How Toll-like receptors signal: what we know and what we don’t know

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Signal transduction pathways activated by Toll-like receptors (TLRs) have continued to be a major focus of research for investigators interested in the initiation of innate immune responses and the induction of pro-inflammatory cytokines and type I interferons during infection. Biochemical details of the major signalling pathways have now been obtained, and the specific signalling pathways activated by different TLRs are being elucidated. New insights into the activation of IRF family members, notably IRF3, IRF5 and IRF7, have been obtained, and interesting spatiotemporal aspects of signalling by MyD88 leading to IRF7 activation revealed. Adapters in TLR signalling are targets for inhibition, both by endogenous regulators and by virally derived proteins. Selective targeting of pathways by anti-inflammatory glucocorticoids also indicates the potential for modulating TLR signalling therapeutically.

Introduction

One of the earliest indications that the body has been infected with an invading microbe is the activation of signalling pathways by Toll-like receptors (TLRs; [1]). TLRs have emerged as the key sensors of microbial products, as they are expressed on sentinel cells in the immune system, most notably dendritic cells and macrophages, where they sense a range of chemicals produced by bacteria, viruses, fungi and protozoa. TLRs can be grouped into families according to the types of ligands they recognize. Lipid-based structures are recognized by TLR2 (in combination with TLR1 or TLR6 as heterodimers) and TLR4 (as a homodimer): the most studied examples of lipid-based recognition are bacterial or mycobacterial lipopeptides, or glycerophosphatidylinositol anchors from parasites, both of which are recognized by TLR2, and bacterial lipopolysaccharide (LPS), which is recognized by TLR4. Viral and/or bacterial nucleic acids are recognized by TLR3, TLR7, TLR8 and TLR9; the most characterised are the recognition of double stranded RNA (dsRNA) by TLR3 and recognition of CpG motifs in DNA by TLR9. Finally, TLR5 and TLR11 recognize proteins from pathogens (flagellin in the case of TLR5 and profilin in the case of TLR11 [only in the mouse]) [2].

The events initiated inside cells when a given TLR is activated have been the subject of much investigation. Many biochemical details of these events have now been elucidated, including the description in the past four years of novel adaptor proteins, protein kinases and transcription factors. Several issues, however, still remain and, in the coming years, we can anticipate new insights into how TLRs work at the molecular level, and the importance of the signals activated for infectious and inflammatory diseases.

Section snippets

Current understanding of the main features of TLR signalling

Broadly speaking, two major pathways are activated by TLRs [3]. The first of these culminates in the activation of the transcription factor NF-κB, which acts as a master switch for inflammation, regulating the transcription of many genes that encode proteins involved in immunity and inflammation. The second leads to activation of the MAP kinases p38 and Jun amino-terminal kinase (JNK), which also participate in increased transcription and regulate the stability of mRNAs that contain AU repeats.

Do we have a molecular understanding of TLR signalling?

A true molecular understanding of how TLRs signal will only be evident when we know the molecular structures of the interfaces between proteins in signalling complexes and of the conformational changes that occur during signalling; however, this is a major technical challenge. The molecular structures of the TIR domains of TLR1, TLR2 and IL-1 receptor accessory protein-like (IL-1RAPL; a member of the IL-1 receptor sub-branch of the TIR domain family) have been solved [17, 18] and, broadly

How do TIR adapters lead to specific signals?

As mentioned above, the engagement of Trif with TBK-1 enables the TLRs that signal via Trif (TLR3 and TLR4) to induce IFN-β. Other specific effects for adapters have been reported: Mal and MyD88 are required for the early activation of NF-κB, whereas Trif and Tram are required for later activation [8, 12, 16, 26]. Two recent studies [27•, 28•] now identify the source of this later activation of NF-κB. It appears that activation of Trif by LPS leads to the rapid production of TNF, which is

Adapters as Achilles’ heels

In addition to forming the basis for specificity in the signalling processes activated by TLRs, adapters also appear to be targets for both exogenous and endogenous inhibition of TLR signalling. Some of these effects are shown in Figure 2. An early demonstration of this was the observation that ST2, which is another member of the IL-1R subgroup of TIR proteins, inhibits TLR signalling in macrophages by sequestering MyD88 and Mal from signalling pathways [34]. A splice variant of MyD88, termed

New transcription factors and new information on old ones

The prolonged focus on NF-κB as a target for TLR signalling was in part due to the large number of genes whose promoters contain NF-κB sites and in part due to the ease of assaying NF-κB. More recently, attention has turned to the IRF family of transcription factors, which are structurally related to NF-κB family members and which have important roles in the regulation of type I interferon production and a growing list of other genes.

Conclusions

The continued focus of research on TLR signalling has provided a lot more information on pathways activated by TLRs, and new layers of complexity and regulation have been revealed. Where this will all lead is still not clear, other than to the identification of an increasingly bewildering array of proteins and interactions leading to highly complex modulation in the expression of large sets of genes that will dynamically alter depending on strength and duration of signal. All of this

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

Acknowledgements

The O’Neill laboratory is supported by grants from Science Foundation Ireland and the Health Research Board.

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