null

Toll-like Receptor Signalling in Neurodegenerative Disease

Toll-Like Receptors

The innate immune response has come under the spotlight in recent years due to its central role in propagating the pathogenesis of several diseases, and specifically in driving neurodegenerative aetiology (1-3). Toll-like receptors (TLRs), the mammalian homologue of the Drosophila melanogaster Toll, are highly conserved innate immune receptors and master regulators of the cellular innate immune response (4-6). Research by several groups in the early 1990s discovered the pivotal role of TLRs in the initiation and propagation of the inflammatory signalling in response to bacterial, viral or microbial nucleic acids, known as Pathogen Associated Molecular Patterns (PAMPs) or Danger Associated Molecular Pattern (DAMPs) (5, 7, 8), primarily focusing on what is now termed TLR4. To date, 13 murine and 11 human TLRs identified (9, 10), with several intracellularly expressed on endosomes (TLR3, TLR7, TLR8 and TLR9), whereas others are characterized as transmembrane receptors (all other TLRs, including TLR2 and TLR4).

Glial Cell Activation

It is widely established that reactive gliosis, a term used to describe the activation of glial cells in response to pathogens (11), is concurrent with neuronal death in the Central Nervous System (CNS) (12-14), with microglial cells eliciting the primary response to pathogenic insult (15, 16) and driving chronic neuroinflammation (17-19). Importantly, microglia highly express TLRs (20, 21). TLR signalling, specifically increased levels of TLR2 and TLR4, is reported in multiple neurodegenerative pathologies (22, 23), with mounting evidence for misfolded protein aggregate induced activation of TLR2 and TLR4 in Alzheimer’s disease (24, 25), Amyotrophic Lateral Sclerosis (2), and Prion disease (26). In the CNS, glial cell crosstalk potentiates a TLR pro-inflammatory feedback loop, with evidence for TLR signaling in initiating the adaptive immune response (27). These studies highlight the importance of TLR-induced inflammation in the chronic neuroinflammatory cascade seen in neurodegenerative aetiology.

TLR signalling

All TLRs signal as dimers, with stimulation of TLR2 and TLR4 leading to the recruitment of extracellular co-receptors CD14 (28) and MD2 (TLR4 only) (29), preceding homodimerization (TLR4/TLR4), or heterodimerization (TLR1/TLR2) (30) and subsequent assembly of a cytoplasmic Toll/Interleukin-1 Receptor (TIR) adaptor domain (31-33). The TIR-adaptor domain is comprised of MyD88 (Myeloid Differentiation Primary Response Gene 88), Mal (MyD88 adaptor-like), TRIF (TIR-domain-containing adapter-inducing interferon-β) and TRAM (34-36). MyD88 facilitates downstream signalling in all TLRs, with the exception of TLR3 (37). Mal (also known as TIRAP) bridges MyD88 to TLR2 and TLR4 (38), with TRAM adapting signals from TLR4 to TRIF (39).

TLR2 and TLR4 activation

Activation of both TLR2 and TLR4 leads to MyD88-dependent signal propagation, whereas TLR4 can additionally initiate a MyD88-independent, also termed TRIF-dependent, signalling pathway, ultimately culminating in the activation of transcription factors, such as NF-κB, Mitogen Activating Protein Kinase (MAPK) and Interferon Regulatory Factor 3 (IRF3), promoting inflammatory gene transcription [reviewed in (21)]. TLR signalling is mediated by a multifaceted network of adaptor proteins, kinases and E3 ubiquitin ligases (40). Ubiquitination is a post-translational modification executed by the formation of polyubiquitin chains on target proteins at different lysine residues modifying the function or stability of the target protein (41-44), and is tightly regulated by a number of E3 ubiquitin ligases that facilitate the polyubiquitination of several critical components inducing TLR-induced pro-inflammatory signalling (42, 45, 46).

