Scientific Report 2003-2004 - Cleveland Clinic Lerner Research ...

The Department of ImmunologyGenetic Analysis of IL-1 Signaling andRegulation of NFκB ActivationAprimary interest in my laboratory is thestudy of interleukin-1 (IL-1)-mediatedsignaling. IL-1 is a major pro-inflammatorycytokine with a wide range of biological activitiesin inflammation. It functionsmainly by activating proteinkinase cascades, leading to theactivation of nuclear factorκB(NFκB) and Jun kinase,which in turn phosphorylatesand activates activationtranscription factor (ATF)and activator protein-1 (AP-1). The IL-1 receptor belongsto the IL-1 receptor/Toll-likereceptor (IL-1R/TLR)superfamily. Human TLRshave recently emerged as keycomponents in the generationof immune and inflammatoryresponses due to their abilityto recognize pathogen-associatedmolecules. Tremendousefforts have beendevoted to understanding themolecular mechanisms bywhich IL-1Rs/TLRs mediatesignaling, with the long-termobjective of developing moreeffective anti-inflammatory small-molecule drugs.We have previously taken a genetic approachto studying IL-1-dependent signaling pathways,using random mutagenesis to generate IL-1-unresponsive cell lines lacking specific componentsof the pathways. Employing the siRNA interferenceapproach in cultured human cells, we havealso successfully abolished the expression ofsignaling components of the pathway. The resultingmutant cell lines have provided us with a uniqueopportunity to study the IL-1R-mediated signalingpathway. We are now focusing on the key stepsthat govern the receptor proximal signaling eventsof the IL-1 pathway, including the IL-1-inducedformation and dissociation of the receptor complexand the translocation and activation of transforminggrowth factor-β-activated kinase 1 in responseto IL-1 stimulation.We recently identified a novel member ofthe IL-1R/TLR superfamily, named Sigirr. Sigirr -/-mice show an enhanced inflammatory state uponstimulation with IL-1 and various Toll ligands,including lipopolysaccharide, CpG DNA anddsRNA, suggesting that Sigirr might be a negativeregulator for the IL-1R/TLR-mediated pathway.Upon stimulation with IL-1, Sigirr interacts withIL-1 receptor, IL-1R-associated kinase and TNFreceptor-associated factor-6, suggesting that Sigirrprobably functions through its interaction with thereceptor complex. We are investigating how Sigirrnegatively regulates the IL-1 pathway via itsXiaoxia Li, Ph.D.interaction with the IL-1 receptor complex.We are also interested in the signalingpathways mediated by the TNF receptor (TNFR)superfamily, whose members exert many importantbiological functions,including cell proliferation/differentiation, inflammation,immune responses, andhomeostasis. These TNFRfamily members functionmainly by activating proteinkinases cascades, leading tothe activation of transcriptionfactors, includingNFκB, ATF and AP-1.Although much effort hasbeen made toward understandingthe molecularmechanisms of signaltransduction mediated bythese receptors, manyquestions still remain. Werecently identified a noveladapter molecule, NFκBactivator 1 (Act1), thatfunctions in signalingpathways mediated by asubset of TNFR familymembers, including CD40and lymphotoxin-β receptor. We are particularlyinterested in the CD40-mediated pathway, since itis not only essential for normal immune responses,but is also associated with several pathologicalconditions of autoimmune and chronic inflammatorydiseases. We have generated Act1 -/- mice, anessential tool to investigate the physiologicalfunction of Act1.THE LI LABORATORYPOSTDOCTORAL FELLOWSZhenfang Jiang, Ph.D.Youcun Qian, Ph.D.TECHNICAL ASSISTANTGrace ChiGRADUATE STUDENTSJinzhong QinDave WaldCOLLABORATORSTim Bird, Ph.D. 1Robert Fairchild, Ph.D. 2Peter Heeger, Ph.D. 2Brian L. Kotzin, M.D. 3Kunihiro Mastumoto, M.D. 4Robert Rickert, Ph.D. 5George R. Stark, Ph.D. 6Paul D. Wightman, Ph.D. 71Immunex Corp., Seattle, WA2Dept. of Immunology, CCF3Depts. of Med. and Immunol.,Univ. of Colorado Health Sci.Ctr., Denver, CO4Dept. of Molec. Biology,Nagoya Univ., Nagoya, Japan5Div. of Biology and UCSDCancer Ctr., Univ. of California/SanDiego, La Jolla, CA6Dept. of Molec. Biology, CCF73M Pharmaceuticals, St.Paul, MNQian, Y., Zhao, Z., Jiang, Z., and X. Li (2002) Role of NFκB activator Act1 in CD40-mediated signaling in epithelial cells. Proc. Natl. Acad Sci. USA 99:9386-9391.Jiang, Z., Ninomiya-Tsuji, J., Qian, Y., Matsumoto, K., and X. Li (2002) Interleukin-1 (IL-1)receptor-associated kinase-dependent IL-1-induced signaling complexes phosphorylateTAK1 and TAB2 at the plasma membrane and activate TAK1 in the cytosol. Mol. Cell. Biol.22:7158-7167.Jiang, Z., Johnson, H.J., Nie, H., Qin, J., Bird, T.A., and X. Li (2003) Pellino 1 is requiredfor interleukin-1 (IL-1)-mediated signaling through its interaction with the IL-1 receptorassociatedkinase 4 (IRAK4)-IRAK-tumor necrosis factor receptor-associated factor 6(TRAF6) complex. J. Biol. Chem. 278:10952-10956.Jiang, Z., Zamanian-Daryoush, M., Nie, H., Silva, A.M., Williams, B.R., and X. Li (2003)Poly(dI x dC)-induced Toll-like receptor 3 (TLR3)-mediated activation of NFκB and MAPkinase is through an interleukin-1 receptor-associated kinase (IRAK)-independent pathwayemploying the signaling components TLR3-TRAF6-TAK1-TAB2-PKR. J. Biol. Chem.278:16713-16719.Zhao, Z., Qian, Y., Wald, D., Xia, Y.F., Geng, J.G., and X. Li (2003) IFN regulatory factor-1 is required for the up-regulation of the CD40-NF-κB activator 1 axis during airwayinflammation. J. Immunol. 170:5674-5680.95