C-type lectin receptor-induced NF-κB activation in innate immune and inflammatory responses

Cellular and Molecular Immunology

Volumn 9, Issue 2, 2012, Pages 105-112

  Kingeter, Lara M ^a   Lin, Xin ^a  

^a UNIVERSITY OF TEXAS   (United States)

Author keywords

C type lectin receptor; innate immunity; NF B

Indexed keywords

C TYPE LECTIN RECEPTOR; CASPASE RECRUITMENT DOMAIN PROTEIN 15; DECTIN 1; DECTIN 2; IMMUNOGLOBULIN ENHANCER BINDING PROTEIN; INTERLEUKIN 10; INTERLEUKIN 12; LECTIN RECEPTOR; MINCLE; RAF PROTEIN; REGULATOR PROTEIN; TOLL LIKE RECEPTOR 2; TUMOR NECROSIS FACTOR ALPHA; UNCLASSIFIED DRUG;

CYTOKINE PRODUCTION; ENZYME ACTIVATION; ENZYME ACTIVITY; HUMAN; IMMUNE RESPONSE; INFLAMMATION; INNATE IMMUNITY; MOLECULAR RECOGNITION; NONHUMAN; PROTEIN BINDING; PROTEIN FUNCTION; PROTEIN PHOSPHORYLATION; PROTEIN PROTEIN INTERACTION; PROTEIN STRUCTURE; PROTEIN TARGETING; REVIEW; SIGNAL TRANSDUCTION;

ANIMALS; CARD SIGNALING ADAPTOR PROTEINS; HUMANS; IMMUNITY, INNATE; INFLAMMATION; INTRACELLULAR SIGNALING PEPTIDES AND PROTEINS; LECTINS, C-TYPE; NF-KAPPA B; PROTEIN-TYROSINE KINASES; RECEPTORS, IGG; RECEPTORS, PATTERN RECOGNITION; SIGNAL TRANSDUCTION; TRANSCRIPTIONAL ACTIVATION;

EID: 84858142445     PISSN: 16727681     EISSN: None     Source Type: Journal    
DOI: 10.1038/cmi.2011.58     Document Type: Review

References (95)