MyD88-dependent signalling

MyD88-dependent signalling results in the activation of the Interleukin-1 Receptor Associated Kinase (IRAK) complex at the membrane. IRAK1, a member of the multi-domain protein kinase IRAK complex containing IRAK4, is phosphorylated upon activation which facilitates the ubiquitin- and phosphorylation- dependent activation of the IκB (IKK) kinase complex, leading to NF-κB transcription (47). Both the kinase activity and adaptor functions of IRAK1 facilitate IRAK4-MyD88 interactions, and are essential for TLR-mediated NF-κB activation (48). The E3 ubiquitin ligase TRAF6 induces ubiquitination of the IRAK complex activating NF-κB (41, 49). Additionally, TRAF6 can initiate NF-κB activation by facilitating the recruitment of the TAK1 (Transforming growth factor beta-activated kinase 1) – TAB1 (TGF-Beta Activated Kinase 2) – TAB2 (TGF-Beta Activated Kinase 2) complex (42) and can directly form ubiquitin chains with the IKK subunit NEMO (also referred to as IKKγ) (50).

The association of Pellino proteins and IRAK1, specifically Peli1-induced K63-linked polyubiquitination of IRAK1, facilitates IRAK activation and mediates downstream signalling to NF-κB and MAPK (43, 51). TLR4-induced dissociation and cytoplasmic translocation of the TRAF6-Peli1-IRAK complex from the membrane bound MyD88-receptor complex is essential for MAPK activation, and is regulated by TRAF3 degradation (52). Peli1-mediated K63-linked polyubiquitination of Cellular Inhibitor of Apoptosis (cIAP2) induces the degradation of TRAF3 via cIAP2-mediated K48-linked polyubiquitination of TRAF3 (53).

MyD88-indepednent signalling

The presence of a MyD88-independent signalling pathway was first reported using MyD88-/- mice (36) stimulated with Lipopolysaccharide (LPS), a well-defined potent agonist of TLR4 (54, 55), and is mediated through engagement of the TIR adaptor domain protein TRIF (31). TRIF interacts with Receptor interacting Protein 1 (RIP1) and Tumour Necrosis Factor α receptor associated factor 6 (TRAF6), to activate NF-ҡB upon and TLR4 stimulation (56, 57). Both TRAF6 and RIP1 facilitate the polyubiquitination of NEMO eliciting downstream phosphorylation and activation of the IKK complex (58).

Additionally, TRIF initiates TRAF3-dependent signalling to TANK-binding kinase 1 (TBK1) and the IKK subunit IKKε, inducing activation of the IRF3 transcription factor (59). Interestingly, TRAF3 has a differential role in the regulation of the MyD88 and TRIF-dependent pathway dictated by the alternative ubiquitination of TRAF3 (52), negatively regulating MyD88 signalling (53), and positively regulating the TRIF-dependent pathway to IRF3 (52). Peli1 has a critical role in the modulation of TRIF-dependent signalling, with Peli1-deficient mice shown to be resistant to TLR4 induced septic shock (60). Furthermore, it was demonstrated that Peli1 interacts with and promotes the phosphorylation of TBK1 resulting in bidirectional communication and enhanced activation of Peli1 (61, 62).

Targeting Neurodegeneration

Targeting the immune system by developing therapeutics for the amelioration of neurodegeneration has been a central focus in recent years; however it has proven difficult due to the ablation of the protective effects associated with the pro-inflammatory response. Recent studies have highlighted the therapeutic potential of targeting TLR4, demonstrating reduced neuronal degeneration upon TLR4 attenuation (63), and misfolded protein induced motoneuron death rescued by TLR4 inhibitors in vitro (64). However, the dual roles of the immune response in pathogenic insult and misfolded proteins need to be considered carefully for further design of immunotherapeutics targeting the CNS. Notwithstanding, TLR signalling holds a fascinating promise for therapeutic modulation of chronic pro-inflammatory signalling in disease.

References:

1. Okun E, Griffioen KJ, Mattson MP. Toll-like receptor signaling in neural plasticity and disease. Trends in neurosciences. 2011;34(5):269-81. Epub 2011/03/23.

2. Zhao W, Beers DR, Henkel JS, Zhang W, Urushitani M, Julien JP, et al. Extracellular mutant SOD1 induces microglial-mediated motoneuron injury. Glia. 2010;58(2):231-43. Epub 2009/08/13.

