The Role of Interleukin 1 Receptor-associated Kinase-4 (IRAK-4) Kinase Activity in IRAK-4-mediated Signaling

Interleukin 1 receptor (IL-1R)-associated kinase-4 (IRAK-4) is required for various responses induced by IL-1R and Toll-like receptor signals. However, the molecular mechanism of IRAK-4 signaling and the role of its kinase activity have remained elusive. In this report, we demonstrate that IRAK-4 is recruited to the IL-1R complex upon IL-1 stimulation and is required for the recruitment of IRAK-1 and its subsequent activation/degradation. By reconstituting IRAK-4-deficient cells with wild type or kinase-inactive IRAK-4, we show that the kinase activity of IRAK-4 is required for the optimal transduction of IL-1-induced signals, including the activation of IRAK-1, NF-κB, and JNK, and the maximal induction of inflammatory cytokines. Interestingly, we also discover that the IRAK-4 kinase-inactive mutant is still capable of mediating some signals. These results suggest that IRAK-4 is an integral part of the IL-1R signaling cascade and is capable of transmitting signals both dependent on and independent of its kinase activity. Interleukin 1 receptor (IL-1R)-associated kinase-4 (IRAK-4) is required for various responses induced by IL-1R and Toll-like receptor signals. However, the molecular mechanism of IRAK-4 signaling and the role of its kinase activity have remained elusive. In this report, we demonstrate that IRAK-4 is recruited to the IL-1R complex upon IL-1 stimulation and is required for the recruitment of IRAK-1 and its subsequent activation/degradation. By reconstituting IRAK-4-deficient cells with wild type or kinase-inactive IRAK-4, we show that the kinase activity of IRAK-4 is required for the optimal transduction of IL-1-induced signals, including the activation of IRAK-1, NF-κB, and JNK, and the maximal induction of inflammatory cytokines. Interestingly, we also discover that the IRAK-4 kinase-inactive mutant is still capable of mediating some signals. These results suggest that IRAK-4 is an integral part of the IL-1R signaling cascade and is capable of transmitting signals both dependent on and independent of its kinase activity. Toll-like receptors (TLRs) 1The abbreviations used are: TLR, Toll-like receptor; IL, interleukin; IL-1R, IL-1 receptor; IRAK, IL-1R-associated kinase-4; JNK, c-Jun NH2-terminal kinase; TRAF, tumor necrosis factor receptor-associated factor 6; PBS, phosphate-buffered saline; EF, embryonic fibroblasts. are critical for receiving signals from molecular patterns associated with microbial pathogens to initiate innate immune responses (1Janeway Jr., C.A. Medzhitov R. Annu. Rev. Immunol. 2002; 20: 197-216Crossref PubMed Scopus (6190) Google Scholar, 2Akira S. Takeda K. Kaisho T. Nat. Immunol. 2001; 2: 675-680Crossref PubMed Scopus (3946) Google Scholar, 3Kimbrell D.A. Beutler B. Nat. Rev. Genet. 2001; 2: 256-267Crossref PubMed Scopus (495) Google Scholar). Interleukin 1 receptor (IL-1R) on the other hand is important for amplifying inflammatory responses triggered by microbial pathogens (4Dinarello C.A. Int. Rev. Immunol. 1998; 16: 457-499Crossref PubMed Scopus (675) Google Scholar). Interestingly, signal transduction pathways mediated by the IL-1R and TLRs are very similar (5O'Neill L.A. Curr. Top. Microbiol. Immunol. 2002; 270: 47-61Crossref PubMed Scopus (250) Google Scholar, 6Martin M.U. Wesche H. Biochim. Biophys. Acta. 2002; 1592: 265-280Crossref PubMed Scopus (342) Google Scholar). One common signaling pathway is initiated by the recruitment of the adaptor protein MyD88 to individual receptors upon ligand binding (7Akira S. Hoshino K. Kaisho T. J. Endotoxin Res. 2000; 6: 383-387Crossref PubMed Scopus (91) Google Scholar). MyD88 in turn is capable of recruiting IL-1 receptor-associated kinase (IRAK, also called IRAK-1) to the receptor complex (8Janssens S. Beyaert R. Martin M.U. Wesche H. Mol. Cell. 2003; 11: 293-302Abstract Full Text Full Text PDF PubMed Scopus (479) Google Scholar). Upon activation and modification, IRAK-1 dissociates from the receptor complex and associates with tumor necrosis factor receptor-associated factor 6 (TRAF6) to trigger downstream signaling pathways (9Cao Z. Xiong J. Takeuchi M. Kurama T. Goeddel D.V. Nature. 