitumor, and other immunoprotective effects. Most PRRs in the innate immune system of vertebrates can be classified into the following five types based on protein domain homology: Toll-like receptors (TLRs), nucleotide oligomerization domain (NOD)-like receptors (NLRs), retinoic acid-inducible gene-I (RIG-I)-like receptors (RLRs), C-type lectin receptors (CLRs), and absent in melanoma-2 (AIM2)-like receptors (ALRs). PRRs are basically composed of ligand recognition domains, intermediate domains, and effector domains. PRRs recognize and bind their respective ligands and recruit adaptor molecules with the same structure through their effector domains, initiating downstream signaling pathways to exert effects. In recent years, the increased researches on the recognition and binding of PRRs and their ligands have greatly promoted the understanding of different PRRs signaling pathways and provided ideas for the treatment of immune-related diseases and even tumors. This review describes in detail the history, the structural characteristics, ligand recognition mechanism, the signaling pathway, the related disease, new drugs in clinical trials and clinical therapy of different types of PRRs, and discusses the significance of the research on pattern recognition mechanism for the treatment of PRR-related diseases. 1234567890();,: Signal Transduction and Targeted Therapy (2021)6:291 ; https://doi.org/10.1038/s41392-021-00687-0 INTRODUCTION host that can recognize pathogenic microorganisms and activate The first line of defense against pathogens that gradually evolved the second signal in time, which are independent of gene in organisms is innate immunity,1 which is divided into two levels: rearrangement. In vertebrates, innate immunity recognizes first, the skin, mucosal tissue, blood–brain barrier, and chemical pathogenic microorganisms and assists in the activation and barrier (e.g. fatty acid, pH, enzyme, and complement system) of expression of second signals that activate the adaptive the host can effectively resist the invasion of general pathogenic immunity.13 microorganisms;2–4 second, the innate immune system of Toll-like receptors (TLRs) are one of the earliest PRRs discovered vertebrates protects the organism through nonspecific immune in the innate immune system, which plays an important role in defense and surveillance by innate immune cells. Innate immune inflammatory responses.14,15 Therefore, here is a brief description cells mainly include monocytes, neutrophils, macrophages, of the development history of PRRs with TLRs as a representative. dendritic cells, natural killer (NK) cells, mast cells, eosinophils, TLRs were first found in Drosophila in the form of genes in 1994. and basophils.5,6 Unlike T cells and B cells, which have high Studies have shown that the function of this gene is related to the specificity, innate immune cells do not express specific antigen formation of the dorsal–ventral axis during the embryonic recognition receptors. Through the recognition and binding of development of Drosophila.16 In 1988, Hashimoto et al. discovered some common molecules on the surface of pathogens, apoptotic that the Toll gene encodes a transmembrane protein and clarified host cells, and damaged senescent cells, pattern recognition the structure of the Toll protein.17 In 1991, Gay et al. found that receptors (PRRs) induce immunoprotective effects, such as anti- Toll protein had structural homology with interleukin-1 (IL-1), a infection and antitumor effects, and participate in the initiation natural immune molecule in mammals, suggesting that the and effect process of specific immune response.7–9 function of Toll may be related to immunity.18 In 1996, Hoffmann In the 1990s, the hypothesis of pathogen-associated molecular team found that Toll plays a role in the resistance of Drosophila to patterns (PAMPs) and PRRs that recognize PAMPs was proposed fungal infection. Toll-activated mutants persistently express by Janeway, which was of epoch-making significance and antifungal peptides, while Toll-deletion mutants, on the contrary, changed research on innate immunity.10 The main point of this lose their ability to arrest fungal infection. It has been found that hypothesis is the connection between the innate immune signal Toll can recognize spatzle (an important protein in the develop- and the initiation of the adaptive immune response. Some unique ment of the dorsal and abdomen of Drosophila) and initiate a and conserved components of pathogenic microorganisms can series of signal transduction to activate the expression of induce the second signal required to activate T cells, so as to antifungal peptide.19 In 1997, Janeway et al. cloned human control the adaptive immunity from being activated under normal TLR4. TLR4 can induce the activation of nuclear factor (NF)-κB and conditions.11,12 In addition, there are a class of receptors in the the expression of the co-stimulatory molecule CD80. This proves 1 Hunan Provincial Tumor Hospital and the Affiliated Tumor Hospital of Xiangya Medical School, Central South University, Changsha, Hunan, China and 2The Key Laboratory of Carcinogenesis of the Chinese Ministry of Health, The Key Laboratory of Carcinogenesis and Cancer Invasion of the Chinese Ministry of Education, Cancer Research Institute, Central South University, Changsha, Hunan, China Correspondence: Minghua Wu ([email protected]) Received: 1 September 2020 Revised: 23 May 2021 Accepted: 22 June 2021 © The Author(s) 2021  Pattern recognition receptors in health and diseases Li and Wu 2 Table 1. Common PRRs in innate immunity Items PRR Domains Cellular PAMP Sources Signaling pathways distribution Toll-like TLR1 LRR Mo, DC, Triacyl lipopeptide Bacteria Most TLRs: MyD88- receptors (TLRs) (TLR1–TLR2) domain–transmembrane Ma, Eo, Ba dependent pathways; TLR2 domain–TIR domain Mo, DC, Lipoteichoic acid Bacteria TLR3: TRIF-dependent (TLR1–TLR2, (extracellular to Ma, Eo, Ba pathways; TLR4: MyD88- intracellular) Arabinomannan Mycobacterium dependent pathways and TLR2–TLR6) Peptidoglycan Bacteria TRIF-dependent pathways Zymosan Fungi Lipoprotein Mycoplasma Pore protein Neisseria TLR3 Mφ, DC, IEC dsRNA Virus TLR4 (MD-2/ Mφ, Lipopolysaccharides Bacteria CD14) DC, Ma, Eo Heat-shock proteins Host TLR5 IEC Flagellin Bacteria TLR6 Mo, DC, Lipoteichoic acid Bacteria (TLR2–TLR6) Ma, Eo, Ba Peptidoglycan Bacteria TLR7 pDC, ssRNA Virus Mφ, Eo Imidazoquinoline Artificially synthesized TLR8 Mφ, N ssRNA Virus TLR9 pDC, Eo, Ba Non-methylated Bacteria, Virus CpG DNA TLR10 pDC, Eo, Ba dsRNA Virus (human) TLR11 Mφ, DC Profilin and related Toxoplasma (mouse) proteins gondii TLR12 DC Profilin and related Toxoplasma (mouse) proteins gondii TLR13 Unknown 23s ribosomal RNA Bacteria (mouse) Nucleotide- NOD1 LRR domain–NBD–effector IEC, cytosol iE-DAP Gram negative RIP2-TAK1-NF-κB pathways binding domains of Mφ bacteria oligomerization NOD2 MDP Gram-negative domain-like bacteria, Gram- receptors (NLRs) positive bacteria RIG-I-like RIG-I (RD)-CTD-DexD/H helicase Cytosol 5’-triphosphorylated Virus MAVS-TRAF6-NF-κB/TBK1 receptors (RLRs) domain–CARD RNA, short- pathways chain dsRNA MDA5 poly IC, long- Virus chain dsRNA LGP2 dsRNA Virus C-type lectin Dectin-1 CTLD–ITAM DC, Mφ β-Glucan Fungus Tyrosine kinase-dependent receptors (CLRs) Dectin-2 α-Mannan Fungus and non-tyrosine kinase- dependent pathways Absent in ALRs HIN-200-PYD Cytosol dsDNA Bacteria Inflammasome–pyroptosis melanoma-2-like receptors (ALRs) LRR leucine-rich repeat, TIR Toll/IL-1R domain, NBD nucleotide-binding domain, RD repressor domain, CTD C-terminal domain, CARD caspase activation and recruitment domain, CTLD C-type lectin-like domains, ITAM immunoreceptor tyrosine-based activation motif, PYD pyrin domain, Mo monocyte, DC dendritic cell, Ma mastocyte, Eo eosinophils, Ba basophils, pDC plasmacytoid dendritic cell, IEC intestinal epithelial cell, N neutrophil, dsRNA double-stranded RNA, ssRNA single-stranded RNA, iE-DAP γ-D-glu-meso-diaminopimelic acid, MDP muramyl dipeptide, MyD88 myeloid differentiation factor 88, TRIF TIR domain-containing adaptor protein-inducing interferon β, RIP2 receptor-interacting serine–threonine protein 2, TAK1 transforming growth factor-β-activated kinase 1, NF-κB nuclear factor κB, MAVS mitochondrial antiviral signaling protein, TRAF6 tumor necrosis factor receptor-associated factor, TBK1 TANK-binding kinase 1 that innate immunity recognizes pathogenic microorganisms and oligomerization domain (NOD)-like receptors (NLRs), retinoic acid- activates the expression of the second signal, which is indis- inducible gene-I (RIG-I)-like receptors (RLRs), C-type lectin recep- pensable for the activation of adaptive immunity.20 Since the tors (CLRs), and absent in melanoma-2 (AIM2)-like receptors (ALRs) discovery of TLR4, many PRRs and their corresponding ligands (Table 1).21 PRRs are representative of immune receptors in innate have been discovered. PRRs can be divided into the following five immunity and exist in various forms. PRRs are not only expressed types based on protein domain homology: TLRs, nucleotide on the cell membrane but also widely distributed in intracellular Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 3 Fig. 1 The signal transduction pathways and structure of TLR-binding ligand complex. TLRs can recognize one or more PAMPs through LRR domain. They usually dimerize themselves and recruit adaptor molecules with the same TIR domain to transmit signals compartment membranes and the cytoplasm.22 Membrane- PRRS AND LIGAND-RECOGNITION MECHANISMS bound