1. Niedergang F, Chavrier P. Regulation of phagocytosis by Rho GTPases. Curr Top Microbiol Immunol 2005; 291: 43-60. (Pubitemid 40674910)
2. Kumar H, Kawai T, Akira S. Pathogen recognition by the innate immune system. Int Rev Immunol 2011; 30: 16-34.
3. Yamamoto M, Takeda K. Current views of Toll-like receptor signaling pathways. Gastroenterol Res Pract. 2010; 2010: 240365.
4. Kawai T, Akira S. TLR signaling. Cell Death Differ 2006; 13: 816-825.
5. Kawai T, Akira S. Toll-like receptors and their crosstalk with other innate receptors in infection and immunity. Immunity 2011; 34: 637-650.
6. Muzio M, Ni J, Feng P, Dixit VM. IRAK (Pelle) family member IRAK-2 and MyD88 as proximal mediators of IL-1 signaling. Science 1997; 278: 1612-1615. (Pubitemid 27516277)
7. Takeuchi O, Kaufmann A, Grote K, Kawai T, Hoshino K, Morr M et al. Cutting edge: preferentially the R-stereoisomer of the mycoplasmal lipopeptide macrophageactivating lipopeptide-2 activates immune cells through a Toll-like receptor 2- and MyD88-dependent signaling pathway. J Immunol 2000; 164: 554-557. (Pubitemid 30043671)
8. 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 2003; 301: 640-643. (Pubitemid 36927946)
9. Loo YM, Gale M Jr. Immune signaling by RIG-I-like receptors. Immunity 2011; 34: 680-692.
10. Kawai T, Takahashi K, Sato S, Coban C, Kumar H, Kato H, et al. IPS-1, an adaptor triggering RIG-I- and Mda5-mediated type I interferon induction. Nat Immunol 2005; 6: 981-988. (Pubitemid 41486182)
11. Meylan E, Curran J, Hofmann K, Moradpour D, Binder M, BartenschlagerRet al. Cardif is an adaptor protein in the RIG-I antiviral pathway and is targeted by hepatitis C virus. Nature 2005; 437: 1167-1172. (Pubitemid 41509355)
12. Seth RB, Sun L, Ea CK, Chen ZJ. Identification and characterization of MAVS, a mitochondrial antiviral signaling protein that activates NF-kappaB and IRF 3. Cell 2005; 122: 669-682. (Pubitemid 41242633)
13. Xu LG, Wang YY, Han KJ, Li LY, Zhai Z, Shu HB. VISA is an adapter protein required for virus-triggered IFN-beta signaling. Mol Cell 2005; 19: 727-740. (Pubitemid 41316668)
14. Martinon F, Tschopp J. Inflammatory caspases: linking an intracellular innate immune system to autoinflammatory diseases. Cell 2004; 117: 561-574. (Pubitemid 38692523)
15. Elinav E, Strowig T, Henao-Mejia J, Flavell RA. Regulation of the antimicrobial response by NLR proteins. Immunity 2011; 34: 665-679.
16. Veldhuizen EJ, van Eijk M, Haagsman HP. The carbohydrate recognition domain of collectins. FEBS J 2011; 278: 3930-3941.
17. Kanazawa N, Tashiro K, Miyachi Y. Signaling and immune regulatory role of the dendritic cell immunoreceptor (DCIR) family lectins: DCIR, DCAR, dectin-2 and BDCA-2. Immunobiology 2004; 209: 179-190. (Pubitemid 39255702)
18. Kanazawa N. Dendritic cell immunoreceptors: C-type lectin receptors for patternrecognition and signaling on antigen-presenting cells. J Dermatol Sci 2007; 45: 77-86. (Pubitemid 46108481)
19. Graham LM, Brown GD. The Dectin-2 family of C-type lectins in immunity and homeostasis. Cytokine 2009; 48: 148-155.
20. Huysamen C, Brown GD. The fungal pattern recognition receptor, Dectin-1, and the associated cluster of C-type lectin-like receptors. FEMS Microbiol Lett 2009; 290: 121-128.
21. Cao W, Zhang L, Rosen DB, Bover L, Watanabe G Bao M et al. BDCA2/Fc epsilon RI gamma complex signals through a novel BCR-like pathway in human plasmacytoid dendritic cellsPLoS Biol 2007; 5: e248.
22. Yamasaki S, Ishikawa E, Sakuma M, Hara H, Ogata K, Saito T. Mincle is an ITAMcoupled activating receptor that senses damaged cells. Nat Immunol 2008; 9: 1179-1188.