3. Nguyen MD, Julien JP, Rivest S. Innate immunity: the missing link in neuroprotection and neurodegeneration? Nature reviews Neuroscience. 2002;3(3):216-27. Epub 2002/05/08.

4. Scheffel J, Regen T, Van Rossum D, Seifert S, Ribes S, Nau R, et al. Toll-like receptor activation reveals developmental reorganization and unmasks responder subsets of microglia. Glia. 2012;60(12):1930-43. Epub 2012/08/23.

5. Medzhitov R, Preston-Hurlburt P, Janeway CA, Jr. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity. Nature. 1997;388(6640):394-7. Epub 1997/07/24.

6. Gillespie SK, Wasserman SA. Dorsal, a Drosophila Rel-like protein, is phosphorylated upon activation of the transmembrane protein Toll. Molecular and cellular biology. 1994;14(6):3559-68. Epub 1994/06/01.

7. Poltorak A, He X, Smirnova I, Liu MY, Van Huffel C, Du X, et al. Defective LPS signaling in C3H/HeJ and C57BL/10ScCr mice: mutations in Tlr4 gene. Science (New York, NY). 1998;282(5396):2085-8. Epub 1998/12/16.

8. Gay NJ, Keith FJ. Drosophila Toll and IL-1 receptor. Nature. 1991;351(6325):355-6. Epub 1991/05/30.

9. Wang PF, Xiong XY, Chen J, Wang YC, Duan W, Yang QW. Function and mechanism of toll-like receptors in cerebral ischemic tolerance: from preconditioning to treatment. Journal of neuroinflammation. 2015;12(1):80. Epub 2015/05/01.

10. Kigerl KA, de Rivero Vaccari JP, Dietrich WD, Popovich PG, Keane RW. Pattern recognition receptors and central nervous system repair. Experimental neurology. 2014;258:5-16. Epub 2014/07/16.

11. Burda JE, Sofroniew MV. Reactive gliosis and the multicellular response to CNS damage and disease. Neuron. 2014;81(2):229-48. Epub 2014/01/28.

12. Prokop S, Miller KR, Heppner FL. Microglia actions in Alzheimer’s disease. Acta neuropathologica. 2013;126(4):461-77. Epub 2013/11/14.

13. Maragakis NJ, Rothstein JD. Mechanisms of Disease: astrocytes in neurodegenerative disease. Nature clinical practice Neurology. 2006;2(12):679-89. Epub 2006/11/23.

14. Sasaki A, Yamaguchi H, Ogawa A, Sugihara S, Nakazato Y. Microglial activation in early stages of amyloid beta protein deposition. Acta neuropathologica. 1997;94(4):316-22. Epub 1997/10/28.

15. Falsig J, van Beek J, Hermann C, Leist M. Molecular basis for detection of invading pathogens in the brain. Journal of neuroscience research. 2008;86(7):1434-47. Epub 2007/12/07.

16. Streit WJ, Mrak RE, Griffin WS. Microglia and neuroinflammation: a pathological perspective. Journal of neuroinflammation. 2004;1(1):14. Epub 2004/08/03.

17. Lee JC, Seong J, Kim SH, Lee SJ, Cho YJ, An J, et al. Replacement of microglial cells using Clodronate liposome and bone marrow transplantation in the central nervous system of SOD1(G93A) transgenic mice as an in vivo model of amyotrophic lateral sclerosis. Biochemical and biophysical research communications. 2012;418(2):359-65. Epub 2012/01/25.

18. Gao HM, Hong JS. Why neurodegenerative diseases are progressive: uncontrolled inflammation drives disease progression. Trends in immunology. 2008;29(8):357-65. Epub 2008/07/05.

19. Kim WG, Mohney RP, Wilson B, Jeohn GH, Liu B, Hong JS. Regional difference in susceptibility to lipopolysaccharide-induced neurotoxicity in the rat brain: role of microglia. The Journal of neuroscience : the official journal of the Society for Neuroscience. 2000;20(16):6309-16. Epub 2000/08/10.

20. Heiman A, Pallottie A, Heary RF, Elkabes S. Toll-like receptors in central nervous system injury and disease: a focus on the spinal cord. Brain, behavior, and immunity. 2014;42:232-45. Epub 2014/07/27.

