The innate immune response is a complex process involving multiple pathogen-recognition receptors, including toll-like receptors (TLRs) and nucleotide-binding oligomerization domain (NOD)-like receptors. Complement is also a critical component of innate immunity. While complement is known to interact with TLR-mediated signals, the interactions between NOD-like receptors and complement are not well understood. Here we report a synergistic interaction between C5a and Nod2 signaling in RAW 264.7 marophages. Long-term treatment with muramyl dipeptide (MDP), a NOD2 ligand, enhanced C5a-mediated expression of chemokine mRNAs in RAW 264.7 cells. This response was dependent on NOD2 expression and was associated with a decrease in expression of C5L2, a receptor for C5a which acts as a negative modulator of C5a receptor (C5aR) activity. MDP amplified C5a-mediated phosphorylation of p38 MAPK. Treatment of RAW264.7 cells with an inhibitor of p38 attenuated the synergistic effects of C5aon MDP-primed cells on MIP-2, but not MCP-1, mRNA. In contrast, inhibition of AKT prevented C5a stimulation of MCP-1, but not MIP-2, mRNA, in MDP-primed cells. Taken together, these data demonstrated a synergistic interaction between C5a and NOD2 in the regulation of chemokine expression in macrophages, associated with a down-regulation of C5L2, a negative regulator of C5a receptor activity.
Raby, A.C., Holst, B., Davies, J., Colmont, C., Laumonnier, Y., Coles, B., Shah, S., Hall, J., Topley, N., Kohl, J., Morgan, B.P. and Labeta, M.O. (2011) TLR activation enhances C5a-induced pro-inflammatory responses by negatively modulating the second C5a receptor, C5L2. European Journal of Immunology, 41, 2741-2752. doi:10.1002/eji.201041350
Chen, N.J., Mirtsos, C., Suh, D., Lu, Y.C., Lin, W.J., McKerlie, C., Lee, T., Baribault, H., Tian, H. and Yeh, W.C. (2007) C5L2 is critical for the biological activities of the anaphylatoxins C5a and C3a. Nature, 446, 203-207. doi:10.1038/nature05559
Bamberg, C.E., Mackay, C.R., Lee, H., Zahra, D., Jackson, J., Lim, Y.S., Whitfeld, P.L., Craig, S., Corsini, E., Lu, B., Gerard, C. and Gerard, N.P. (2010) The C5a receptor (C5aR) C5L2 is a modulator of C5aR-mediated signal transduction. Journal of Biological Chemistry, 285, 7633-7644. doi:10.1074/jbc.M109.092106
Cui, W., Simaan, M., Laporte, S., Lodge, R. and Cianflone, K. (2009) C5a- and ASP-mediated C5L2 activation, endocytosis and recycling are lost in S323I-C5L2 mutation. Molecular Immunology, 46, 3086-3098. doi:10.1016/j.molimm.2009.06.007
Hajishengallis, G. and Lambris, J.D. (2010) Crosstalk pathways between Toll-like receptors and the complement system. Trends in Immunology, 31, 154-163. doi:10.1016/j.it.2010.01.002
Akira, S., Uematsu, S. and Takeuchi, O. (2006) Pathogen recognition and innate immunity. Cell, 124, 783-801. doi:10.1016/j.cell.2006.02.015
Inohara, N. and Nunez, G. (2003) NODs: Intracellular proteins involved in inflammation and apoptosis. Nature Reviews Immunology, 3, 371-382. doi:10.1038/nri1086
Ogura, Y., Bonen, D.K., Inohara, N., Nicolae, D.L., Chen, F.F., Ramos, R., Britton, H., Moran, T., Karaliuskas, R., Duerr, R.H., Achkar, J.P., Brant, S.R., Bayless, T.M., Kirschner, B.S., Hanauer, S.B., Nunez, G. and Cho, J.H. (2001) A frameshift mutation in NOD2 associated with susceptibility to Crohn’s disease. Nature, 411, 603-606. doi:10.1038/35079114
Hugot, J.P., Chamaillard, M., Zouali, H., Lesage, S., Cezard, J.P., Belaiche, J., Almer, S., Tysk, C., O’Morain, C. A., Gassull, M., Binder, V., Finkel, Y., Cortot, A., Modigliani, R., Laurent-Puig, P., Gower-Rousseau, C., Macry, J., Colombel, J.F., Sahbatou, M. and Thomas, G. (2001) Association of NOD2 leucine-rich repeat variants with susceptibility to Crohn’s disease. Nature, 411, 599-603. doi:10.1038/35079107
Strober, W., Murray, P.J., Kitani, A. and Watanabe, T. (2006) Signalling pathways and molecular interactions of NOD1 and NOD2. Nature Reviews Immunology, 6, 9-20. doi:10.1038/nri1747
Girardin, S.E., Boneca, I.G., Viala, J., Chamaillard, M., Labigne, A., Thomas, G., Philpott, D.J. and Sansonetti, P.J. (2003) Nod2 is a general sensor of peptidoglycan through muramyl dipeptide (MDP) detection. Journal of Biological Chemistry, 278, 8869-8872. doi:10.1074/jbc.C200651200