23. Kanazawa N, Tashiro K, Inaba K, Miyachi Y. Dendritic cell immunoactivating receptor, a novel C-type lectin immunoreceptor, acts as an activating receptor through association with Fc receptor gamma chain. J Biol Chem 2003; 278: 32645-32652 (Pubitemid 37055703)
24. Kanazawa N, Tashiro K, Inaba K, Lutz MB, Miyachi Y. Molecular cloning of human dectin-2. J Invest Dermatol 2004; 122: 1522-1524.
25. Brown GD, Gordon S. Immune recognition. A new receptor for beta-glucans. Nature 2001; 413: 36-37. (Pubitemid 32843478)
26. Brown GD, Taylor PR, Reid DM, Willment JA, Williams DL, Martinez-Pomares L et al. Dectin-1 is a major beta-glucan receptor on macrophages. J Exp Med 2002; 196: 407-412. (Pubitemid 34875669)
27. McGreal EP, Rosas M, Brown GD, Zamze S, Wong SY, Gordon S et al. The carbohydrate-recognition domain of Dectin-2 is a C-type lectin with specificity for high mannose. Glycobiology 2006; 16: 422-430.
28. Sato K, Yang XL, Yudate T, Chung JS, Wu J, Luby-Phelps K et al. Dectin-2 is a pattern recognition receptor for fungi that couples with the Fc receptor gamma chain to induce innate immune responses. J Biol Chem 2006; 281: 38854-38866. (Pubitemid 46042012)
29. Saijo S, Ikeda S, Yamabe K, Kakuta S, Ishigame H, Akitsu A et al. Dectin-2 recognition of alpha-mannans and induction of Th17 cell differentiation is essential for host defense against Candida albicans. Immunity 2010; 32: 681-691.
30. Ishikawa E, Ishikawa T, Morita YS, Toyonaga K, Yamada H, Takeuchi O et al. Direct recognition of the mycobacterial glycolipid, trehalose dimycolate, by C-type lectin Mincle. J Exp Med 2009; 206: 2879-2888.
31. Schoenen H, Bodendorfer B, Hitchens K, Manzanero S, Werninghaus K, Nimmerjahn F et al. Cutting edge: Mincle is essential for recognition and adjuvanticity of the mycobacterial cord factor and its synthetic analog trehalos e-dibehenate. J Immunol 2010; 184: 2756-2760.
32. Yamasaki S, Matsumoto M, Takeuchi O, Matsuzawa T, Ishikawa E, Sakuma M et al. C-type lectin Mincle is an activating receptor for pathogenic fungus, Malassezia. Proc Natl Acad Sci USA 2009; 106: 1897-1902.
33. Gross O, Gewies A, Finger K, Schafer M, Sparwasser T, Peschel C et al. Card9 controls a non-TLR signalling pathway for innate anti-fungal immunity. Nature 2006; 442: 651-656. (Pubitemid 44215314)
34. Oeckinghaus A, Hayden MS, Ghosh S. Crosstalk in NF-kappaB signaling pathways. Nat Immunol 2011; 12: 695-708.
35. Vallabhapurapu S, Karin M. Regulation and function of NF-kappaB transcription factors in the immune system. Annu Rev Immunol 2009; 27: 693-733.
36. Chen ZJ, Parent L, Maniatis T. Site-specific phosphorylation of IkappaBalpha by a novel ubiquitination-dependent protein kinase activity. Cell 1996; 84: 853-862. (Pubitemid 26106855)
37. Tang ED, Wang CY, Xiong Y, Guan KL. A role for NF-kappaB essential modifier/ IkappaB kinase-gamma (NEMO/IKKgamma) ubiquitination in the activation of the IkappaB kinase complex by tumor necrosis factor-alpha. J Biol Chem 2003; 278: 37297-37305. (Pubitemid 37175246)
38. Zhou H, Wertz I, O'Rourke K, Ultsch M, Seshagiri S, Eby M et al. Bcl10 activates the NF-kappaB pathway through ubiquitination of NEMO. Nature 2004; 427: 167-171. (Pubitemid 38094958)
39. Delhase M, Hayakawa M, Chen Y, Karin M. Positive and negative regulation of IkappaB kinase activity through IKKbeta subunit phosphorylation. Science 1999; 284: 309-313.
40. Schmid JA, Birbach A. IkappaB kinase beta (IKKbeta/IKK2/IKBKB)-a key molecule in signaling to the transcription factor NF-kappaB. Cytokine Growth Factor Rev 2008; 19: 157-165. (Pubitemid 351382281)
41. Chen Z, Hagler J, Palombella VJ, Melandri F, Scherer D, Ballard D et al. Signalinduced site-specific phosphorylation targets I kappa B alpha to the ubiquitinproteasome pathway. Genes Dev 1995; 9: 1586-1597.