1996; 383: 443-446Crossref PubMed Scopus (1122) Google Scholar, 10Cao Z. Henzel W.J. Gao X. Science. 1996; 271: 1128-1131Crossref PubMed Scopus (773) Google Scholar, 11Takaesu G. Ninomiya-Tsuji J. Kishida S. Li X. Stark G.R. Matsumoto K. Mol. Cell. Biol. 2001; 21: 2475-2484Crossref PubMed Scopus (159) Google Scholar, 12Qian Y. Commane M. Ninomiya-Tsuji J. Matsumoto K. Li X. J. Biol. Chem. 2001; 276: 41661-41667Abstract Full Text Full Text PDF PubMed Scopus (177) Google Scholar), including the activation of NF-κB and various stress kinases such as c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK). The apparently simple cascade of MyD88→ IRAK→ TRAF6 became slightly more complex recently with a number of interesting discoveries. Starting from the top of the signaling cascade, three MyD88-like adaptor proteins, Mal (TIRAP), TRIF (TICAM-1), and TRAM (TICAM-2), were reported to play a role in mediating TLR signals (13O'Neill L.A. Fitzgerald K.A. Bowie A.G. Trends Immunol. 2003; 24: 286-290Abstract Full Text Full Text PDF PubMed Scopus (416) Google Scholar). Although MyD88 is important in mediating signals for most TLRs and IL-1R (7Akira S. Hoshino K. Kaisho T. J. Endotoxin Res. 2000; 6: 383-387Crossref PubMed Scopus (91) Google Scholar), Mal helps transmit MyD88-dependent signals induced by TLR2 and TLR4 (14Horng T. Barton G.M. Flavell R.A. Medzhitov R. Nature. 2002; 420: 329-333Crossref PubMed Scopus (685) Google Scholar, 15Yamamoto M. Sato S. Hemmi H. Sanjo H. Uematsu S. Kaisho T. Hoshino K. Takeuchi O. Kobayashi M. Fujita T. Takeda K. Akira S. Nature. 2002; 420: 324-329Crossref PubMed Scopus (817) Google Scholar). TRIF is critical in mediating TLR3 and TLR4 signals, particularly interferon-responsive pathways (through IRF-3) that are MyD88-independent (16Yamamoto M. Sato S. Hemmi H. Hoshino K. Kaisho T. Sanjo H. Takeuchi O. Sugiyama M. Okabe M. Takeda K. Akira S. Science. 2003; 301: 640-643Crossref PubMed Scopus (2510) Google Scholar, 17Hoebe K. Du X. Georgel P. Janssen E. Tabeta K. Kim S.O. Goode J. Lin P. Mann N. Mudd S. Crozat K. Sovath S. Han J. Beutler B. Nature. 2003; 424: 743-748Crossref PubMed Scopus (1034) Google Scholar), and TRAM associates with TLR4 specifically to signal along a TRIF-dependent pathway (18Yamamoto M. Sato S. Hemmi H. Uematsu S. Hoshino K. Kaisho T. Takeuchi O. Takeda K. Akira S. Nat. Immunol. 2003; 4: 1144-1150Crossref PubMed Scopus (827) Google Scholar, 19Fitzgerald K.A. Rowe D.C. Barnes B.J. Caffrey D.R. Visintin A. Latz E. Monks B. Pitha P.M. Golenbock D.T. J. Exp. Med. 2003; 198: 1043-1055Crossref PubMed Scopus (932) Google Scholar, 20Oshiumi H. Sasai M. Shida K. Fujita T. Matsumoto M. Seya T. J. Biol. Chem. 2003; 278: 49751-49762Abstract Full Text Full Text PDF PubMed Scopus (328) Google Scholar). Studies of IL-1R/TLR signals at the level of IRAKs have also made important progress over the past two years. There are a total of four IRAK family members: IRAK-1, IRAK-2, IRAK-M, and IRAK-4 (10Cao Z. Henzel W.J. Gao X. Science. 1996; 271: 1128-1131Crossref PubMed Scopus (773) Google Scholar, 21Muzio M. Ni J. Feng P. Dixit V.M. Science. 1997; 278: 1612-1615Crossref PubMed Scopus (984) Google Scholar, 22Wesche H. Gao X. Li X. Kirschning C.J. Stark G.R. Cao Z. J. Biol. Chem. 1999; 274: 19403-19410Abstract Full Text Full Text PDF PubMed Scopus (340) Google Scholar, 23Li S. Strelow A. Fontana E.J. Wesche H. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 5567-5572Crossref PubMed Scopus (543) Google Scholar). As deletion of the prototypical member IRAK-1 in mice reveals a partial defect in IL-1R/TLR signaling (24Kanakaraj P. Schafer P.H. Cavender D.E. Wu Y. Ngo K. Grealish P.F. Wadsworth S.A. Peterson P.A. Siekierka J.J. Harris C.A. Fung-Leung W.P. J. Exp. Med. 1998; 187: 2073-2079Crossref PubMed Scopus (177) Google Scholar, 25Thomas J.A. Allen J.L. Tsen M. Dubnicoff T. Danao J. Liao X.C. Cao Z. Wasserman S.A. J. Immunol. 1999; 163: 978-984PubMed Google Scholar), it was initially speculated that other IRAK proteins might compensate for the absence of IRAK-1. However, mice lacking IRAK-M exhibit enhanced inflammatory responses induced by IL-1 or TLR ligands, suggesting that IRAK-M may instead be a negative signal regulator (26Kobayashi K. Hernandez L.D. Galan J.E. Janeway Jr., C.A. Medzhitov R. Flavell R.A. Cell. 2002; 110: 191-202Abstract Full Text Full Text PDF PubMed Scopus (1150) Google Scholar). Furthermore, we have previously reported that, in IRAK-4-deficient mice, the signals mediated by IL-1R and most TLRs are severely impaired, indicating that IRAK-4 plays an essential non-redundant role in these signaling pathways (27Suzuki N. Suzuki S. Duncan G.S. Millar D.G. Wada T. Mirtsos C. Takada H. Wakeham A. Itie A. Li S. Penninger J.M. Wesche H. Ohashi P.S. Mak T.W. Yeh W.C. Nature. 2002; 416: 750-756Crossref PubMed Scopus (666) Google Scholar). IRAK-4 was shown to be primarily involved in MyD88-dependent pathways including signals triggered by IL-18 and LPS (28Suzuki N. Chen N.J. Millar D.G. Suzuki S. Horacek T. Hara H. Bouchard D. Nakanishi K. Penninger J.M. Ohashi P.S. Yeh W.C. J. Immunol. 2003; 170: 4031-4035Crossref PubMed Scopus (55) Google Scholar, 29Suzuki N. Suzuki S. Eriksson U. Hara H. Mirtosis C. Chen N.J. Wada T. Bouchard D. Hwang I. Takeda K. Fujita T. Der S. Penninger J.M. Akira S. Saito T. Yeh W.C. J. Immunol. 2003; 171: 6065-6071Crossref PubMed Scopus (36) Google Scholar), but the relationship between IRAK-4 and IRAK-1 and the requirement of the kinase activity of IRAK-4 in various signaling events remain to be addressed. In this report, we demonstrate that IRAK-4 is an integral component of the IL-1R signaling complex induced by IL-1 stimulation. The presence of IRAK-4 is required for the recruitment of IRAK-1 to the receptor complex and for the activation and subsequent degradation of IRAK-1 protein. Reconstitution of IRAK-4-deficient cells with wild type IRAK-4, but not kinase-inactive IRAK-4, is capable of restoring IL-1 responses. Interestingly, kinase-inactive IRAK-4 still supports partial cytokine production in IL-1-stimulated mutant cells. These results provide genetic evidence that IRAK-4 is required for IRAK-1 function and that IRAK-4 can transmit signals both through phosphorylation of downstream substrates and through protein-protein interactions that are independent of its kinase activity. Biological Reagents—Recombinant mouse IL-1β was purchased from R&D. Polyclonal rabbit antiserum to murine IRAK-4 was raised against a peptide corresponding to the COOH-terminal amino acids 439–459 of mouse IRAK-4 protein. Polyclonal rabbit antiserum to murine IRAK-1 was raised against a peptide corresponding to the COOH-terminal amino acids 692–712 of mouse IRAK-1 protein. Anti-mouse CD121a (IL-1 receptor, type I/p80) monoclonal antibody, anti-human JNK1 polyclonal antibody, and anti-IκB-α polyclonal antibody were purchased from BD Biosciences, Santa Cruz Biotechnology, and Cell Signaling, respectively. 