PRRs and PRRs in the cytoplasm are basically composed Toll-like receptors of ligand recognition domains, intermediate domains, and effector TLRs are membrane-bound signal receptors and are important domains.23,24 PRRs activate downstream signaling pathways PRRs in the innate immune system of vertebrates.15,33 Such through recognition of their ligands. The activation of down- receptor molecules usually have two functions, one is to bind stream signaling pathways can produce many effects: recruiting specifically to the ligand, and the other is to transmit signals. The and releasing cytokines, chemokines, hormones, and growth corresponding signal transduction will amplify the effect of anti- factors; inducing chronic inflammation; forming an inflammatory pathogen infection, so that the immune cells active in the microenvironment; initiating innate immune killing and subse- inflammatory response can be activated through the transcription quent acquired immune response,9 maintaining the balance of of genes, and produce and secrete a variety of pro-inflammatory host microecology; and eliminating dead or mutated cells. and antiviral factors.34–36 Up to now, 10 functional TLRs (TLR1–10) PAMPs are the specific and highly conserved molecular have been found in humans and 12 (TLR1–9 and TLR11–13) in structures shared by the same kind of pathogenic microorgan- mice.37–41 TLR10 in mice is not functional due to the insertion of isms,25,26 including lipids, proteins, and nucleic acids, such as reverse transcriptase.42 TLRs recognize PAMPs in different lipopolysaccharides (LPS), lipoteichoic acid (LTA), and bacterial subcellular structures. The cellular localization of TLRs determines DNA.27,28 PAMPs are essential for pathogen survival and usually the types of ligands and the recognition mechanism. Some TLRs have unique molecular or subcellular characteristics that are not (TLR1, 2, 4, 5, 6, 10) are expressed on the surface of immune cells found in host cells. Therefore, innate immune cells can recognize in the form of heterodimers or homodimers, mainly recognizing PAMPs via PRRs, distinguish “self” and “non-self,”29 and respond to the membrane components of pathogenic microorganisms, such pathogens and their products. However, the host will produce as lipids, lipoproteins, and proteins; others (TLR3, 7, 8, 9) are some proteins and metabolites after being stimulated by its own expressed in the form of homodimers, which mainly recognize the tissue damage, cell necrosis, and other factors.30 These molecules nucleic acids of microorganisms (Fig. 1).43 are called damage-associated molecular pattern (DAMP).31 PRRs TLRs are type I transmembrane glycoproteins and are com- can also recognize such molecules, activate natural immunity, and posed of an extracellular region, a transmembrane region, and an cause inflammation.32 intracellular region.44 The extracellular region contains leucine-rich With advances in research on PRRs structure and distribution at repeats (LRRs), which are responsible for the recognition of different levels, the role of PRRs in the innate immune regulatory specific ligands and perform extracellular pattern recognition. The network has become clearer. The recognition and binding of PRRs intracellular domain contains the same Toll/IL-1R (TIR) domain as to their ligands is critical in initiating the innate immune response. IL-1R, which plays a role in signal transduction. The extracellular Therefore, the study of PRR-mediated pattern recognition region of TLRs contains LRRs, which mediate the pattern mechanisms will help to elucidate the signaling pathways and recognition of TLRs (Fig. 1).45 In 2007, researchers used X-ray mechanisms of disease and provide new targets and methods for crystal diffraction to analyze and determine the structure of the the treatment of diseases. In this review, we describe the structural TLR–ligand complex,46 which provided a deeper understanding of characteristics, ligand recognition mechanism, the signaling the LRR domain. The LRR domain is shaped like a horseshoe, and pathway, the related disease, new drugs in clinical trials, and each module consists of a conserved leucine motif and a variable clinical therapy of different types of PRRs in detail. We focus on the region. The “LxxLxLxxN (L leucine, x any amino acid, N different domains and ligand recognition mechanisms between asparagine)” motif is composed of 20–30 amino acids and is on PRRs, which can not only provide new ideas for the definition, role, the concave surface of the horseshoe-like structure.47–50 The and clinical application of PRRs but also promote the study of the horseshoe-shaped N-terminus and C-terminus contain disulfide role of the innate immune system in related diseases and even bridges formed by cysteine clusters51,52 to protect the hydro- tumors. phobic core. After TLRs recognize and bind the corresponding Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 4 Fig. 2 Crystal structure of TLRs with ligands. a Crystal structure of the TLR1–TLR2 heterodimer induced by binding of a tri-acylated lipopeptide (PDB 2Z7X). TLR2 initiates immune responses by recognizing di-acylated and tri-acylated lipopeptides. The ligand specificity of TLR2 is controlled by whether it heterodimerizes with TLR1 or TLR6. Binding of the tri-acylated lipopeptide (red) induced the formation of M- type crystal structures of the TLR1 (pale yellow) and TLR2 (slate) ectodomains. b Crystal structure of TLR2–TLR6–Pam2CSK4 complex (PDB 3A79). Binding of the di-acylated lipopeptide, Pam2CSK4 (red), induced the formation of M-type crystal structures of the TLR2 (slate) and TLR6 (pale green) ectodomains. c Crystal structure of mouse TLR4/MD2/LPS complex (PDB 3VQ2). After LPS (red) binds with the TLR4 (yellow)/MD2 (gray) complex, the hydrophobic pocket of MD2 is used to bridge the two TLR4–MD2–LPS complexes to form a spatially symmetrical M-type structure. Mouse TLR4/MD2/LPS exhibited an complex similar to the human TLR4/MD2/LPS complex. d Crystal structure of the N-terminal fragment of zebrafish TLR5 in complex with Salmonella flagellin (PDB 3V47). Two TLR5 (cyan)–flagellin (firebrick) 1:1 heterodimers assemble into a 2:2 tail-to-tail signaling complex to function PAMPs and endogenous ligands, the TIR domains conduct signals module (Fig. 2a, b).63 Researches on the structure of ligand by binding to different receptor adaptor proteins in the complexes can significantly promote the discovery of small cytoplasmic region.53,54 The TIR domain has three conserved molecule agonists/antagonists targeting PRRs. A recent study amino acid sequences, which are called 1,2,3 cassettes. Depending revealed the activation mechanism of atypical agonists for on the different adaptor proteins, TLRs signaling can be divided TLR1–TLR2. Diprovocim is a recently found small molecule into myeloid differentiation factor 88 (MyD88)-dependent and activator for TLR1–TLR2, but it has no structural similarity with MyD88-independent pathways (Fig. 1).55,56 the tri-acylated lipopeptide complex. It also interacts with Exploring the pattern recognition mechanisms of TLRs is very TLR1–TLR2 in the same binding pocket as typical lipopeptide valuable for understanding innate immunity and some tumor- ligand.64 Crystal structure analysis revealed that double-stranded igenesis mechanisms. Therefore, researchers used X-ray crystal RNA (dsRNA) binds to the LRR domains of the N-terminus and C- diffraction to determine the crystallographic structure of the terminus of TLR3.65,66 Different from the way that other TLRs extracellular domain of TLRs and the ligand complex. Although directly recognize ligands,67–69 TLR4 specifically recognizes LPS in the ligand complexes have different structures, all these combination with two auxiliary molecules, myeloid differentiation complexes have similar M-type crystal structures (Fig. 2).50,51 factor 2 (MD2) and the LRR structural protein CD14. LPS is TLR1 or TLR6 can form TLR1/TLR2 and TLR6/TLR2 heterodimers transported by LPS-binding protein to CD14 on the cell membrane with TLR2 to recognize tri-acylated lipopeptide and di-acylated of monocytes and macrophages to form a complex and then lipopeptide,57 respectively. After recognizing the appropriate interacts with TLR4/MD2.70 After LPS binds with the TLR4/MD2 ligands, TLR2 can form an M-type structure with the extracellular complex, the hydrophobic pocket of MD2 is used to bridge the region of TLR1 and TLR6, and the pocket structure formed binds to two TLR4–MD2–LPS complexes to form a spatially symmetrical M- the ligand.58–61 The crystal structure of TLR1–TLR2–tri-acylated type TLR4–MD2–LPS dimer,71 and then conformational changes lipopeptide complex is similar to that of TLR2–TLR6–di-acylated affect their respective functional domains and transmit signals lipopeptide complex, but there are important structural differ- (Fig. 2c). In addition to binding to LPS, TLR4 is also involved in the ences between TLR1 and TLR6 in the ligand-binding site and recognition of natural products (carnosic acid, paclitaxel) and dimerization surface. The ligand-binding pocket of TLR1–TLR2 is pneumolysin.72–74 TLR5 is the most conserved and important PRR, located in the interface between the central and C-terminal which is usually stimulated by bacterial flagellin. In the form of domain, and TLR1–TLR2–tri-acylated lipopeptide is stabilized by homodimer, TLR5 plays a major role in the primary defense of non-covalent bonds such as hydrogen bonds, hydrophobic invasive pathogens and immune homeostasis regulation.75 interactions, and