42. Scherer DC, Brockman JA, Chen Z, Maniatis T, Ballard DW. Signal-induced degradation of I kappa B alpha requires site-specific ubiquitination. Proc Natl Acad Sci USA 1995; 92: 11259-11263. (Pubitemid 3015352)
43. Vermeulen L, de Wilde G, Notebaert S, Vanden Berghe W, Haegeman G. Regulation of the transcriptional activity of the nuclear factor-kappaB p65 subunit. Biochem Pharmacol 2002; 64: 963-970. (Pubitemid 35232270)
44. Chen LF, Greene WC. Shaping the nuclear action of NF-kappaB. Nat Rev Mol Cell Biol 2004; 5: 392-401.
45. Sun SC. Non-canonical NF-kappaB signaling pathway. Cell Res 2011; 21: 71-85.
46. Underhill DM, Goodridge HS. The many faces of ITAMs. Trends Immunol 2007; 28: 66-73. (Pubitemid 46176733)
47. Kerrigan AM, Brown GD. Syk-coupled C-type lectins in immunity. Trends Immunol 2011; 32: 151-156.
48. Mocsai A, Ruland J, Tybulewicz VL. The SYK tyrosine kinase: a crucial player in diverse biological functions. Nat Rev Immunol 2010; 10: 387-402.
49. Underhill DM, Rossnagle E, Lowell CA, Simmons RM. Dectin-1 activates Syk tyrosine kinase in a dynamic subset of macrophages for reactive oxygen production. Blood 2005; 106: 2543-2550. (Pubitemid 41510832)
50. Rogers NC, Slack EC, Edwards AD, Nolte MA, Schulz O, Schweighoffer E et al. Sykdependent cytokine induction by Dectin-1 reveals a novel pattern recognition pathway for C type lectins. Immunity 2005; 22: 507-517. (Pubitemid 40545115)
51. Robinson MJ, Osorio F, Rosas M, Freitas RP, Schweighoffer E, Gross O et al. Dectin-2 is a Syk-coupled pattern recognition receptor crucial for Th17 responses to fungal infection. J Exp Med 2009; 206: 2037-2051.
52. LeibundGut-Landmann S, Gross O, Robinson MJ, Osorio F, Slack EC, Tsoni SV et al. Syk- and CARD9-dependent coupling of innate immunity to the induction of T helper cells that produce interleukin 17. Nat Immunol 2007; 8: 630-638. (Pubitemid 46785127)
53. Hara H, Ishihara C, Takeuchi A, Imanishi T, Xue L, MorrisSWet al. The adaptor protein CARD9 is essential for the activation of myeloid cells through ITAM-associated and Toll-like receptors. Nat Immunol 2007; 8: 619-629. (Pubitemid 46785126)
54. Thome M. CARMA1, BCL-10 and MALT1 in lymphocyte development and activation. Nat Rev Immunol 2004; 4: 348-359. (Pubitemid 38649277)
55. Wegener E, Krappmann D. CARD-Bcl10-Malt1 signalosomes: missing link to NFkappaBSci STKE 2007; 2007: pe21
56. Hara H, Saito T. CARD9 versus CARMA1 in innate and adaptive immunity. Trends Immunol 2009; 30: 234-242.
57. Bi L, Gojestani S, Wu W, Hsu YM, Zhu J, Ariizumi K et al. CARD9 mediates dectin-2- induced IkappaBalpha kinase ubiquitination leading to activation of NF-kappaB in response to stimulation by the hyphal form of Candida albicans. J Biol Chem 2010; 285: 25969-25977.
58. Gorjestani S, Yu M, Tang B, Zhang D, Wang D, Lin X. Phospholipase Cgamma2 (PLCgamma2) is a key component in Dectin-2 signaling pathway, mediating antifungal innate immune responses. J Biol Chem 2011; 286: 43651-43659.
59. Gringhuis SI, den Dunnen J, Litjens M, van der Vlist M, Wevers B, Bruijns SC et al. Dectin-1 directs T helper cell differentiation by controlling noncanonical NF-kappaB activation through Raf-1 and Syk. Nat Immunol 2009; 10: 203-213.
60. 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. Proc Natl Acad Sci USA 2000; 97: 13766-13771.
61. Gantner BN, Simmons RM, Canavera SJ, Akira S, Underhill DM. Collaborative induction of inflammatory responses by dectin-1 and Toll-like receptor 2. J Exp Med 2003; 197: 1107-1117. (Pubitemid 36570066)