3T3-like immortalized mouse embryonic fibroblasts were maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum. Cloning and Expression Vectors—Human wild type and kinase-inactive IRAK-4 (K213A/K214A) vectors were obtained from H. Wesche (Tularik Inc.). Murine IRAK-4 cDNA was obtained by PCR with a universal mouse cDNA library as template. Expression vectors for IRAK-4 were constructed by inserting PCR-generated cDNA fragments in the mammalian retroviral expression vector, pBabe-puro. Murine IRAK-4 (K213A/K214A) was constructed using the QuikChange site-directed mutagenesis kit (Stratagene). Cytokine Enzyme-linked Immunosorbent Assay—For detection of secreted IL-6 protein, MEFs (5 × 104/well) were incubated overnight in 24-well plates in 10% fetal bovine serum-Dulbecco's modified Eagle's medium. Cells were treated for 24 h with IL-1β (10 ng/ml) in fresh medium. Supernatants were then collected and analyzed for the presence of IL-6 using a commercial enzyme-linked immunosorbent assay kit (from BD Biosciences) using recombinant mouse IL-6 as a standard. Transfection-based Reporter Gene Assays—Cells were seeded at a density of 8 × 104/well in 6-well plates 24 h prior to transfection. For experiments with the NF-κB-dependent endothelial leukocyte adhesion molecule promoter, cells were transfected using the polycationic transfection reagent Superfect (Qiagen) with 0.2 μg of cytomegalovirus-lacZ/0.8 μg of pELAM-luciferase reporter and the indicated amounts of expression constructs. 42 h after transfection, cells were left untreated or stimulated with 10 ng/ml IL-1β for 6 h. Cells then were washed twice with PBS and lysed in 200 μl of reporter lysis buffer (Promega) at room temperature for 30 min. Luciferase activity in 20 μl of extract was measured immediately using the luciferase assay system (Promega) and a luminometer (Spectrafluor Plus, TECAN) according to the manufacturers' instructions. β-Galactosidase activity was measured in 20 μl of extract with 4 mg/ml ONPG (o-nitrophenyl-β-galactopyranoside, Sigma) in a 0.067 m of sodium phosphate buffer (pH 7.5). The optical density then was measured at 405 nm. Fold activation of reporter activity was calculated for each sample by dividing the luciferase activity in the experimental sample (normalized to β-galactosidase activity) by the luciferase activity in the unstimulated control. Stable Transfections—pBabe-puro (24Kanakaraj P. Schafer P.H. Cavender D.E. Wu Y. Ngo K. Grealish P.F. Wadsworth S.A. Peterson P.A. Siekierka J.J. Harris C.A. Fung-Leung W.P. J. Exp. Med. 1998; 187: 2073-2079Crossref PubMed Scopus (177) Google Scholar), a murine leukemia virus-based retroviral vector, was used to transduce the target genes. 16 h prior to transfection, the packaging cell line (Phoenix) was seeded at 5 × 106 cells/10-cm plate. Cells were grown to 60–70% confluency and transfected with empty pBabe-puro, pBabe-mIRAK-4, or pBabe-mIRAK-4 K213A/K214A vectors using a calcium-phosphate method. Viral supernatants were collected 48 h after transfection and then added to wild type or irak-4–/– cells. The infected cells were grown for 24 h and then subjected to selection in the same medium containing puromycin (2 μg/ml). Immunoprecipitation and Western Blot Analyses—Cells (2 × 106) were seeded on 10-cm plates and incubated with or 10 ng/ml IL-1β for the indicated of For the plates were immediately in an washed with PBS, and then in The cells were and the cell was lysed for 30 on in μl of lysis buffer containing 20 and a containing 1 10 and 10 The cells were for at in an and the was were with 20 of protein or Biosciences) for 30 at 4 on a were from by the of antibody and 30 μl of by for at 4 on a The were washed in lysis and the were in sample were on for IRAK-1 and IRAK-4 and 10% for and and to The were incubated with to or JNK, at in PBS were with or Biosciences) and proteins were with the system Biosciences) according to the instructions. were as previously (27Suzuki N. Suzuki S. Duncan G.S. Millar D.G. Wada T. Mirtsos C. Takada H. Wakeham A. Itie A. Li S. Penninger J.M. Wesche H. Ohashi P.S. Mak T.W. Yeh W.C. Nature. 2002; 416: 750-756Crossref PubMed Scopus (666) Google Scholar). × 106 cells were left untreated or stimulated with 10 ng/ml IL-1β for the indicated by the of were using the protein assay using bovine as the standard. 