ionic interactions near the ligand-binding Although the heterodimeric structure of TLR5a–TLR5b in zebrafish pocket.62 In TLR6, the side chains of amino acid residues block and the crystallographic structure of TLR5–flagellin complex has the ligand-binding pocket, resulting in a pocket less than half the been clearly reported, the lack of biochemical and structural length of TLR1. In addition, the TLR2–TLR6 heterodimer is mainly information of fish TLR hinders the understanding of flagellin- regulated by the surface exposed residues of the LRR11–14 based therapies. In the future, experimental TLR5–flagellin Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 5 but it must be a complete viral ssRNA.92 The basic process of NOD2 activation and signal transduction is as follows: after pathogenic bacteria are phagocytosed by macrophages, they first form phagosomes, and then fuse with lysosomes to become phagolysosomes. Under the action of lysosomal enzymes, bacterial cell wall components are decomposed into peptidogly- can, which can be degraded into a cell wall peptide with immunomodulatory activity and enter the cytosol, thereby activating NOD2.93 In general, the LRR domain of the NLR molecule folds to form a U-shaped configuration with the central NACHT domain, which inhibits its multimerization and makes the NLRs inactive.94 Once PAMPs directly or indirectly bind to the LRRs, the NLR molecule change their conformation, exposing the NACHT oligomerization domain, which triggers oligomeriza- tion, and the NLR molecule is activated.95 At the same time, the N- Fig. 3 The ligand recognition mechanism of NLRs. The combination terminal effector domain is exposed, and through homotypic of PAMP and LRR changes the conformation of NLRs from self- inhibition to activation interactions, downstream adaptor molecules and signaling proteins with the same structure are recruited to initiate the corresponding signal transduction (Fig. 3).96 Although NOD1 and complex structure modeling and computational simulation should NOD2 do not have transmembrane domains, studies have shown be used to study flagellin-mediated interactions between various that they are recruited into the plasma membrane and endosomal pathogens and host immune receptors (Fig. 2d).76 It has been membrane, which is necessary for signal transduction.97 In this reported that TLR1–6 each exist as monomers in solution, and process, palmitoylation plays a vital role. The modification of dimerization occurs only when the ligand is bound; in contrast, NOD1/2 protein under the action of palmitoyltransferase ZDHHC5, TLR8 and TLR9 exist as preformed dimers, and the binding of which makes NOD1/2, possess the characteristics of rapid and ligands induces conformational changes in preformed dimers (Fig. reversible localization changes, which is necessary for membrane 1).49,77,78 Lee revealed that TLR10 binds dsRNA in vitro at recruitment and inflammatory signal transduction.98 This study endosomal pH, indicating that dsRNA is a ligand of TLR10. The gives us a good enlightenment that the modification of PRRs may recognition of dsRNA by TLR10 recruits MyD88, thereby transdu- play a key role in the regulation of host innate immune signal. cing signals and inhibiting interferon regulatory factor 7 (IRF7)- dependent type I interferon (IFN) production.79 In mice, TLR11 and RIG-I-like receptors TLR12 are the main effector molecules to recognize Toxoplasma RLRs are also intracellular PRRs. In innate antiviral immunity, in gondii. The recognition of T. gondii profilin by TLR11 depends on addition to the recognition of viral nucleic acids by TLR7 and TLR9, the parasite-specific, surface-exposed motif in TgPRF consisting of most other types of cells recognize viral nucleic acids through an acidic loop and a β-hairpin.80–82 RLRs to induce antiviral immune responses.99,100 The currently discovered RLR family members mainly include three: RIG-I, NOD-like receptors melanoma differentiation-associated gene 5 (MDA5), and labora- The growth cycle of some pathogenic microorganisms involves tory of genetics and physiology 2 (LGP2) (Fig. 4).101 infection of the cytoplasm. For example, viral genes are often RIG-I was first discovered in acute promyelocytic leukemia cells transcribed and translated in the cytoplasm, and virus particles are induced by retinoic acid. In 2004, it was found that RIG-I could assembled. In addition, some bacteria and parasites have a series induce the expression of a reporter gene in the IFN-β promoter of escape mechanisms, such as making holes in the phagosome region, which confirmed its antiviral activity.102 The structure of membrane and entering the cytoplasm. Therefore, pathogens and the RIG-I protein consists of three parts.103–105 The middle part is their components, as well as other components produced by the DexD/H helicase domain, which is the common domain of the infection and injury, will appear in the cytoplasm,83 which requires RLR family, and has ATPase and helicase activities.106–108 The N- the recognition of PRRs in the body. NLRs are intracellular PRRs, terminus of the RIG-I protein is composed of two caspase composed of three domains:84,85 one is the central nucleotide- activation and recruitment domains in series,109 which are binding domain (NBD), also known as the NACHT domain responsible for transmitting signals downstream.110 The C- (synthesized by the abbreviations of the following four kinds of terminus is composed of the repressor domain (RD) and the C- NLR members: NAIP, CIITA, HETE, TP1), which is shared by the NLR terminal domain (CTD), which can regulate its own state.106,111 The family and is very important for nucleic acid binding and former can inhibit the activation of the receptor, and the latter is oligomerization of NLRs (Fig. 3); LRRs at the C-terminus, which responsible for the recognition of viral RNA.112,113 In the resting are used to identify ligands; and the N-terminal effector domain, state, CARD, CTD, and the helicase domain are folded, and RIG-I is which is the protein interaction domain, such as the caspase in a self-inhibited state. During viral infection, the CTD of RIG-I activation and recruitment domain (CARD) or the pyrin domain recognizes viral RNA and undergoes a conformational change.114 (PYD).86–89 According to the different N-terminal effector domains, RIG-I uses ATP hydrolase activity to expose and activate the CARD the NLRs family can be divided into five subfamilies: the NLRC and multimerize, thereby recruiting downstream signaling linker subfamily, which contain CARDs; the NLRP subfamily, which molecules (Fig. 4).115–117 contain PYDs; the NLRB subfamily, which contain baculovirus The structure and functions of MDA5 are similar to those of RIG-I, inhibitor of apoptosis protein repeats; the NLRA subfamily, which with the DexD/H helicase domain in the middle, two CARD at the contain acidic activation domains; and the NLRX subfamily N-terminus, and a CTD at the C-terminus; however, MDA5 lacks the containing other NLR effector domains.85 RD, and so it does not have self-inhibitory functions. In contrast to Among the NLRs family, the most in-depth study has focused other RLRs, LGP2 does not have CARD,118,119 and so it cannot on NOD1 and NOD2 proteins. NOD1 mainly recognizes the recruit molecules of the same structure to transmit signals, but it diaminopimelic acid (γ-D-glu-meso-diaminopimelic acid (iE-DAP)) can regulate the recognition of viral nucleic acids by RIG-I and of the cell wall of Gram-negative bacteria.90,91 In addition to MDA5, thereby preventing RLR-mediated resistance.120–123 LGP2 recognizing muramyl dipeptide (MDP) in all bacterial cell walls, can negatively regulate RIG-I-mediated recognition of viral dsRNA, NOD2 can also recognize single-stranded RNA (ssRNA) of the virus, reduce the production of IFNs and inflammatory factors, and Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 6 Fig. 4 Structural features and ligand recognition mechanism of RLRs. The structure and functions of MDA5 are similar to those of RIG-I. However, MDA5 lacks the repressor domain, so it does not have self-inhibitory functions. LGP2 does not have CARD, and so it cannot transmit signals. The combination of viral RNA and CTD changes the conformation of RLRs ultimately inhibit the antiviral innate immune response.124 LGP2 is C-type lectin receptors also critical in the antiviral response mediated by MDA5.125 LGP2 CLRs, which belong to phagocytic PRRs, are also a popular type of exhibits a concentration-dependent conversion between MDA5- receptor under study.143 The function of phagocytic receptor is specific enhancement and interference.126 The latest research different from the receptor that activates cells by signal revealed a mechanistic basis for LGP2-mediated regulation of transduction. It recognizes and binds to PAMPs through PRRs MDA5 antiviral innate immune responses. LGP2 facilitates MDA5 and places pathogens in cytoplasmic vesicles for direct digestion fiber assembly and is incorporated into the fibers, forming hetero- and elimination to control infection.144 CLRs are a class of oligomers with MDA5.127 In addition, LGP2 can significantly induce receptors that recognize carbohydrates on the surface of the exposure of the CARD domain of MDA5.128 Under bacterial pathogenic microorganisms with the participation of Ca+.145 It is infection of the Indian major carp Labeo rohita, LGP2 gene expressed on macrophages, dendritic cells (DCs), and certain expression was significantly increased after dsRNA and various tissue cells. The ability