Elsevier

Cytokine

Volume 42, Issue 2, May 2008, Pages 145-151
Cytokine

Review Article
LPS/TLR4 signal transduction pathway

https://doi.org/10.1016/j.cyto.2008.01.006Get rights and content

Abstract

The stimulation of Toll-like receptor 4 (TLR4) by lipopolysaccharide (LPS) induces the release of critical proinflammatory cytokines that are necessary to activate potent immune responses. LPS/TLR4 signaling has been intensively studied in the past few years. Here we review molecules involved in TLR4-mediated signaling, including players that are involved in the negative regulation of this important pathway.

Introduction

The Toll protein first discovered in Drosophila, was shown to be essential for determining the dorsal–ventral patterning during embryogenesis [1], [2] and an early form of the innate immune system [3], [4]. The mammalian Toll-like receptors (TLRs) are germline-encoded receptors expressed by cells of the innate immune system that are stimulated by structural motifs characteristically expressed by bacteria, viruses and fungi known as pathogen-associated molecular patterns (PAMPs) [5], [6]. Importantly, TLR interactions trigger the expression of proinflammatory cytokines as well as the functional maturation of antigen presenting cells of the innate immune system [6], [7].

Many PAMPs have been defined that interact with particular TLRs. For example, the TLR2/TLR6 heterodimer can be stimulated by several bacterial components, such as lipoteichoic acid (LTA) and peptidoglycan (PG). Viral DNA is rich in unmethylated CpG motifs, which stimulates TLR9. While TLR3 interacts with viral double-stranded RNA, TLR7/8 can sense guanosine- or uridine-rich single-stranded RNA from viruses. Collectively, the innate immune system utilizes TLRs and cytosolic sensors (RIG-I, MDA5, ets.) to detect viruses [8]. Thus, using TLR as critical sensors, the innate immune system has devised a way to decode the type of invading pathogen and trigger an appropriate effective immune response.

Evidence suggests that several PAMPs can stimulate TLR4. These molecules include lipopolysaccharide (LPS) from Gram-negative bacteria, fusion (F) protein from respiratory syncytial virus (RSV) and the envelope protein from mouse mammary tumor virus (MMTV) [9], [10]. In addition, endogenous molecules can also interact directly or indirectly with TLR4, such as heat-shock proteins, hyaluronic acid and β-defensin 2 [11], [12], [13].

LPS is one of the best studied immunostimulatory components of bacteria and can induce systemic inflammation and sepsis if excessive signals occur [14]. LPS is an important structural component of the outer membrane of Gram-negative bacteria. LPS consists of three parts: lipid A, a core oligosaccharide, and an O side chain [15], [16]. Lipid A is the main PAMP of LPS. Using the C3H/HeJ mouse strain which is known to have a defective response to LPS, Beutler’s group demonstrated that TLR4 is an important sensor for LPS [17].

LPS stimulation of mammalian cells occurs through a series of interactions with several proteins including the LPS binding protein (LBP), CD14, MD-2 and TLR4 [18], [19]. LBP is a soluble shuttle protein which directly binds to LPS and facilitates the association between LPS and CD14 [20], [21]. CD14 is a glycosylphosphatidylinositol-anchored protein, which also exists in a soluble form. CD14 facilitates the transfer of LPS to the TLR4/MD-2 receptor complex and modulates LPS recognition [22]. MD-2 is a soluble protein that non-covalently associates with TLR4 but can directly form a complex with LPS in the absence TLR4 [23], [24], [25]. Although no evidence suggests that TLR4 can bind LPS directly, TLR4 can enhance the binding of LPS to MD-2 [26]. Therefore LPS stimulation of TLR4, includes the participation of several molecules, and the currently favoured model is outlined in Fig. 1 [19], [27].

Upon LPS recognition, TLR4 undergoes oligomerization and recruits its downstream adaptors through interactions with the TIR (Toll-interleukin-1 receptor) domains. TIR domains contain three highly conserved regions, which mediate protein–protein interactions between the TLRs and signal transduction adaptor proteins. The TIR domain of TLR4 is critical for signal transduction, because a single point mutation in the TIR domain can abolish the response to LPS [17]. There are five TIR domain-containing adaptor proteins: MyD88 (myeloid differentiation primary response gene 88), TIRAP (TIR domain-containing adaptor protein, also known as Mal, MyD88-adapter-like), TRIF (TIR domain-containing adaptor inducing IFN-β), TRAM (TRIF-related adaptor molecule), and SARM (sterile α and HEAT-Armadillo motifs-containing protein) [28]. Different TLRs use different combinations of adaptor proteins to determine downstream signaling. Interestingly, TLR4 is the only known TLR which utilizes all these adaptor proteins.