62. Brown GD, Herre J, Williams DL, Willment JA, Marshall AS, Gordon S. Dectin-1 mediates the biological effects of beta-glucans. J Exp Med 2003; 197: 1119-1124. (Pubitemid 36570067)
63. DiCarlo EF, Kahn LB. Inflammatory diseases of the bones and joints. Semin Diagn Pathol 2011; 28: 53-64.
64. Rosenzweig HL, Clowers JS, Nunez G, Rosenbaum JT, Davey MP. Dectin-1 and NOD2 mediate cathepsin activation in zymosan-induced arthritis in mice. Inflamm Res 2011; 60: 705-714.
65. Esteban A, Popp MW, Vyas VK, Strijbis K, Ploegh HL, Fink GR. Fungal recognition is mediated by the association of dectin-1 and galectin-3 in macrophages. Proc Natl Acad Sci USA 2011; 108: 14270-1425.
66. Jouault T, El Abed-El Behi M, Martinez-Esparza M, Breuilh L, Trinel PA, Chamaillard M et al. Specific recognition of Candida albicans by macrophages requires galectin-3 to discriminate Saccharomyces cerevisiae and needs association with TLR2 for signaling. J Immunol 2006; 177: 4679-4687. (Pubitemid 44469801)
67. Geijtenbeek TB, Torensma R, van Vliet SJ, van Duijnhoven GC, Adema GJ, van Kooyk Y et al. Identification of DC-SIGN, a novel dendritic cell-specific ICAM-3 receptor that supports primary immune responses. Cell 2000; 100: 575-585.
68. Lozach PY, Kuhbacher A, Meier R, Mancini R, Bitto D, Bouloy M et al. DC-SIGN as areceptor for phleboviruses. Cell Host Microbe 2011; 10: 75-88.
69. Geijtenbeek TB, Kwon DS, Torensma R, van Vliet SJ, van Duijnhoven GC, Middel J et al. DC-SIGN, a dendritic cell-specific HIV-1-binding protein that enhances transinfection of T cells. Cell 2000; 100: 587-597.
70. Alvarez CP, Lasala F, Carrillo J, Muniz O, Corbi AL, Delgado R. C-type lectins DC-SIGN and L-SIGN mediate cellular entry by Ebola virus in cis and in trans. J Virol 2002; 76: 6841-6844. (Pubitemid 34633551)
71. Lozach PY, Lortat-Jacob H, de Lacroix de Lavalette A, Staropoli I, Foung S, Amara A et al. DC-SIGN and L-SIGN are high affinity binding receptors for hepatitis C virus glycoprotein E2. J Biol Chem 2003; 278: 20358-20366. (Pubitemid 36799236)
72. Tassaneetrithep B, Burgess TH, Granelli-Piperno A, Trumpfheller C, Finke J, Sun W et al. DC-SIGN (CD209) mediates dengue virus infection of human dendritic cells. J Exp Med 2003; 197: 823-829. (Pubitemid 36437860)
73. Tailleux L, Schwartz O, Herrmann JL, Pivert E, Jackson M, Amara A et al. DC-SIGN is the major Mycobacterium tuberculosis receptor on human dendritic cells. J Exp Med 2003; 197: 121-127. (Pubitemid 36110263)
74. Cambi A, Gijzen K, de Vries JM, Torensma R, Joosten B, Adema GJ et al. The C-type lectin DC-SIGN (CD209) is an antigen-uptake receptor for Candida albicans on dendritic cells. Eur J Immunol 2003; 33: 532-538. (Pubitemid 36267619)
75. Geijtenbeek TB, van Vliet SJ, Koppel EA, Sanchez-Hernandez M, Vandenbroucke- Grauls CM, Appelmelk B et al. Mycobacteria target DC-SIGN to suppress dendritic cell function. J Exp Med 2003; 197: 7-17. (Pubitemid 36110253)
76. Nigou J, Zelle-Rieser C, Gilleron M, Thurnher M, Puzo G. Mannosylated lipoarabinomannans inhibit IL-12 production by human dendritic cells: evidence for a negative signal delivered through the mannose receptor. J Immunol 2001; 166: 7477-7485. (Pubitemid 32525626)
77. H-17 immunity by dectin-1 and dectin-2PLoS PathogPLoS Pathog 2011; 7: e1001259.
78. Cheng SC, van de Veerdonk FL, Lenardon M, Stoffels M, Plantinga T, SmeekensS et al. The dectin-1/inflammasome pathway is responsible for the induction of protective Thelper 17 responses that discriminate between yeasts and hyphae of Candida albicans. J Leukoc Biol 2011; 90: 357-366.
79. Cunha C, Di Ianni M, Bozza S, Giovannini G, Zagarella S, Zelante T, et al. Dectin-1 Y238X polymorphism associates with susceptibility to invasive aspergillosis in hematopoietic transplantation through impairment of both recipient- and donordependent mechanisms of antifungal immunity. Blood 2010; 116: 5394-5402.