21. Doyle SL, O’Neill LA. Toll-like receptors: from the discovery of NFkappaB to new insights into transcriptional regulations in innate immunity. Biochemical pharmacology. 2006;72(9):1102-13. Epub 2006/08/26.

22. Casula M, Iyer AM, Spliet WG, Anink JJ, Steentjes K, Sta M, et al. Toll-like receptor signaling in amyotrophic lateral sclerosis spinal cord tissue. Neuroscience. 2011;179:233-43. Epub 2011/02/10.

23. Letiembre M, Liu Y, Walter S, Hao W, Pfander T, Wrede A, et al. Screening of innate immune receptors in neurodegenerative diseases: a similar pattern. Neurobiology of aging. 2009;30(5):759-68. Epub 2007/10/02.

24. Reed-Geaghan EG, Savage JC, Hise AG, Landreth GE. CD14 and toll-like receptors 2 and 4 are required for fibrillar A{beta}-stimulated microglial activation. The Journal of neuroscience : the official journal of the Society for Neuroscience. 2009;29(38):11982-92. Epub 2009/09/25.

25. Jin JJ, Kim HD, Maxwell JA, Li L, Fukuchi K. Toll-like receptor 4-dependent upregulation of cytokines in a transgenic mouse model of Alzheimer’s disease. Journal of neuroinflammation. 2008;5:23. Epub 2008/05/31.

26. Trudler D, Farfara D, Frenkel D. Toll-like receptors expression and signaling in glia cells in neuro-amyloidogenic diseases: towards future therapeutic application. Mediators of inflammation. 2010;2010. Epub 2010/08/14.

27. Olson JK, Miller SD. Microglia initiate central nervous system innate and adaptive immune responses through multiple TLRs. Journal of immunology (Baltimore, Md : 1950). 2004;173(6):3916-24. Epub 2004/09/10.

28. Jiang Q, Akashi S, Miyake K, Petty HR. Lipopolysaccharide induces physical proximity between CD14 and toll-like receptor 4 (TLR4) prior to nuclear translocation of NF-kappa B. Journal of immunology (Baltimore, Md : 1950). 2000;165(7):3541-4. Epub 2000/10/18.

29. Shimazu R, Akashi S, Ogata H, Nagai Y, Fukudome K, Miyake K, et al. MD-2, a molecule that confers lipopolysaccharide responsiveness on Toll-like receptor 4. The Journal of experimental medicine. 1999;189(11):1777-82. Epub 1999/06/08.

30. Ozinsky A, Underhill DM, Fontenot JD, Hajjar AM, Smith KD, Wilson CB, et al. The repertoire for pattern recognition of pathogens by the innate immune system is defined by cooperation between toll-like receptors. Proceedings of the National Academy of Sciences of the United States of America. 2000;97(25):13766-71. Epub 2000/11/30.

31. Yamamoto M, Sato S, Hemmi H, Hoshino K, Kaisho T, Sanjo H, et al. Role of adaptor TRIF in the MyD88-independent toll-like receptor signaling pathway. Science (New York, NY). 2003;301(5633):640-3. Epub 2003/07/12.

32. Horng T, Barton GM, Flavell RA, Medzhitov R. The adaptor molecule TIRAP provides signalling specificity for Toll-like receptors. Nature. 2002;420(6913):329-33. Epub 2002/11/26.

33. Fitzgerald KA, Palsson-McDermott EM, Bowie AG, Jefferies CA, Mansell AS, Brady G, et al. Mal (MyD88-adapter-like) is required for Toll-like receptor-4 signal transduction. Nature. 2001;413(6851):78-83. Epub 2001/09/07.

34. O’Neill LA, Bowie AG. The family of five: TIR-domain-containing adaptors in Toll-like receptor signalling. Nature reviews Immunology. 2007;7(5):353-64. Epub 2007/04/26.

35. Yamamoto M, Sato S, Hemmi H, Uematsu S, Hoshino K, Kaisho T, et al. TRAM is specifically involved in the Toll-like receptor 4-mediated MyD88-independent signaling pathway. Nature immunology. 2003;4(11):1144-50. Epub 2003/10/14.