Studies using knockout mice have revealed important roles for these adaptors in TLR4 signaling. MyD88 was first described as a myeloid differentiation primary response gene [29]. It was later suggested to be the critical adaptor in the interleukin-1 receptor (IL-1R) signaling pathway [30], [31]. Because both the IL-1R family and the TLR family contained TIR domains, studies were also done to determine whether MyD88 was involved in TLR-mediated signaling pathways. MyD88-deficient mice were shown to be resistant to LPS-induced septic shock, and MyD88-deficient macrophages failed to produce proinflammatory cytokines after LPS stimulation, despite the ability to activate nuclear factor-κB (NF-κB) [32]. In addition, the expression of Type I interferons and interferon-inducible genes was not impaired in MyD88-deficient macrophages [33]. This demonstrated an important role for MyD88 downstream of IL-1R and TLR signaling, but also indicated that other molecules are involved in the induction of a subset of LPS induced responses.

TIRAP/Mal was cloned through a computer-based search for proteins containing TIR domains [34], [35]. TIRAP-deficient mice were generated in subsequent studies and had a phenotype similar to MyD88 knockout mice [36], [37]. TIRAP also contains a phosphatidylinositol 4,5-bisphosphate (PIP2) binding domain, which mediates TIRAP recruitment to the plasma membrane. TIRAP then facilitates the association between MyD88 and the TLR4 cytoplasmic domain to initiate MyD88-dependent downstream signaling [38].

TRAM was also cloned by homology of the TIR domain [39], while TRIF was cloned using multiple approaches [40], [41], [42]. Studies using knockout mice indicated that TRIF and TRAM mediate MyD88-independent signaling and will be discussed in detail later [39], [42], [43]. Studies suggest that TRAM associates with the plasma membrane through myristoylation, and is essential for TLR4 signal transduction [44] (Fig. 1). SARM was suggested to function as an inhibitor of TRIF-mediated signaling in the human HEK293 cell line [45]. However, the role of SARM in vivo is still unclear.

Section snippets

TLR4 signal transduction

TLR4 signaling has been divided into MyD88-dependent and MyD88-independent (TRIF-dependent) pathways. Based on studies using MyD88-deficient macrophages, the MyD88-dependent pathway was shown to be responsible for proinflammatory cytokine expression, while the MyD88-independent pathway mediates the induction of Type I interferons and interferon-inducible genes (Fig. 1).

Negative regulation of TLR4 signaling pathway

Because TLR4 stimulation can induce potent responses such as sepsis, inhibitory pathways are necessary to protect the host from inflammation-induced damage. TLR4 signaling can be regulated at multiple levels by many negative regulators. Typically mice lacking these key regulators exhibit enhanced TLR4 responses [74]. RP105 (radioprotective 105), ST2L (also known as IL1Rl) and SIGIRR (single immunoglobulin IL-1R-related molecule) are expressed on the cell surface and their inhibitory functions

Conclusion

Recent studies have provided tremendous insights into LPS/TLR4 signaling pathway. The knowledge from this pathway also provides models for how other TLR signaling pathways may be regulated. Because improper regulation of LPS/TLR4 signaling has the potential to induce massive inflammation and cause acute sepsis or chronic inflammatory disorders, it is important to further explore this pathway and evaluate novel targets to counteract these conditions [90], [91].

References (91)

  • M. Koziczak-Holbro et al.

    IRAK-4 kinase activity is required for interleukin-1 (IL-1) receptor- and toll-like receptor 7-mediated signaling and gene expression

    J Biol Chem

    (2007)
  • E. Lye et al.

    The role of interleukin 1 receptor-associated kinase-4 (IRAK-4) kinase activity in IRAK-4-mediated signaling

    J Biol Chem

    (2004)
  • S.E. Keating et al.

    IRAK-2 participates in multiple toll-like receptor signaling pathways to NFκB via activation of TRAF6 ubiquitination

    J Biol Chem

    (2007)
  • N. Cusson-Hermance et al.

    Rip1 mediates the Trif-dependent Toll-like receptor 3- and 4-induced NF-κB activation but does not contribute to interferon regulatory factor 3 activation

    J Biol Chem

    (2005)
  • B. Guo et al.

    Modulation of the interferon antiviral response by the TBK1/IKKi adaptor protein TANK

    J Biol Chem

    (2007)
  • P.N. Moynagh

    TLR signalling and activation of IRFs: revisiting old friends from the NF-kappaB pathway

    Trends Immunol

    (2005)
  • A.G. Bowie et al.

