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Structural Characterization of Nucleic Acid-sensing Toll-like Receptors

8 The innate immune system senses pathogen-associated or cell damageassociated structurally conserved molecules through various pattern recognition receptors (PPRs) [1]. Nucleic acids that are released from viruses, bacteria or dead cells during infection or tissue damage are principal ligands to PPRs. PRR involved in nucleic acid recognition can be divided into two groups on the basis of cellular localization: several members of membrane-bound Tolllike receptors (TLRs) and cytoplasmic PPRs such as Nod-like receptors (NLRs), RIG-I like receptors (RLRs), and the DNA recognition receptor cyclic GMP-AMP synthase (cGAS). TLRs, evolutionarily conserved membrane-spanning recep tors homologous to the Drosophila Toll protein, are mostly expressed in macrophages and dendritic cells. The recognition of pathogen-associated molecular patterns (PAMPs) or damage-associated molecular patterns (DAMPs) by TLRs is critical for the activation of the transcription factor NF-κB or IRFs, leading to the production of pro-inflammatory cytokines or type I interferons (Fig. 1). TLRs are glycosylated type I integral membrane receptors with N-terminal extracellular leucine-rich repeats (LRRs), a transmembrane domain, and a C-terminal cytoplasmic domain (Fig. 2(a)). The LRR domain contains binding sites for PAMPs or DAMPs, while the cytoplasmic domain, known as the Toll/IL-1 receptor (TIR) region, activates downstream signaling cascades by interacting with adaptor proteins such as MyD88 and TRIF [2]. Biochemical and structural studies on TLR extracellular domains suggest the following mechanism of TLR activation. Inactive TLRs exist as monomers, and l igand binding induces dimerization of the extracellular TLR domain, producing a typical “m”-shaped structure, where the intracellular C-terminal regions of the two TLR protomers are positioned in close proximity. Subsequent dimerization of the intracellular TIR domain is followed by the recruitment of adaptor proteins that execute signal transduction. Viral/bacterial nucleic acids are potent stimulators of innate immunity. In humans, nearly half the TLRs recognize nucleic acid ligands. TLR3 responds to double-stranded (ds) RNA, while TLR7 and TLR8, which are closely related, recognize singlestranded (ss)RNA, and TLR9 senses DNA with unmethylated cytosine phosphate-guanosine (CpG) motifs (Fig. 2(b)). In addition, mouse TLR13 detects bacterial 23S ribosomal RNA. In contrast to other TLRs, TLR3 and TLRs 7–9 are expressed on endosomal membranes, limiting the recognition of self-derived ligands that are released from dying cells (Fig. 1). In addition, the members of the TLR7 subfamily comprising TLRs 7, 8, and 9 contain a characteristic long inserted loop region (known as the Z-loop) composed of approximately 30 amino acid residues, and localized in the region between LRR14 and LRR15 (Fig. 2(a,b)). Z-loop processing is required for the activation of these TLRs. Structural Characterization of Nucleic Acid-sensing Toll-like Receptors Review Article