36. Kawai T, Adachi O, Ogawa T, Takeda K, Akira S. Unresponsiveness of MyD88-deficient mice to endotoxin. Immunity. 1999;11(1):115-22. Epub 1999/08/06.

37. Takeda K, Akira S. TLR signaling pathways. Seminars in immunology. 2004;16(1):3-9. Epub 2004/01/31.

38. Sasai M, Yamamoto M. Pathogen recognition receptors: ligands and signaling pathways by Toll-like receptors. International reviews of immunology. 2013;32(2):116-33. Epub 2013/04/11.

39. Oshiumi H, Sasai M, Shida K, Fujita T, Matsumoto M, Seya T. TIR-containing adapter molecule (TICAM)-2, a bridging adapter recruiting to toll-like receptor 4 TICAM-1 that induces interferon-beta. The Journal of biological chemistry. 2003;278(50):49751-62. Epub 2003/10/02.

40. Verstrepen L, Bekaert T, Chau TL, Tavernier J, Chariot A, Beyaert R. TLR-4, IL-1R and TNF-R signaling to NF-kappaB: variations on a common theme. Cellular and molecular life sciences : CMLS. 2008;65(19):2964-78. Epub 2008/06/07.

41. Conze DB, Wu CJ, Thomas JA, Landstrom A, Ashwell JD. Lys63-linked polyubiquitination of IRAK-1 is required for interleukin-1 receptor- and toll-like receptor-mediated NF-kappaB activation. Molecular and cellular biology. 2008;28(10):3538-47. Epub 2008/03/19.

42. Lamothe B, Besse A, Campos AD, Webster WK, Wu H, Darnay BG. Site-specific Lys-63-linked tumor necrosis factor receptor-associated factor 6 auto-ubiquitination is a critical determinant of I kappa B kinase activation. The Journal of biological chemistry. 2007;282(6):4102-12. Epub 2006/12/01.

43. Schauvliege R, Janssens S, Beyaert R. Pellino proteins: novel players in TLR and IL-1R signalling. Journal of cellular and molecular medicine. 2007;11(3):453-61. Epub 2007/07/20.

44. Alkalay I, Yaron A, Hatzubai A, Orian A, Ciechanover A, Ben-Neriah Y. Stimulation-dependent I kappa B alpha phosphorylation marks the NF-kappa B inhibitor for degradation via the ubiquitin-proteasome pathway. Proceedings of the National Academy of Sciences of the United States of America. 1995;92(23):10599-603. Epub 1995/11/07.

45. Wertz IE, Dixit VM. Signaling to NF-kappaB: regulation by ubiquitination. Cold Spring Harbor perspectives in biology. 2010;2(3):a003350. Epub 2010/03/20.

46. Schauvliege R, Janssens S, Beyaert R. Pellino proteins are more than scaffold proteins in TLR/IL-1R signalling: a role as novel RING E3-ubiquitin-ligases. FEBS letters. 2006;580(19):4697-702. Epub 2006/08/04.

47. Cao Z, Henzel WJ, Gao X. IRAK: a kinase associated with the interleukin-1 receptor. Science (New York, NY). 1996;271(5252):1128-31. Epub 1996/02/23.

48. Gottipati S, Rao NL, Fung-Leung WP. IRAK1: a critical signaling mediator of innate immunity. Cellular signalling. 2008;20(2):269-76. Epub 2007/09/25.

49. Qian Y, Commane M, Ninomiya-Tsuji J, Matsumoto K, Li X. IRAK-mediated translocation of TRAF6 and TAB2 in the interleukin-1-induced activation of NFkappa B. The Journal of biological chemistry. 2001;276(45):41661-7. Epub 2001/08/24.

50. Sebban-Benin H, Pescatore A, Fusco F, Pascuale V, Gautheron J, Yamaoka S, et al. Identification of TRAF6-dependent NEMO polyubiquitination sites through analysis of a new NEMO mutation causing incontinentia pigmenti. Human molecular genetics. 2007;16(23):2805-15. Epub 2007/08/31.