80. Taylor PR, Tsoni SV, Willment JA, Dennehy KM, Rosas M, Findon H et al. Dectin-1 is required for beta-glucan recognition and control of fungal infection. Nat Immunol 2007; 8: 31-38. (Pubitemid 46242356)
81. Werner JL, Metz AE, Horn D, Schoeb TR, Hewitt MM, Schwiebert LM et al. Requisite role for the dectin-1 beta-glucan receptor in pulmonary defense against Aspergillus fumigatus. J Immunol 2009; 182: 4938-4946.
82. Saijo S, Fujikado N, Furuta T, Chung SH, Kotaki H, SekiKet al. Dectin-1 is required for host defense against Pneumocystis carinii but not against Candida albicans. Nat Immunol 2007; 8: 39-46. (Pubitemid 46242357)
83. Ferwerda B, Ferwerda G, Plantinga TS, Willment JA, van Spriel AB, Venselaar H et al. Human dectin-1 deficiency and mucocutaneous fungal infections. N Engl J Med 2009; 361: 1760-1767.
84. Sousa Mda G, Reid DM, Schweighoffer E, Tybulewicz V, Ruland J, Langhorne J et al. Restoration of pattern recognition receptor costimulation to treat chromoblastomycosis, a chronic fungal infection of the skin. Cell Host Microbe 2011; 9: 436-443.
85. Glocker EO, Hennigs A, Nabavi M, Schaffer AA, Woellner C, Salzer U et al. A homozygous CARD9 mutation in a family with susceptibility to fungal infections. N Engl J Med 2009; 361: 1727-1735.
86. Unger WW, van Kooyk Y. 'Dressed for success' C-type lectin receptors for the delivery of glyco-vaccines to dendritic cells. Curr Opin Immunol 2011; 23: 131-137.
87. Bonifaz LC, Bonnyay DP, Charalambous A, Darguste DI, Fujii S, Soares H et al. In vivo targeting of antigens to maturing dendritic cells via the DEC-205 receptor improves T cell vaccination. J Exp Med 2004; 199: 815-824. (Pubitemid 38375201)
88. Demangel C, Zhou J, Choo AB, Shoebridge G, Halliday GM, Britton WJ. Single chain antibody fragments for the selective targeting of antigens to dendritic cells. Mol Immunol 2005; 42: 979-985. (Pubitemid 40489023)
89. Trumpfheller C, Finke JS, Lopez CB, Moran TM, Moltedo B, Soares H et al. Intensified and protective CD41 T cell immunity in mice with anti-dendritic cell HIV gag fusion antibody vaccine. J Exp Med 2006; 203: 607-617
90. Nchinda G, Kuroiwa J, Oks M, Trumpfheller C, Park CG, Huang Y et al. The efficacy of DNA vaccination is enhanced in mice by targeting the encoded protein to dendritic cells. J Clin Invest 2008; 118: 1427-1436. (Pubitemid 351500436)
91. Idoyaga J, Lubkin A, Fiorese C, Lahoud MH, Caminschi I, Huang Y et al. Comparable T helper 1 (Th1) and CD8 T-cell immunity by targeting HIV gag p24 to CD8 dendritic cells within antibodies to Langerin, DEC205, and Clec9A. Proc Natl Acad Sci USA 2011; 108: 2384-2389.
92. Mukhopadhaya A, Hanafusa T, Jarchum I, Chen YG, Iwai Y, Serreze DV et al. Selective delivery of beta cell antigen to dendritic cells in vivo leads to deletion and tolerance of autoreactive CD81 T cells in NODmice. Proc Natl Acad Sci USA 2008; 105: 6374-6379. (Pubitemid 351758353)
93. Karumuthil-Melethil S, Perez N, Li R, Vasu C. Induction of innate immune response through TLR2 and dectin 1 prevents type 1 diabetes. J Immunol 2008; 181: 8323-8334.
94. He LZ, Crocker A, Lee J, Mendoza-Ramirez J, Wang XT, Vitale LA et al. Antigenic targeting of the human mannose receptor induces tumor immunity. J Immunol 2007; 178: 6259-6267. (Pubitemid 46717409)
95. Ramakrishna V, Treml JF, Vitale L, Connolly JE, O'Neill T, Smith PA et al. Mannose receptor targeting of tumor antigen pmel17 to human dendritic cells directs antimelanoma T cell responses via multiple HLA molecules. J Immunol 2004; 172: 2845-2452.