of CLRs to recognize carbohydrates existing PAMPs were stimulated, indicating that LGP2 can act as an antiviral on self and non-self structures is mediated by carbohydrate and antibacterial cytoplasmic receptor.129 recognition domain (CRD).146 The CRD of CLRs is a compact Although the RLR family members have similar structures, they spherical structure, and this region is called C-type lectin-like recognize the RNA of different viruses through ligand-recognition domain (CTLD).147,148 Depending on the location of the protein on domains.130 Both RIG-I and MDA5 can recognize viral dsRNA, but the cell membrane, CLRs are divided into transmembrane their recognition depends on the length of the dsRNA.131 RIG-I receptors and secretory receptors.146,149,150 The main representa- mainly recognizes viruses with relatively short dsRNA (1000 bp).132 “Extracellular pattern recognition molecules”).151 Transmembrane Additionally, RIG-I mediates the antiviral response by recognizing receptors can be divided into type I and type II according to their the 5’-triphosphate RNA of viruses.133 The 5’-terminal triphosphate topological structure.152,153 The N-terminal of type I receptors group can be recognized by RIG-I as a non-self component, but points to extracellular and contains multiple CRDs, while the N- after posttranslational modification, this molecule cannot be terminal of type II receptors points to intracellular and contains recognized by RIG-I.134 Because host cell RNA needs to undergo only one CRD.154,155 It has been shown that the vast majority of different degrees of processing and modification after synthesis in CLRs are involved in the presentation of antigens as active the nucleus, these results indicate that RIG-I can distinguish viral membrane-associated receptors, and CLRs are mainly expressed dsRNA from endogenous RNA. In the cell, RIG-I mainly recognizes on antigen-presenting cells such as DCs and macrophages.145 influenza virus,135 vesicular stomatitis virus,136 Sendai virus, and CLRs are circular structures connected by two disulfide bonds.156 Japanese encephalitis virus,137,138 while MDA5 mainly recognizes CLRs contain at least one CTLD outside the cell, while the small RNA viruses, such as poliovirus.139,140 MDA5 also participates intracellular domain is different. in the synthesis of the dsRNA analog polycytidylic acid (poly I:C). Mannose receptors (MRs) belong to membrane CLRs, which are Previous studies have shown that filamentous fibers are formed single-chain transmembrane molecules.157–159 The extracellular during the recognition of ligands by RIG-I and MDA5, and segment of MR consists of two parts: one is the proximal membrane signaling pathways are initiated from the tail and inside of the end with eight consecutive CTLDs, which is responsible for the viral dsRNA, respectively.141 endocytosis and transport of the ligand; the other is the distal Although it is mentioned in the “Toll-like receptors” section that membrane end of the cysteine-rich lectin domain, which recognizes TLR3, TLR7, TLR8, and TLR9 specifically recognize virus-derived sulfation of carbohydrate conjugates.160 The endogenous ligands of nucleic acid molecules and bacterial nuclear components, they MR are lysosomal hydrolase and myeloperoxidase, as well as the mainly appear in the endosomal membrane. RLRs can not only be mannan-rich structure expressed by pathogens.161,162 expressed in cells infected by various viruses but also can directly Dendritic cell-associated C-type lectin (Dectin)-1 and Dectin-2 recognize and perceive the virus products and virus particles that are typical representatives of the CLR family.163,164 Dectin-1 is a exist in the cytosol. Its antiviral significance cannot be ignored.142 type II transmembrane protein expressed in DCs, macrophages, Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 7 neutrophils, and monocytes.165 The extracellular region is a CTLD. collectin, and ficolin.203 They generally function in two ways: The intracellular tail is connected to an immunoreceptor tyrosine- one is that they recognize various pathogenic factors and based activation motif (ITAM),166 indicating that the receptor also eliminate them through complement activation,204,205 opsoniza- has a signal transduction function. Dectin-1 can identify a variety tion,206 aggregation, and neutralization of inflammatory regula- of fungi,167 including yeast,168 Candida albicans,169,170 Pneumo- tion; the other is that they interact with cell-related PRRs and cystis carinii,171,172 Cryptococcus,173,174 and Aspergillus.175,176 The regulate their functions to jointly regulate innate immune ligand of Dectin-1 is β-1,3-glucan, which can activate downstream response.207 signals through tyrosine kinase-dependent and tyrosine kinase- Pentraxin is characterized by the aggregation of five molecules independent pathways after recognition and binding of the and is highly conserved in evolution, including two families of ligand.177,178 Glycosylation is an important modification of the short molecules and long molecules.208–211 The family of short posttranslational modification of proteins (including antibo- molecules is called acute phase proteins, which is represented by dies),179 which can significantly change the structure and function C-reactive protein (CRP)212–214 and serum amyloid P compo- of proteins or antibodies, so it is also a key mechanism for the nent215,216 in humans and mice, respectively. These molecules are immune system to regulate biological activity.180 Abnormal mainly produced by the liver under the stimulation of inflamma- glycosylation is usually associated with malignant tumors.179 tory signals and interleukins. They are non-specific proteins that Therefore, the identification of molecules that bind glycosylated reflect the systemic inflammatory response. Serum levels increase glycans can provide a new way for the treatment of human rapidly after the body is infected or injured. CRP generally binds to infectious and malignant diseases. Studies have found that Dectin- phosphocholine expressed on the surface of pathogenic micro- 1 can recognize aromatic amino acids adjacent to the N-terminal organisms in a Ca+-dependent manner.217 SAA can bind to the asparagine at the glycosylation site as well as the core fucose on outer membrane protein A of bacteria and interact with IgG antibodies, which do not compete for the same protein TLRs.218,219 In clinic, SAA and CRP are usually used as auxiliary binding site for β-glucan, so Dectin-1 can regulate the immune diagnostic indicators for infectious diseases, but studies have response induced by IgG by combining with core fucose.181 shown that they also have diagnostic value in non-infectious Dectin-2, which is different from Dectin-1, does not contain the diseases and can be used as disease classification markers.220,221 ITAM sequence and has no signal transduction function.182 Dectin- The representative of the pentraxin long molecule family is 2 mainly recognizes α-mannan in the fungal cell wall and PTX3,222 which is unique in that it has a long N-terminal domain. recognizes the Schistosoma mansoni egg antigen.183,184 The PTX3 is produced by dendritic cells, monocyte macrophages, molecular mechanism by which Dectin-2 recognizes the binding epithelial cells, smooth muscle cells (SMCs), and endothelial cells ligand has always been the focus of research. Decout et al.185 under the regulation of a variety of inflammatory factors.223 PTX3 found that the stimulation of Dectin-2 by purified Mycobacterium is involved in the defense of selected pathogens and the tuberculosis mannose-capped lipoarabinomannan requires the regulation of inflammation.224–226 Due to its expression increases (α1 → 2)-linked mannosides forming the cap. Besides, Dectin-2 sharply under the conditions of inflammatory stimulation, PTX3 can also recognize lipoglycans from other bacterial species.185,186 can become a biomarker of general acute inflammation and a From the perspective of the relationship between the structure variety of tumors.227 In coronavirus disease 2019 (COVID-19) and function of the above two ligands, dimannoside caps and patients, circulating and lung bone marrow monocytes and multivalent interaction are necessary for Dectin-2 to recognize endothelial cells express high levels of PTX3, and PTX3 plasma binding ligands and conduct signals.187 concentration can serve as an independent strong prognostic indicator of short-term mortality in COVID-19.228,229 AIM2-like receptors Collectin mainly includes mannose-binding lectin (MBL) and ALRs are a new type of PRRs that can recognize intracellular surfactant protein (SP).151,230 MBL is formed by connecting DNA.188,189 The C-terminus is the DNA-binding domain HIN-200, multiple homotrimers. Each component of the trimer includes a and the N-terminus is the PYD.189–192 The HIN-200 domain CRD, an alpha helix, and a main stem formed by spirals of recognizes double-stranded DNA and binds to it. The N-terminal collagen.231,232 The main stem of collagen gathers each trimer into PYD binds to the PYD of apoptosis-associated speck-like protein bundles. MBL is composed of six CRDs.151 The end of CRD can containing CARD (ASC),193,194 thereby promoting the formation of identify the sugar structure on the surface of various pathogens, inflammasomes and the maturation and release of IL-1β and IL- such as mannose, fucose, glucose, etc.233–235 The pathogens 18.195 Both the DNA-binding affinity of AIM2 and the activity of its involved include yeast, parasites, Gram bacteria, and so