10 μg of extract was incubated with an and containing two The was in a total of 20 μl of binding buffer (5 1 μg of and 10% for 20 at room were on a and complex was by proteins in total cell were at 4 using polyclonal antibody Santa Cruz activity in the was using as a in the presence of The of total protein in the was by Western using the polyclonal IRAK-4 to the IL-1R the role of IRAK-4 in IL-1 three independent of IRAK-4-deficient embryonic fibroblasts and wild type were As shown in the defect in IL-6 production was in of the These independent were in the experiments in this to the of have previously that signals downstream of MyD88 on IRAK-4, of TRAF6 is capable of NF-κB independent of These results suggest that IRAK-4 in the signaling cascade between MyD88 and TRAF6 in a similar as IRAK-1 (5O'Neill L.A. Curr. Top. Microbiol. Immunol. 2002; 270: 47-61Crossref PubMed Scopus (250) Google Scholar, S. Beyaert R. Martin M.U. Wesche H. Mol. Cell. 2003; 11: 293-302Abstract Full Text Full Text PDF PubMed Scopus (479) Google Scholar). IRAK-4 is recruited to the receptor complex as MyD88 and IRAK-1. As shown in as as 1 after IL-1 IRAK-4 is recruited to the IL-1R-associated Interestingly, IRAK-1 that dissociates from the receptor complex and is M.U. Wesche H. Biochim. Biophys. Acta. 2002; 1592: 265-280Crossref PubMed Scopus (342) Google Scholar), IRAK-4 with IL-1R for to after stimulation and the total protein level of IRAK-4 not to this In a of IL-1R-associated IRAK-4 was at after IL-1 stimulation the of this IRAK-4 IRAK-1 and IRAK-1 shown to with IRAK-4 and after IL-1 stimulation S. Strelow A. Fontana E.J. Wesche H. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 5567-5572Crossref PubMed Scopus (543) Google Scholar), we the absence of IRAK-4 the function of IRAK-1 in IL-1 the recruitment of IRAK-1 to the IL-1R complex in to IL-1 is severely in cells lacking IRAK-4 to its recruitment to the IL-1R complex in wild type IRAK-1 in and then in K. S. B. N. Beyaert R. J. J. Exp. Med. 2003; PubMed Scopus Google Scholar). However, in the absence of IRAK-4, IRAK-1 not activation or degradation IL-1 These results suggest that IRAK-4 is essential for the recruitment and activation of IRAK-1. is interesting to that the level of IRAK-1 expression is enhanced in IRAK-4-deficient cells with wild type this not in responses. The of IRAK-4 on IL-1 IRAK-4 can function as a kinase and downstream substrates such as IRAK-1 S. Strelow A. Fontana E.J. Wesche H. Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 5567-5572Crossref PubMed Scopus (543) Google Scholar, C. D. J. Cao P. Li S. Wesche H. Martin M.U. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar), it is that the kinase activity of IRAK-4 plays a role in IL-1 signal of wild type IRAK-4, but not kinase-inactive IRAK-4, in three independent IRAK-4-deficient cell was to the IL-1-induced as measured by NF-κB-dependent reporter is shown in The of the NF-κB signal was to the of wild type IRAK-4 In of kinase-inactive IRAK-4 expression in severely IL-1 responses with wild type the requirement of IRAK-4 kinase activity for IL-1R we of IRAK-4 cells wild type IRAK-4, or kinase-inactive type cells were as the for IL-1 responses. we measured IL-1-induced cytokine production and that mutant cells with wild type IRAK-4 were capable of IL-6 to the same level as the wild type standard. the other mutant cells with kinase-inactive IRAK-4 to of the cytokine that cells with wild type IRAK-4 The level of IL-6 production induced by the kinase-inactive IRAK-4, severely was IRAK-4 mutant cells with of the of a partial cytokine mediated by kinase-inactive IRAK-4 by transfection experiments on three IRAK-4 cell not also wild type and kinase-inactive IRAK-4 using transfected cells as as and similar results were obtained not protein expression of wild type and kinase-inactive IRAK-4 in were and with similar expression were for these experiments we also wild type and kinase-inactive IRAK-4 to mouse cells and kinase-inactive IRAK-4 the cytokine induced by IL-1 NF-κB and activation have reported to play a role in IL-6 production G. K. E. G. 2000; PubMed Scopus Google Scholar, D. Z. Flavell R.A. T. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar). we these signaling events in IRAK-4 cells with wild type or kinase-inactive As with wild type IRAK-4 the of mutant cells with kinase-inactive IRAK-4 in severely NF-κB and activation induced by IL-1 and These results suggest that the kinase activity of IRAK-4 plays an important role in IL-1 signals to NF-κB and also the kinase activity of IRAK-4 plays a role in the activation of IRAK-1 in to with wild type IRAK-4 of mutant with kinase-inactive IRAK-4 was of restoring IRAK-1 degradation is dependent on its after IL-1 stimulation The common signaling cascade of MyD88→ IRAK→ TRAF6 is for mediating signals induced by the IL-1R and most TLR family the of TLR3 and S. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar). of this signaling pathway is have for and inflammatory In this report, we the function of IRAK-4 and its with IRAK-1, both kinases that play important in IL-1R and TLR signaling demonstrate that IRAK-4 is essential for signals at by in the recruitment of IRAK-1 to the receptor complex and in the activation of IRAK-1. The kinase activity of IRAK-4 is required for mediating IL-1-induced NF-κB and activation and for the optimal induction of inflammatory cytokines. In cells with kinase-inactive IRAK-4, the IRAK-4 for signaling complex activity but can still signal with other signaling these interactions are capable of a partial cytokine to IL-1 stimulation. The signals for this partial are still One may in the kinase activity of IRAK-1 M.U. Wesche H. Biochim. Biophys. Acta. 2002; 1592: 265-280Crossref PubMed Scopus (342) Google Scholar, C. D. J. Cao P. Li S. Wesche H. Martin M.U. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). of IRAK-1 may be at the receptor or may function in the in to downstream signals. it is interesting to that signaling other IRAK-1 may signals independent of IRAK-4 kinase activity. These may more on protein interactions and to of proteins provide important to this Upon IL-1 IRAK-4 with the IL-1R for a of in IRAK-1 associates with the receptor in cells. IRAK-4 can transmit signals from the receptor complex IRAK-1 is to be addressed. is that are IRAK-4 substrates other IRAK-1 that can IL-1 signals. the of IRAK-4 in the IL-1R complex may as a negative signal at to inflammatory responses. with the the of IRAK-4 shown to IL-1 signals in cell A. C. S. M. J. Exp. Med. 2003; 198: PubMed Scopus Google Scholar). The relationship between IRAK-4 and two other IRAK family and IRAK-M, is that or IRAK-M can with IRAK-4 and may as substrates for these two kinase-inactive IRAK family might to signaling in a IRAK-1 or as negative in the IRAK-M are interesting to mutant mice and cells lacking individual or of IRAK to the molecular IRAK results the requirement for the kinase activity of IRAK-4 in IL-1R signaling are from by J. Z. Y. J.L. Li X. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). from using kinase-inactive IRAK-4 to a cell line from a IRAK-4-deficient C. A. M. J. K. C. S. J. C. C. R. D. G. A. H. S. S. R. A. H. S. R.A. A. J.L. Science. 2003; PubMed Scopus Google that the kinase activity of IRAK-4 is for its signaling The for this are but are three in experimental mouse embryonic fibroblasts and the of cell is that three the requirement for the kinase activity of IRAK-4 between it be very interesting from the of the of signaling as as these Wesche for IRAK-4 expression and for are also to and for and to for

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