Pauleau, A.L. and Murray, P.J. (2003) Role of nod2 in the response of macrophages to toll-like receptor agonists. Molecular and Cellular Biology, 23, 7531-7539. doi:10.1128/MCB.23.21.7531-7539.2003
Kobayashi, K.S., Chamaillard, M., Ogura, Y., Henegariu, O., Inohara, N., Nunez, G. and Flavell, R.A. (2005) Nod2-dependent regulation of innate and adaptive immunity in the intestinal tract. Science, 307, 731-734. doi:10.1126/science.1104911
McMullen, M.R., Pritchard, M.T., Wang, Q., Millward, C.A., Croniger, C.M. and Nagy, L.E. (2005) Early growth response-1 transcription factor is essential for ethanol-induced Fatty liver injury in mice. Gastroenterology, 128, 2066-2076. doi:10.1053/j.gastro.2005.02.065
Fritz, J.H., Girardin, S.E., Fitting, C., Werts, C., Mengin-Lecreulx, D., Caroff, M., Cavaillon, J.M., Philpott, D.J. and Adib-Conquy, M. (2005) Synergistic stimulation of human monocytes and dendritic cells by Toll-like recaptor 4 and NOD1-and NOD2-activating agonists. European Journal of Immunology, 35, 2459-2470. doi:10.1002/eji.200526286
Argast, G.M., Fausto, N. and Campbell, J.S. (2005) Inhibition of RIP2/RIck/CARDIAK activity by pyridinyl imidazole inhibitors of p38 MAPK. Molecular and Cellular Biology, 268, 129-140. doi:10.1007/s11010-005-3701-0
Song, W.C. (2012) Crosstalk between complement and toll-like receptors. Toxicologic Pathology, 40, 174-182. doi:10.1177/0192623311428478
Hawlisch, H., Belkaid, Y., Baelder, R., Hildeman, D., Gerard, C. and Kohl, J. (2005) C5a negatively regulates toll-like receptor 4-induced immune responses. Immunity, 22, 415-426. doi:10.1016/j.immuni.2005.02.006
Ward, P.A. (2010) The harmful role of c5a on innate immunity in sepsis. Journal of Innate Immunity, 2, 439-445. doi:10.1159/000317194
Cain, S.A. and Monk, P.N. (2002) The orphan receptor C5L2 has high affinity binding sites for complement fragments C5a and C5a des-Arg(74). Journal of Biological Chemistry, 277, 7165-7169. doi:10.1074/jbc.C100714200
Okinaga, S., Slattery, D., Humbles, A., Zsengeller, Z., Morteau, O., Kinrade, M.B., Brodbeck, R.M., Krause, J.E., Choe, H.R., Gerard, N.P. and Gerard, C. (2003) C5L2, a nonsignaling C5A binding protein. Biochemistry, 42, 9406-9415. doi:10.1021/bi034489v
Rittirsch, D., Flierl, M.A., Nadeau, B.A., Day, D.E., Huber-Lang, M., Mackay, C.R., Zetoune, F.S., Gerard, N.P., Cianflone, K., Kohl, J., Gerard, C., Sarma, J.V. and Ward, P.A. (2008) Functional roles for C5a receptors in sepsis. Nature Medicine, 14, 551-557. doi:10.1038/nm1753
Hao, J., Meng, L.Q., Xu, P.C., Chen, M. and Zhao, M.H. (2012) p38MAPK, ERK and PI3K signaling pathways are involved in C5a-primed neutrophils for ANCA-mediated activation. PLoS One, 7, e38317. doi:10.1371/journal.pone.0038317
Cao, Q., McIsaac, S.M. and Stadnyk, A.W. (2012) Human colonic epithelial cells detect and respond to C5a via apically expressed C5aR through the ERK pathway. American Journal of Physiology: Cell Physiology, 302, C1731-C1740. doi:10.1152/ajpcell.00213.2011
Yan, C., Zhu, M., Staiger, J., Johnson, P.F. and Gao, H. (2012) C5a-regulated CCAAT/enhancer-binding proteins beta and delta are essential in Fcgamma receptor-mediated inflammatory cytokine and chemokine production in macrophages. Journal of Biological Chemistry, 287, 3217-3230. doi:10.1074/jbc.M111.280834
Marks, D.J., Harbord, M.W., MacAllister, R., Rahman, F.Z., Young, J., Al-Lazikani, B., Lees, W., Novelli, M., Bloom, S. and Segal, A.W. (2006) Defective acute inflammation in Crohn’s disease: A clinical investigation. Lancet, 367, 668-678. doi:10.1016/S0140-6736(06)68265-2
Bonen, D.K., Ogura, Y., Nicolae, D.L., Inohara, N., Saab, L., Tanabe, T., Chen, F.F., Foster, S. J., Duerr, R. H., Brant, S.R., Cho, J.H. and Nunez, G. (2003) Crohn’s disease-associated NOD2 variants share a signaling defect in response to lipopolysaccharide and peptidoglycan. Gastroenterology, 124, 140-146. doi:10.1053/gast.2003.50019
Johswich, K., Martin, M., Bleich, A., Kracht, M., Dittrich-Breiholz, O., Gessner, J.E., Suerbaum, S., Wende, E., Rheinheimer, C. and Klos, A. (2009) Role of the C5a receptor (C5aR) in acute and chronic dextran sulfate-induced models of inflammatory bowel disease. Inflammatory Bowel Disease, 15, 1812.