on.236–240 inflammasome depend on dsDNA, and it can assemble into When the distance of each CRD between the same trimer or filamentous structures along dsDNA. However, without dsDNA, it adjacent trimers is 45 Å, it is most conducive to ligand binding.241 can also form filaments at high protein concentrations.196–198 ALRs The other family members include A and D,242 which exist on the can not only participate in the innate immune response but also surface of the alveoli and are important innate immune defense regulate apoptosis, which is related to the occurrence and molecules in the lungs. Both of them are composed of N-terminal development of tumors.199 region, CRD, neck region, collagen-like region, and other parts.243 CRD recognizes and binds glycosyl groups. The biological Extracellular soluble pattern recognition molecules significance is that they can selectively identify microbial The initiation of the innate immune response depends on the carbohydrate structures that are harmful to themselves.244,245 recognition of PAMPs by pattern recognition molecules (PRMs), The domain of ficolin is similar to collectin, but it recognizes a including cell PRRs and extracellular soluble PRMs. They are a class variety of bacteria with a fibrinogen-type carbohydrate recogni- of free receptors that can play an antibacterial effect in serum.200 tion structure.246,247 Its ligands are N-acetylglucosamine and LTA, a Although the pattern recognition of innate immunity does not cell wall component of Gram-positive bacteria.248,249 have the antigen specificity of the adaptive immune response, some PRMs produced by the body after infection by pathogenic microorganisms will exist in the serum. Once the new pathogens SIGNALING PATHWAYS OF PRRS invade, they can also bind to the pathogen like an antibacterial There are three main types of molecules involved in signal molecule and play an effective function. Unlike cell-related PRRs, transduction: protein kinases, adaptor proteins, and transcription extracellular soluble PRMs are an important part of non-specific factors. Although PRRs are activated by their respective ligands in humoral immunity.201 Extracellular soluble PRMs are composed of different subcellular structures with different mechanisms, the different molecular families, mainly including pentraxin,202 three main types of molecules involved in signal transduction Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 8 Fig. 5 Pattern recognition receptor-mediated NF-κB signaling. The NF-κB protein can regulate gene expression and affect various biological processes, including innate and adaptive immunity, inflammation, stress response, B cell development, and lymphoid organ formation. TLRs, NLRs, RLRs, and CLRs can generally phosphorylate IκB protein, which inhibits the activation of NF-κB protein, thereby promoting the transcription and activation of inflammatory genes have similar structures and functions, and the signals they receptors are activated, self-dimerize, and recruit downstream transmit are cross-talking, which can converge into several receptor-interacting serine–threonine protein 2 (RIP2) through its common signaling pathways. CARD.268 Activated RIP2 gathers downstream TAK1, TAK1-binding protein 1, and the NF-κB essential modulator/IKKα/IKKβ complex, The NF-κB signaling and the former activates IKKα/IKKβ,269 thereby activating the The transcription factor NF-κB is named after it was first transcription of NF-κB and promoting the release of pro- discovered to be involved in the transcription of B cell κ chain inflammatory factors. genes.250 NF-κB is a heterodimer composed of two molecules, p50 When virus invades cells, RIG-I and MDA5 recognize the and p65, and is inactive due to binding to the inhibitory protein corresponding viral RNA through the CTD and undergo con- IκB under normal conditions. NF-κB plays a key role in the process formation changes.270 Activated RIG-I and MDA-5 induce down- of cellular inflammation and immune response,251,252 and its stream signal transduction by binding with mitochondrial antiviral mediated signal pathways are commonly seen in the activation of signaling protein (MAVS). MAVS is an important adaptor protein various immune cells, including signal transduction initiated by for downstream signal transduction. The N-terminus contains a PRRs in innate immunity (Fig. 5).253 CARD-like domain, which binds to RIG-I and MDA-5 through the In the signal transduction initiated by TLRs,45 after TLRs CARD–CARD interaction.271,272 The proline-enriched domain in recognize and bind the corresponding PAMPs and DAMPs, the MAVS can interact with a series of downstream signaling TIR domains conduct signals by binding to different receptor molecules, such as TRAF3 and 6,273 and activate the protein adaptor proteins in the cytoplasmic region.254,255 Depending on kinase IKK, which causes phosphorylation of IκB,265 and then IκB is the different adaptor proteins, TLR signaling can be divided into ubiquitinated and degraded by proteases, activating the NF-κB MyD88-dependent and MyD88-independent pathways.256 MyD88 pathway.274 has a TIR domain at the C-terminus and a death domain at the N- Different from other typical PRR-mediated signaling pathways, terminus and is the linker molecule in most TLR signal transduction spleen tyrosine kinase (Syk) can be activated by associating with pathways.257 The current research indicated that, in the MyD88- the phosphorylated ITAM motif of CLRs.275 In the Dectin-1/Syk dependent pathway, MyD88 signaling mainly leads to the pathway, Syk activates protein kinase C-δ, which mediates the production of pro-inflammatory cytokines, such as tumor necrosis phosphorylation of CARD9.276 This allows CARD9 to bind to B cell factor (TNF), IL-6, IL-1, and chemokines.258–260 The C-terminus of lymphoma 10277 and para-aspase mucosa-associated lymphoid MyD88 binds to the intracellular TIR domain of TLRs, and the N- tissue lymphoma translocation protein 1, forming a three terminus of MyD88 recruits IL-1R-related kinase 4 (IRAK4)261 and molecular structure that can typically activate NF-κB.278 activates IRAK1 and IRAK2 through autophosphorylation of its central kinase domain. Then ubiquitin ligase TNF receptor- The mitogen-activated protein kinase (MAPK) signaling associated factor 6 (TRAF6) is recruited to form a complex with MAPK is a group of serine–threonine protein kinases that can be transforming growth factor (TGF)-β-activated kinase 1 (TAK1) and activated by different extracellular stimuli,279 such as cytokines, two TAK-binding proteins (TAB1 and TAB4). TRAF6 is degraded due neurotransmitters, hormones, cell stress, and cell adhesion. The to its own ubiquitination.262,263 The TAK1–TAB1–TAB4 complex MAPK pathway is one of the common intersections of signal activates the IκB kinase (IKK) complex through phosphorylation. transduction pathways, such as cell proliferation, stress, inflamma- The latter phosphorylates IκB and degrades itself by ubiquitination. tion, differentiation, functional synchronization, transformation, NF-κB is released and translocated to the nucleus, thereby and apoptosis.280,281 It is an important transmitter of signals from regulating the transcription of inflammatory genes.264,265 the cell surface to the inside of the nucleus. In the signal pathway mediated by NLRs, when the bacterial In the MyD88-dependent pathway of TLRs, IRAK-1 is activated component invades the cell, NOD1 and NOD2 recognize the by phosphorylation and interacts with TRAF6. In addition to bacterial iE-DAP and MDP, respectively.266,267 And then NOD-like activating the IKK complex, it can also cause the activation of Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 9 Fig. 6 Pattern recognition receptor-mediated TBK1-IRF-3 signaling. Intracellular induction of pathogens is carried out through the detection of foreign molecular components (including cytoplasmic viral and bacterial nucleic acids). Once detected, the innate immune system induces type I interferon (IFN) production through the TANK-binding kinase 1 (TBK1)-interferon regulatory factor-3/7 (IRF-3/7) pathway. IRF-3/7 can be activated through two innate immune antiviral signal pathways, TLR3/TLR4-TIR domain-containing adaptor protein-inducing interferon β (TRIF) and RIG-I-MAVS, and then dimerize and merge into the nucleus to work MAPKs (c-Jun N-terminal kinase (JNK), p38 MAPK).282 In addition, IRF-3 phosphorylation and subsequent dimerization induce IRF- when bacterial components invade cells, NLRs are activated, 3 nuclear translocation, leading to type I IFN gene expression recruiting downstream CARD9, thereby activating p38, JNK, and (Fig. 6).192,293,294 finally activating the MAPK pathway283 to promote the release of pro-inflammatory factors. The inflammasome signaling Inflammasome is the multi-protein complex assembled by PRRs in The TBK1–IRF-3 signaling the cytoplasm and is an important part of the innate immune IRF-3 is a key transcription factor that promotes the synthesis of system.295 The inflammasome can recognize PAMPs or DAMPs type I IFN and plays an important role in the antiviral innate and recruit and activate Caspase-1. The activated Caspase-1 immune response.284 IRF-3 can be activated through two innate spliced proIL-1β/proIL-18 into the corresponding mature cyto- immune antiviral signal pathways, TLR3/TLR4-TIR domain- kine.193,296 There are five main types of inflammasomes that have containing adaptor protein-inducing interferon β (TRIF) and RIG- been discovered, namely, NLRP1 inflammasome,297 NLRP3 inflam- I-MAVS,285 and