    The role of Toll-like receptors in the host response to viruses

    Mol Immunol

    (2005)
  • J. Qin et al.

    SIGIRR inhibits interleukin-1 receptor- and toll-like receptor 4-mediated signaling through different mechanisms

    J Biol Chem

    (2005)
  • C. Fearns et al.

    Triad3A regulates ubiquitination and proteasomal degradation of RIP1 following disruption of Hsp90 binding

    J Biol Chem

    (2006)
  • H. Wesche et al.

    IRAK-M is a novel member of the Pelle/interleukin-1 receptor-associated kinase (IRAK) family

    J Biol Chem

    (1999)
  • K. Kobayashi et al.

    IRAK-M is a negative regulator of Toll-like receptor signaling

    Cell

    (2002)
  • M.P. Hardy et al.

    The murine IRAK2 gene encodes four alternatively spliced isoforms, two of which are inhibitory

    J Biol Chem

    (2004)
  • D. Morisato et al.

    Signaling pathways that establish the dorsal–ventral pattern of the Drosophila embryo

    Annu Rev Genet

    (1995)
  • S. Cherry et al.

    Host-pathogen interactions in Drosophila: new tricks from an old friend

    Nat Immunol

    (2006)
  • C.A. Janeway et al.

    Innate immune recognition

    Annu Rev Immunol

    (2002)
  • E.A. Kurt-Jones et al.

    Pattern recognition receptors TLR4 and CD14 mediate response to respiratory syncytial virus

    Nat Immunol

    (2000)
  • J.C. Rassa et al.

    Murine retroviruses activate B cells via interaction with toll-like receptor 4

    Proc Natl Acad Sci USA

    (2002)
  • K. Ohashi et al.

    Cutting edge: heat shock protein 60 is a putative endogenous ligand of the toll-like receptor-4 complex

    J Immunol

    (2000)
  • C. Termeer et al.

    Oligosaccharides of Hyaluronan activate dendritic cells via toll-like receptor 4

    J Exp Med

    (2002)
  • A. Biragyn et al.

    Toll-like receptor 4-dependent activation of dendritic cells by beta-defensin 2

    Science

    (2002)
  • B. Beutler et al.

    Innate immune sensing and its roots: the story of endotoxin

    Nat Rev Immunol

    (2003)
  • C.R. Raetz et al.

    Lipopolysaccharide endotoxins

    Annu Rev Biochem

    (2002)
  • S.I. Miller et al.

    LPS, TLR4 and infectious disease diversity

    Nat Rev Microbiol

    (2005)
  • A. Poltorak et al.

    Defective LPS signaling in C3H/HeJ and C57BL/10ScCr mice: mutations in Tlr4 gene

    Science

    (1998)
  • T.L. Gioannini et al.

    Regulation of interactions of Gram-negative bacterial endotoxins with mammalian cells

    Immunol Res

    (2007)
  • P.S. Tobias et al.

    Isolation of a lipopolysaccharide-binding acute phase reactant from rabbit serum

    J Exp Med

    (1986)
  • S.D. Wright et al.

    Lipopolysaccharide (LPS) binding protein opsonizes LPS-bearing particles for recognition by a novel receptor on macrophages

    J Exp Med

    (1989)
  • S.D. Wright et al.

    CD14, a receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein

    Science

    (1990)
  • R. Shimazu et al.

    MD-2, a molecule that confers lipopolysaccharide responsiveness on Toll-like receptor 4

    J Exp Med

    (1999)
  • Y. Nagai et al.

    Essential role of MD-2 in LPS responsiveness and TLR4 distribution

    Nat Immunol

    (2002)
  • T.L. Gioannini et al.

    Isolation of an endotoxin-MD-2 complex that produces Toll-like receptor 4-dependent cell activation at picomolar concentrations

    Proc Natl Acad Sci USA

    (2004)
  • H. Mitsuzawa et al.

    Recombinant soluble forms of extracellular TLR4 domain and MD-2 inhibit lipopolysaccharide binding on cell surface and dampen lipopolysaccharide-induced pulmonary inflammation in mice

    J Immunol

    (2006)
  • L.A. O’Neill et al.

    The family of five: TIR-domain-containing adaptors in Toll-like receptor signalling

    Nat Rev Immunol

    (2007)
  • K.A. Lord et al.

    Nucleotide sequence and expression of a cDNA encoding MyD88, a novel myeloid differentiation primary response gene induced by IL6

    Oncogene

    (1990)
  • M. Muzio et al.

    IRAK (Pelle) family member IRAK-2 and MyD88 as proximal mediators of IL-1 signaling

    Science

    (1997)
  • Cited by (0)

    View full text