51. Moynagh PN. The Pellino family: IRAK E3 ligases with emerging roles in innate immune signalling. Trends in immunology. 2009;30(1):33-42. Epub 2008/11/22.

52. Tseng PH, Matsuzawa A, Zhang W, Mino T, Vignali DA, Karin M. Different modes of ubiquitination of the adaptor TRAF3 selectively activate the expression of type I interferons and proinflammatory cytokines. Nature immunology. 2010;11(1):70-5. Epub 2009/11/10.

53. Xiao Y, Jin J, Chang M, Chang JH, Hu H, Zhou X, et al. Peli1 promotes microglia-mediated CNS inflammation by regulating Traf3 degradation. Nature medicine. 2013;19(5):595-602. Epub 2013/04/23.

54. Chow JC, Young DW, Golenbock DT, Christ WJ, Gusovsky F. Toll-like receptor-4 mediates lipopolysaccharide-induced signal transduction. The Journal of biological chemistry. 1999;274(16):10689-92. Epub 1999/04/10.

55. Hoshino K, Takeuchi O, Kawai T, Sanjo H, Ogawa T, Takeda Y, et al. Cutting edge: Toll-like receptor 4 (TLR4)-deficient mice are hyporesponsive to lipopolysaccharide: evidence for TLR4 as the Lps gene product. Journal of immunology (Baltimore, Md : 1950). 1999;162(7):3749-52. Epub 1999/04/14.

56. Kawai T, Akira S. Antiviral signaling through pattern recognition receptors. Journal of biochemistry. 2007;141(2):137-45. Epub 2006/12/28.

57. Sato S, Sugiyama M, Yamamoto M, Watanabe Y, Kawai T, Takeda K, et al. Toll/IL-1 receptor domain-containing adaptor inducing IFN-beta (TRIF) associates with TNF receptor-associated factor 6 and TANK-binding kinase 1, and activates two distinct transcription factors, NF-kappa B and IFN-regulatory factor-3, in the Toll-like receptor signaling. Journal of immunology (Baltimore, Md : 1950). 2003;171(8):4304-10. Epub 2003/10/08.

58. Shembade N, Harhaj EW. Elucidating dynamic protein-protein interactions and ubiquitination in NF-kappaB signaling pathways. Methods in molecular biology (Clifton, NJ). 2015;1280:283-95. Epub 2015/03/05.

59. Kawai T, Takeuchi O, Fujita T, Inoue J, Muhlradt PF, Sato S, et al. Lipopolysaccharide stimulates the MyD88-independent pathway and results in activation of IFN-regulatory factor 3 and the expression of a subset of lipopolysaccharide-inducible genes. Journal of immunology (Baltimore, Md : 1950). 2001;167(10):5887-94. Epub 2001/11/08.

60. Chang M, Jin W, Sun SC. Peli1 facilitates TRIF-dependent Toll-like receptor signaling and proinflammatory cytokine production. Nature immunology. 2009;10(10):1089-95. Epub 2009/09/08.

61. Murphy M, Xiong Y, Pattabiraman G, Qiu F, Medvedev AE. Pellino-1 positively regulates Toll-like Receptor (TLR) 2 and TLR4 signaling and is suppressed upon induction of endotoxin tolerance. The Journal of biological chemistry. 2015. Epub 2015/06/18.

62. Smith H, Liu XY, Dai L, Goh ET, Chan AT, Xi J, et al. The role of TBK1 and IKKepsilon in the expression and activation of Pellino 1. The Biochemical journal. 2011;434(3):537-48. Epub 2011/01/06.

63. Lee M, McGeer E, McGeer PL. Activated human microglia stimulate neuroblastoma cells to upregulate production of beta amyloid protein and tau: implications for Alzheimer’s disease pathogenesis. Neurobiology of aging. 2015;36(1):42-52. Epub 2014/08/30.

64. Paola M, Sestito SE, Mariani A, Memo C, Fanelli R, Freschi M, et al. Synthetic and natural small molecule TLR4 antagonists inhibit motoneuron death in cultures from ALS mouse model. Pharmacological research. 2015. Epub 2015/12/08.

7th Oct 2021 Sinead Kinsella PhD

Recent Posts