then dimerize and merge into the nucleus to work masome,298 NLRC4 inflammasome,299,300 IPAF inflammasome, and (Fig. 6).286 AIM2 inflammasome.301 Known inflammasomes generally contain The adaptor protein in the MyD88-independent pathway is TRIF. ASC, caspase protease, and a protein of the NLR family (e.g., The TRIF axis mainly induces the expression of type I IFNs.287 After NLRP3) or HIN-200 family protein (e.g., AIM2). Taking NLRP3 as an the receptor is recognized and combined with the ligand, the example,302 the dimerization of NLRP3 under the action of pathway is activated by TRIF and TRAF3, leading to the intracellular PAMPs or DAMPs makes the two PYDs to polymerize. recruitment of IKKε/TANK-binding kinase 1 (TBK1),288 phosphor- With the help of homotype interaction, NLRP3 binds and activates ylation of IRF3, and the activation of type I IFN genes, which the ASC complex with both PYD and CARD domains, which promotes the expression of IFN-α and IFN-β, and exerts antiviral reactivates the effector complex composed of CARD and caspase- effects (Fig. 6).289–291 1. In this way, NLRP3 (LRR + NACHT + PYD), ASC (PYD + CARD), RLRs such as RIG-I and MDA5 can detect viral nucleic acid. and the effector complex (CARD + Caspase-1) together constitute MDA5 and RIG-I will interact with the shared caspase recruit- the inflammasome, which produces important pro-inflammatory ment domain to induce MAVS to dimerize and bind to factors.303–305 After AIM2 recognizes cytoplasmic dsDNA, it also TRAF3.134,140,292 In turn, TRAF3 recruits the adaptor proteins uses inflammasomes to produce IL-1β and IL-18. After AIM2 TANK, NAP1, and SINTBAD. TANK connects upstream RLR signal recognizes cytoplasmic dsDNA, it also produces IL-1β and IL-18 transduction to TBK1, which induces phosphorylation of IRF-3. through the inflammasome pathway (Fig. 7).306 Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 10 Fig. 7 Pattern recognition receptor-mediated inflammasome signaling. One way for pathogenic microorganisms to induce inflammation is by activating inflammasomes, which are multi-protein complexes assembled by PRRs in the cytoplasm and activate caspase-1 and subsequent activation of pro-inflammatory cytokines IL-1β and IL-18. The inflammasome complex usually contains cytoplasmic PRRs, adaptor protein (ASC), and pro-caspase-1. Many different inflammasome complexes have been detected, each with unique PRRs and activation triggers Innate immunity not only plays a role in controlling the We can see from the data in GEPIA316 (http://gepia.cancer-pku. infection and spread of pathogens in the early stage of infection cn/) that TLR level is significantly increased in tumors, including but also plays an important role in initiating and regulating glioblastoma multiforme, brain lower-grade glioma, kidney renal adaptive immunity.12,307 Innate immune cells produce different clear cell carcinoma, acute myeloid leukemia (AML), and types of cytokines through signal transduction initiated after PRRs pancreatic adenocarcinoma (PAAD). NOD1/2 are highly expressed recognize PAMPs, which directly affect the differentiation of T in AML and PAAD, while RLRs are highly expressed in AML, PAAD, helper type 1 (Th1), Th2, Th17, and other subgroups in adaptive diffuse large B cell lymphoma, head and neck squamous cell immunity.15 For example, pathogenic microorganisms activate carcinoma, and thymoma. macrophages to secrete IL-6, TGF-β, IL-23, and other cytokines, which promote Th17 response, leading to excessive immune- Colorectal cancer (CRC). CRC, including colon cancer and rectal inflammatory effects and tissue damage, or activate NK cells to cancer, is one of the most common gastrointestinal malignancies secrete IFN-γ, and then activate macrophages to secrete IL-12, in clinical practice, and it is also one of the cancers that seriously promote the differentiation of Th0 into Th1, promote cellular endanger human health.317,318 Intestinal mucosal epithelial cells immune response, and effectively eliminate viral infections.13,308 and immune cells recognize intestinal microorganisms and their Therefore, the immune system is a system of mutual influence. products through TLRs.319–322 TLR2 can recognize peptidoglycans Any anti-infection process is completed by mutually activating or and lipopeptides that infect intestinal epithelial bacteria and inhibiting of different components. These components are as produce anti-infection and other immune-protective effects.323 small as each cytokine and as large as the immune system. Studies have shown that the expression level of TLR2 protein in colon cancer is significantly upregulated compared with normal epithelial tissues,324 and the use of TLR2 agonists significantly PRR-RELATED DISEASES enhances the proliferation, migration, and invasion capabilities of PRRs and cancers colon cancer cells.325 TLR4 is highly expressed on the surface of The inflammatory microenvironment of tumor constitutes the colon cancer cells. After stimulation and activation, it can induce a barrier for tumor growth, which is conducive to tumor formation variety of immunosuppressive factors, thus promoting the and development.309 PRRs are widely expressed in a variety of proliferation and immune escape of colon cancer cells.326 First, tumor tissues, such as colon cancer, lung cancer, breast cancer, TLR4 can produce trophic factors and vascular growth factors gastric cancer, melanoma, and so on.310,311 The activation of through the TLR4/MyD88/NF-κB signaling pathway, thereby PRRs on the surface of tumor cells can induce the expression of promoting tumor cell invasion.327,328 Second, TLR4 can promote a large number of cytokines, chemokines, hormones, and tumor proliferation through TLR4/Cyclooxygenase 2 (COX2)/ vascular-promoting factors, which is one of the important prostaglandin E2 (PGE2). PGE2 is an important cell growth and factors to induce the formation of tumor inflammatory micro- regulatory factor. After binding to specific receptors, it plays a key environment and promote the development of tumor.312,313 At role in mediating a series of cell activities, such as cell proliferation, the same time, the activation of PRRs on immune cells can differentiation, and apoptosis, and has immunosuppressive and induce antigen-presenting cells including DCs, tumor-associated anti-inflammatory effects. COX2 is the rate-limiting enzyme of macrophages, and B cells to activate tumor-specific T cell prostaglandin synthesis, and it is also highly expressed in responses or enhance the antitumor effects of phagocytes. inflammation, tumor, and other pathological states.329 Hsu These also indicate that the role of PRRs in immunotherapy et al.330 found that knocking out the mouse TLR4 gene against tumors is very important and might represent a new significantly reduced the expression of COX2 and PGE2 in the strategy for patients with tumors.314,315 intestinal mucosa; after administration of PGE2, the expression of Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 11 COX2 in the intestinal mucosa increased significantly and of HCC remains uncertain. Song et al.347 found that the deficiency promoted the occurrence of intestinal tumors. After administra- of TLR4, TLR9, and their downstream molecule MyD88 in a mouse tion of PGE2, the expression of COX2 in the intestinal mucosa model characterized by hepatic deletion of TAK1 could block the increased significantly and promoted the occurrence of intestinal liver inflammation–fibrosis–cancer axis and reduce liver injury and tumors. At the same time, it has also been found to promote the tumor growth. For TLR3, the downregulation of TLR3 in HCC expression of amphiregulin and epidermal growth factor receptor patients leads to poor prognosis (e.g., defective immune cell (EGFR) in the intestinal mucosa. Finally, the study showed that the recruitment and lack of killing of transformed hepatocytes), leading abnormal expression of TLR4 in CRC caused by chronic inflamma- to protection of transformed hepatocytes from apoptosis, thereby tion of the intestine can significantly enhance the expression of promoting the occurrence of liver cancer.348 Therefore, the PGE2, the upregulation of COX2, and the phosphorylation of EGFR expression of TLR3 may become a useful clinical treatment in intestinal mucosal cells, thereby positive feedback promotes the monitoring marker. proliferation of tumor cells. TLR5 also plays an important role in In addition to the above reasons, there is now more and more tumor immunotherapy.331 In mouse xenograft models of human evidence that the imbalance of the gut–hepatic axis may also play colon cancer, flagellin around the tumor activates TLR5 to inhibit a role in the occurrence of HCC.349 Zhou et al.350 discovered that tumor growth and promote tumor apoptosis.332 In addition, NOD2 acts as a bacterial sensor, linking gut-derived microorgan- TLR9 is expressed on the surface of the mesentery, which isms to the occurrence of HCC through a known mechanism and a maintains intestinal homeostasis and repairs intestinal damage newly discovered mechanism. The known mechanism is that by generating an immune response.333 TLR9 relies on the MyD88 NOD2 activates NF-κB, JAK2/STAT3, and MAPK pathways in a RIP2- pathway to induce downstream signals to recruit many inflam- dependent manner, leading to liver inflammation.351 It is worth matory factors, such as IL-8, TGF-β, PGE2, and other immunosup- noting that activated NOD2 can also act as the initiator of the pressive molecules,334 leading to the continuous development of nuclear autophagy