Tolerance induced by the lipopeptide Pam3Cys is due to ablation of IL-1R-associated kinase-1.
Stimulation of the human monocytic cell line Mono Mac 6 with the synthetic lipopeptide (S)-(2,3-bis(palmitoyloxy)-(2RS)-propyl)-N-palmitoyl-(R)-Cys-(S)-Ser(S)-Lys(4)-OH, trihydrochloride (Pam(3)Cys) at 10 microg/ml induces a rapid expression of the TNF gene in a TLR2-dependent fashion. Preculture of the cells with Pam(3)Cys at 1 microg/ml leads to a reduced response after subsequent stimulation with Pam(3)Cys at 10 microg/ml, indicating that the cells have become tolerant to Pam(3)Cys. The CD14 and TLR2 expression is not decreased on the surface of the tolerant cells, but rather up-regulated. Analysis of the NF-kappaB binding in Pam(3)Cys-tolerant cells shows a failure to mobilize NF-kappaB-p50p65 heterodimers, while NF-kappaB-p50p50 homodimers remain unchanged. Pam(3)Cys-tolerant cells showed neither IkappaBalpha-Ser(32) phosphorylation nor IkappaBalpha degradation but MyD88 protein was unaltered. However, IRAK-1 protein was absent in Pam(3)Cys-induced tolerance, while IRAK-1 mRNA was still detectable at 30% compared with untreated cells. In contrast, in LPS-tolerized cells, p50p50 homodimers were induced, IRAK-1 protein level was only partially decreased, and p50p65 mobilization remained intact. It is concluded that in Mono Mac 6 monocytic cells, inhibition of IRAK-1 expression at the mRNA and protein levels is the main TLR-2-dependent mechanism responsible for Pam(3)Cys-induced tolerance, but not for TLR-4-dependent LPS-induced tolerance.
Institute for Immunology, University of Muenchen, Muenchen, Germany.
Maciej
M
Siedlar
Marion
M
Frankenberger
Elke
E
Benkhart
Terje
T
Espevik
Martina
M
Quirling
Korbinian
K
Brand
Marek
M
Zembala
Loems
L
Ziegler-Heitbrock
D048868:Adaptor Proteins, Signal Transducing; D000943:Antigens, Differentiation / Q000276:immunology / Q000378:metabolism; D015153:Blotting, Western; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D006801:Humans; D020672:I-kappa B Proteins / Q000276:immunology / Q000378:metabolism; D007108:Immune Tolerance; D053592:Interleukin-1 Receptor-Associated Kinases; D018950:Lipopolysaccharide Receptors / Q000276:immunology / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology / Q000494:pharmacology; D008074:Lipoproteins / Q000276:immunology / Q000494:pharmacology*; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism; D009000:Monocytes / Q000187:drug effects* / Q000276:immunology; D053594:Myeloid Differentiation Factor 88; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000276:immunology / Q000378:metabolism; D011494:Protein Kinases / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000276:immunology / Q000378:metabolism; D011971:Receptors, Immunologic / Q000276:immunology / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000276:immunology; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D000943:Antigens, Differentiation; D020672:I-kappa B Proteins; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008074:Lipoproteins; C507779:MYD88 protein, human; D008562:Membrane Glycoproteins; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D011494:Protein Kinases; D053592:Interleukin-1 Receptor-Associated Kinases