pathway that does not depend on RIP2, thereby inflammation, resulting in immune escape, and promoting the promoting the degradation of the nuclear component lamin A/C, unlimited proliferation of tumor cells. After TLR9 recognizes the leading to damage to DNA damage repair mechanisms and exogenous ligand, it upregulates the expression of NF-κB signaling increased genomic instability, which eventually leads to the factor. Once this pathway is opened, it may induce the secretion occurrence of HCC.350 Meanwhile, the study found that the of matrix metalloproteinase-13 (MMP-13) and the activation of expression of ALRs was negatively correlated with tumor volume, intercellular adhesion molecule-1, thus promoting the metastasis stage, and metastasis of HCC patients. The researchers proposed of tumor cells.335,336 At the same time, the metastatic tumor cells that overexpression of ALRs in HCC cells could increase the are better adapted and combined with the cell matrix at the expression of caspase-1 and IL-1a, and the release of lactate metastasis, and the stability of tumor cell metastasis is enhanced. dehydrogenase was also observed,352 which was a marker of the Although TLRs have been shown to enhance colon cancer initiation of apoptosis. Thus, ALRs may play an antitumor role by metastasis, inhibiting these receptors cannot completely hinder promoting tumor cell apoptosis. tumor progression. Surprisingly, NOD1 is highly expressed in human CRC and its cell lines. After being activated by C12-iE-DAP, Breast cancer. Breast cancer is one of the most common it mainly enhances the adhesion, migration, and metastasis of CRC malignant tumors in the female population. It has a strong ability cells through the p38 MAPK pathway.337 to invade and metastasize.353 It can metastasize to the liver, lung, brain, bone, and other organs, forming complications and Hepatocellular carcinoma (HCC). HCC is the most common type of increasing the difficulty of treatment. Studies have shown that primary liver cancer. Among its many influencing factors, the promotion of the TLR2 signaling pathway on the metastasis inflammation is one of the main reasons that induce liver and invasion ability of human breast cancer cells is achieved by cancer.338 The expression of TLR2 in liver cancer tissues is upregulating the secretion of inflammatory cytokines.354 LTA, a significantly higher than that in normal liver tissues, and the TLR2 specific ligand, can significantly promote the secretion of expression of TLR2 protein is related to some mutant genes that tumor metastasis-related factors IL-6, TGF-β, and vascular lead to the occurrence of HCC, such as p53, PIK3CA, and endothelial growth factor (VEGF) in breast cancer cells,355 thereby β-catenin.339–342 In addition, Chew et al. revealed that the promoting the proliferation and metastatic invasion of breast expression of TLR3 has independent effects on tumor parenchyma cancer cells, and this promotion is related to the level of TLR2 and infiltrating NK cells, and the expression of these two parts is expression.356,357 In addition, the activation of TLR4 can increase related to inhibiting tumor cell proliferation, promoting tumor cell the secretion of IL-6 and IL-10 of cancer cells and induce the death, and prolonging the survival rate of patients.343 This production of more MMP-2, MMP-9, and VEGF,358,359 which can indicates that TLR3 may directly act on tumor parenchymal cells, significantly enhance the invasion ability of breast cancer. It has promote the recruitment and activation of NK cells, and exert been reported that activation of TLR4 on metastatic breast cancer antitumor effects. More and more evidences show that LPS plays a cells can regulate the expression of integrin, which can promote role in the development of HCC. Zhou et al.344 found that LPS its adhesion and invasion.360 activates the TLR4–AKT–SOX2 signaling pathway of liver cancer cell lines to improve the ability of cancer stem cells; Lin et al.345 found Head and neck squamous cell carcinoma. Squamous cell carci- that there is a positive feedback loop of COX-2/PGE2/signal noma, also known as epidermal carcinoma, is a malignant tumor transducer and activator of transcription factor 3 (STAT3) activated occurring in the epidermis or adnexal cells.361,362 It is more by LPS in liver cancer cells, which regulates the expression of genes common in the parts covered by squamous epithelium, such as related to tumor proliferation, differentiation, and apoptosis. In the skin, mouth, lip, esophagus, cervix, vagina, etc.363,364 TLR2, TLR4, latest research on the treatment of liver cancer, it is found that the and TLR9 are expressed in primary tumors, neck metastases, and antitumor effect of TLR9 agonist combined with anti-PD-1 antibody recurrent tumors of oral tongue squamous cell carcinoma or anti-PD-L1 is significantly better than single-agent therapy.346 (OTSCC), and their expression varies from the tumor surface to The activation of TLR9 inherent in liver cancer cells regulates the the invasive front, which may be one of the important factors to autoarylation and ubiquitination of poly(ADP-ribose) polymerase-1 promote the invasion of OTSCC.365,366 NOD1 and NOD2 genes are and the phosphorylation of STAT3, which together upregulate the expressed in the human oral squamous cell carcinoma (OSCC) cell expression of PD-L1 and eventually induce immune escape. line YD-10B, and they may trigger immune responses through the Although TLRs have been reported to be associated with chronic MAPK pathway. Surprisingly, the study revealed that stimulation inflammation of the liver, whether they promote the development by the NOD2 agonist MDP can inhibit cell growth by inducing Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 12 apoptosis. These findings provide the potential value of MDP as a susceptible to infection.394 This phenomenon is related to multiple new candidate for OSCC antitumor drugs.367 organ failure and immunodeficiency and is usually manifested as insufficient antibacterial activity of neutrophils.395 The neutrophil Respiratory diseases function to resist microbial infections needs to generate reactive Aspergillus fumigatus is a fungus widely distributed in nature. It can oxygen species through NADPH oxidase 2. Rolas et al. found that, easily invade the respiratory tract and cause bronchitis and in patients with alcoholic liver cirrhosis, the lack of catalytic core pneumonia in patients.368,369 For the allergic lung inflammation flavocytochrome b558 (gp91phox, p22phox) and p47phox of the caused by Aspergillus, the recognition of PAMPs by the body’s NADPH enzyme may be a new factor that patients are susceptible dendritic cells is mainly negatively regulated through the TLR2- to infection. What is surprising is that the activation of TLR7/8 can MyD88 pathway. The results showed that PAMPs recognized by reverse the expression and activity of the deficient gp91phox, TLR2 upregulated IL-10 and decreased the recruitment of which provides a direction for restoring the antibacterial response pulmonary eosinophils, thus downregulating Th2 response.370,371 of immunodeficiency patients.396,397 In allergic asthma, TLR9–IL-2 affects the Th2 response by regulating the expression of IL-17A, so small molecule inhibitors Cardiovascular diseases targeting TLR9 may become a new treatment strategy.372 Atherosclerosis is a chronic inflammatory disease caused by plaques composed of lipids, cholesterol, calcium, and other Nervous diseases substances in blood vessels.398,399 It has been reported that TLRs The connection between innate immunity and nervous system is are extensively and deeply involved in the process of athero- becoming more and more complex and close.373–375 Studies have sclerosis.400,401 TLR7 has been identified as a good prognosis shown that NOD1/NOD2 may be a new target for the treatment of marker for patients with severe atherosclerosis. TLR7 in the stress-related gut–brain diseases.376 The gut–brain axis is a plaque produced in atherosclerotic lesions will secrete IL-10 and biochemical signal of the digestive tract and central nervous system, TNF-α after being stimulated by ligand. Studies have found that which affects all events from brain development to the progression TLR7 may regulate inflammation in atherosclerosis by inhibiting of neurological diseases. The hypothalamic–pituitary–adrenal axis the effects of pro-inflammatory cytokines.402 In addition to the (HPA) is one of the main pathways of gut–brain axis signal production of plaque, cell-free DNA (cfDNA) is also released in transmission.377 It has been reported that the immune system plays atherosclerotic lesions. TLR9 recognizes cfDNA and plays a key a key role in brain function and stress response. NLRs are PRRs role in the development of vascular inflammation and athero- expressed in the gut and brain. The lack of NOD1 and NOD2 affects sclerosis by promoting the pro-inflammatory activation of the serotonergic signaling of gut and brain, the proliferation of macrophages.403 The formation of initial atherosclerotic plaques hippocampal cells, and the maturation of neurons, which makes the is caused by the interaction of macrophages and endothelial mice lacking both of them vulnerable to HPA overactivation under cells, macrophage infiltration, and other factors that lead to an stimulation, thus showing anxiety, cognitive impairment, and increase in neointima.404,405 It is worth noting that studies have depression.378–380 TLR4 is very important in the process of shown that TLR5-mediated activation of NADPH oxidase 4 neuropathic pain caused by infection and sterile neuronal injury. (Nox4) can regulate the migration of SMCs and promote the TLR4 exerts its effects through the activation and nuclear localization expression of pro-inflammatory cytokines, which may contribute of NF-κB and the production of pro-inflammatory cytokines, which to the formation of atherosclerotic plaques. This indicates that can activate pain receptors to cause neuropathic pain.381,382 Relevant the flagellin–TLR5–Nox4 cascade is of great significance in studies have revealed for the first time that lysozyme acts as an atherosclerotic intimal hyperplasia.406,407 In addition, TLR2 endogenous ligand for activating TLR4 in sterile nerve injury, thereby activates p38 and extracellular signal-regulated kinase 1/ promoting neuronal excitement and neuropathic pain.383 The 2 signals, thereby upregulating IL-6-mediated receptor activator identification of lysozyme as DAMPs has improved our under- of NF-κB ligand and downregulating osteoprotegerin. These will standing of neuroinflammation and opened up prospects for the make the cartilage formation of vascular SMC transdifferentiate, treatment of neuropathic pain. leading to vascular calcification.408 The expression of TLR3 is reduced in the lung tissue and endothelial cells of patients with Digestive diseases pulmonary hypertension, and its deficiency increases the Newborns with chronic obstructive jaundice make their livers susceptibility to apoptosis and pulmonary hypertension.409 In prone to cholestatic liver disease, and biliary atresia (BA) accounts addition to TLRs, the lack of NOD1 and NOD2 can lead to lipid for half of the cases.384 Viruses have always been considered as deposition of atherosclerotic plaques and the reduction of the causative pathogen of this disease, and the role of TLRs in the inflammatory cell infiltration, so it has been identified as pre- pathogenesis and progression of BA has been determined.385 disease factors.410 Subsequent studies have shown that activation of TLR7 can Human brain pericytes (HBPs) are an important component of induce type 1 IFN signal transduction, apoptosis, and dysplasia of the microvascular wall and contribute to the integrity of the the neonatal liver and biliary system. This new discovery reveals blood–brain barrier (BBB).411 It has been found that TLR4 and the pathogenesis of neonatal cholestatic liver disease.386 NOD1 are expressed in HBP, which also reveals that HBP has the The expression of TLR5 is closely related to various infectious ability to sense systemic infection or blood-borne PAMPs. HBP can diseases caused by bacteria.387,388 The lack of TLR5 can cause perceive Gram-negative bacteria to protect BBB through different changes in the intestinal flora and cause colitis.389 The pathways mediated by TLR4 and NOD1, or it can trigger paracrine protein–protein interaction between TLR5 and flagellin plays an signaling pathways by releasing chemokines and cytokines, important role in pathogen defense, immune diseases, and leading to the destruction of BBB.412,413 These are all new insights tumors. 4-((4-benzyl-5-(pyridin4yl)-4H-1,2,4-triazol-3-yl)thio)pyrido that PRRs are involved in the body’s inflammatory response and [3’,2’:4,5]thieno[3,2-d] Pyrimidine (TH1020) is a small molecule may have an impact on the treatment of diseases. inhibitor identified through high-throughput screening, which can disrupt the association between TLR5 and flagellin, and provides a Endocrine diseases lead compound for new therapies against TLR5.390,391 In order to adapt to the constantly changing internal and external Alcoholic liver disease is a liver disease caused by long-term environment and maintain the relative stability of the internal heavy drinking. The initial stage usually manifests as fatty liver, environment, the human body must rely on the cooperation and which can then develop into alcoholic hepatitis, liver fibrosis, and regulation of the nervous, endocrine, and immune systems,414,415 cirrhosis.392,393 Patients with advanced alcoholic cirrhosis are more so that the activities of various organs and systems are Signal Transduction and Targeted Therapy (2021)6:291  Pattern recognition receptors in health and diseases Li and Wu 13 coordinated, and jointly shoulder all the life phenomena of the most of the agonists and antagonists of TLRs are only in the body. The immune system is not only regulated by the other two clinical development stage, which are studied comprehensively in systems but also affects them through chemical information PRRs (Table 2). In addition, microRNA (miRNA), exosomes, and molecules and receptors.416,417 combination therapy also have certain potential in this field.438,439 In patients with autoimmune thyroid disease, the expression and activation of TLR2, 3, and 9 are significantly increased.418,419 In PRR agonists for therapy addition, it has been reported that TLR9 negatively regulates Imiquimod is the first targeted drug for TLRs. It is a specific agonist pancreatic islet development and β cell differentiation, providing of the TLR7 receptor. It can induce the production of IFN-α, IL-6, new directions for diabetes prevention and treatment strate- and TNF-α to achieve the purpose of regulating immunity and gies.420 In diet-induced obesity, TLR2 and TLR4 inhibit the treating tumors.440 Selgantolimod, a novel TLR8 agonist, can replication of β cells and affect the nuclear abundance of the activate TLR8 and elicit cytokine responses in patients with cell cycle regulators cyclin D2 and Cdk4. Therefore, targeting chronic hepatitis B infection.441,442 Further studies with longer TLR2–TLR4 may alleviate the failure of β cells in diabetic durations of selgantolimod treatment are required to evaluate patients.421 efficacy. The ligand of TLR9 is unmethylated DNA, and CpG One of the most common complications of diabetes is diabetic oligodinucleotide (CpG ODN) that mimics this structure has good foot ulcers, and the reason why it is not easy to get better is the antitumor potential (e.g., IMO-2055, MGN-1703, MGN-1704).443 inflammation of the wound.422,423 Singh et al. believe that The research of MGN-1703 on the treatment of advanced CRC is in changes in the expression level of intracellular TLRs may be one phase III. MGN-1703 can induce a potent type I IFN response in the of the reasons for the continuous inflammation of chronic intestine, which is related to the subtle changes of intestinal wounds, such as diabetic foot ulcers, and may hinder wound flora.444 Clinically, PRR agonists can be potentially used not only as healing in patients with type 2 diabetes mellitus (T2DM).424 The therapeutic agents to treat but also as adjuvants in conjunction basis of these pro-inflammatory effects is that intracellular TLRs with other immunotherapies.445,446 Due to the instability and drug activate dendritic cells and B cells to produce IFN I and III, resistance of existing drugs, some adjuvants targeting PRRs aggravating the inflammatory response.425–427 The inflammatory emerge as the times require. A TLR3-specific adjuvant, ARNAX, phase of the wound healing cascade is the decisive stage of can enhance DC priming and CTL proliferation without cytokine wound development, and studies have found that certain toxicity.447 And the conjugated STING and TLR1/2 agonist members of innate immunity play an important role in the Pam3CSK4-CDGSF can be used as an effective adjuvant for pathogenesis of chronic wound healing abnormalities.428,429 The constructing vaccines to enhance antitumor immunotherapy.448 signaling pathway of intracellular TLRs is shown to be involved in NLR agonists can promote the processing and secretion of IL-1β, some chronic inflammatory diseases, such as systemic lupus which is very important for activating many immune cells.449,450 erythematosus (SLE), multiple sclerosis, hepatitis, and T2DM.430 Therefore, NLR agonists can be served as effective components of vaccines and immune stimulants. Skeletal diseases The class II major histocompatibility complex transactivator The expression level of NOD2 in human osteoarthritis (OA) (CIITA) is a single member of the NLRA subfamily.451 As the main cartilage is significantly higher than that of normal cartilage, and regulator of major histocompatibility complex II, its own its combined action with TLR2 contributes to the pro-catabolic modification is very important in the occurrence and development gene expression induced by 29-kDa amino terminal (matrix of tumors. Both the deacetylation of histones and the demethyla- degradation product in synovial fluid of patients with OA) in tion of DNA will downregulate the transcription of CIITA.452 human chondrocytes.431,432 This indicates that the NOD2 and TLR2 Decitabine (5-aza-2’-deoxycytidine), Entinostat (MS-275), and cross-regulatory pathway may be a target to prevent the Trichostatin A target CIITA to promote its expression recovery in development of arthritis. More and more evidences indicate that tumors. In addition, andrographolide, arglabin, formononetin, the interaction between different signal pathways may be a new mangiferin, and other botanicals can be used to treat tumors by way for the occurrence and development of diseases. The latest inhibiting NLRP3 inflammasome.453–457 Representative NLR ago- research combines three biological processes in skeletal muscle nists like Mifamurtide have been approved for clinical trials of non- during exercise, innate immune response, autophagy protein metastatic sarcoma. homeostasis, and adenosine