Activation of stress-activated protein kinase/c-Jun N-terminal kinase, but not NF-kappaB, by the tumor necrosis factor (TNF) receptor 1 through a TNF receptor-associated factor 2- and germinal center kinase related-dependent pathway

A key step by which tumor necrosis factor (TNF) signals the activation of nuclear factor-kappaB (NF-kappaB) and the stress-activated protein kinase (SAPK, also called c-Jun N-terminal kinase or JNK) is the recruitment to the TNF receptor of TNF receptor-associated factor 2 (TRAF2). However, the subsequent steps in TRAF2-induced SAPK and NF-kappaB activation remain unresolved. Here we report the identification of a TNF-responsive serine/threonine protein kinase termed GCK related (GCKR) that likely signals via mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase kinase 1 (MEKK1) to activate the SAPK pathway. TNF, TRAF2, and ultraviolet (UV) light, which in part uses the TNF receptor signaling pathway, all increased GCKR activity. A TRAF2 mutant, which inhibits both TRAF2-induced NF-kappaB and SAPK activation, blocked TNF-induced GCKR activation. Finally, interference with GCKR expression impeded TRAF2- and TNF-induced SAPK activation but not that of NF-kappaB. This suggests a divergence in the TNF signaling pathway that leads to SAPK and NF-kappaB activation, which is located downstream of TRAF2 but upstream of GCKR.     

Binding of interleukin-18 to the interleukin-1 receptor homologous receptor IL-1Rrp1 leads to activation of signaling pathways similar to those used by interleukin-1

Interleukin-18 (IL-18) is an inflammatory cytokine that has been shown to enhance a variety of Th1 type T cell responses. Because IL-18 is homologous to IL-1, we tested binding of IL-18 to the known IL-1R family members. We could show binding of IL-18 to the orphan receptor IL-1Rrp1 but not to other IL-1R homologous proteins. IL-1Rrp1 and IL-1RI share highly conserved domains within their cytoplasmic regions. Comparison of the IL-1 and IL-18 signaling mechanisms showed that they activate identical cytoplasmic messengers. IL-18, like IL-1, induced association of its receptor with IRAK and subsequent recruitment of TRAF6. IL-18 activated p38 MAP kinase, jun kinase, and beta casein kinase (TIP kinase), an apparently novel kinase previously thought to be specifically activated by IL-1 and tumor necrosis factor (TNF). IL-18 activated NF-kappaB in EL4/6.1 thymoma cells but not in COS-7 cells, even though the latter presumably contain all components required for the IL-1 signaling pathway. From our binding and signaling studies, we conclude that the IL-18 receptor complex consists of IL-18, the IL-1Rrp1, and another thus far unidentified receptor molecule.     

Overview of interleukin-18: more than an interferon-gamma inducing factor

Initially described in 1989 as interferon-gamma (IFN-gamma) inducing factor (IGIF), interleukin-18 (IL-18) is a novel pro-inflammatory cytokine that is clearly more than an inducer of IFN-gamma. The cytokine possesses several biological properties such as activation of nuclear factor-kappaB (NF-kappaB), Fas ligand expression, the induction of both CC and CXC chemokines, and increased production of competent human immunodeficiency virus. Most activities are due to a receptor complex that recruits the IL-1 receptor-activating kinase (IRAK), leading to translocation of NF-kappaB. This property and others support the concept that IL-18 is related to the IL-1 family. Indeed, one of the IL-18 receptor chains is the IL-1 receptor-related protein, a member of the IL-1R family. In addition, IL-18 is structurally similar to IL-1beta and like IL-1beta is first synthesized as a leaderless precursor requiring the IL-1beta converting enzyme for cleavage into an active molecule. The biology of IL-18 is reviewed in the overview and the implication for a role for this cytokine in disease is presented.     

Role of nuclear factor-kappaB activation in cytokine- and sphingomyelinase-stimulated inducible nitric oxide synthase gene expression in vascular smooth muscle cells

Inflammatory cytokines, such as interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF alpha), are known to activate sphingomyelinase (SMase) and nuclear factor-kappaB (NF-kappaB) in certain cell types, which also stimulate inducible nitric oxide synthase (iNOS) gene in vascular smooth muscle cells (VSMCs). However, it remains unknown whether the SMase pathway is involved in iNOS gene expression in VSMCs. Therefore, the present study was designed to examine whether SMase induces iNOS gene expression via the NF-kappaB activation pathway similar to that of IL-1beta and TNF alpha in cultured rat VSMCs. Neutral SMase, although less potently than IL-1beta and TNF alpha, stimulated nitrite/nitrate (NOx) production, and iNOS messenger RNA and protein expression, as assessed by Northern and Western blot analyses, respectively. Neutral SMase, IL-1beta, and TNF alpha activated NF-kappaB, as revealed by electrophoretic mobility shift assay, and its nuclear translocation, as demonstrated by immunocytochemical study. Neutral SMase potentiated NOx production, iNOS expression, and NF-kappaB activation stimulated by TNF alpha, but not by IL-1beta. Aldehyde peptide proteasome inhibitors completely blocked NOx production, iNOS expression, NF-kappaB activation, and its nuclear translocation induced by cytokines and neutral SMase. IL-1beta and TNF alpha, but not neutral SMase, caused a transient decrease in IkappaB-alpha protein levels, whereas IkappaB-beta protein expression was not affected by either agent. Proteasome inhibitors prevented cytokine-mediated IkappaB-alpha degradation. Several cell-permeable ceramide analogs (C2, C6, and C8), hydrolysis products of sphingomyelin, activated NF-kappaB less potently than neutral SMase, but had no effect on NOx production. These results demonstrate an essential role of NF-kappaB activation in mediation of neutral SMase-induced iNOS expression, but distinct from the proteasome-mediated IkappaB-alpha degradation by cytokines, suggesting the possible involvement of an additional signaling pathway(s).     

Signal transduction by the IL-1 type I receptor: evidence for the involvement of a receptor-coupled protein kinase

A novel serine/threonine specific protein kinase was found to be associated with the type I IL-1 receptor in the murine T cell lines D10N and EL-4. This kinase was identified in immunoprecipitates from IL-1 stimulated T-cells by its ability to phosphorylate exogenous substrates in the presence of radiolabeled ATP. An endogenous protein, most likely a member of the IL-1 R1 complex, was also phosphorylated. The activation of the kinase is specific for IL-1, neither TNF nor phorbol esters were able to activate the IL-1 RI associated kinase activity. The IL-1 receptor antagonist had no intrinsic activity and inhibited the activation of the kinase. The activation of the kinase was rapid and detectable after 30 seconds of IL-1 stimulation. A minimal model of the IL-RI signal transduction complex is discussed, presenting this novel serine/threonine kinase as a constituent of the complex.     

Differential regulation of IkappaB kinase alpha and beta by two upstream kinases, NF-kappaB-inducing kinase and mitogen-activated protein kinase/ERK kinase kinase-1

NF-kappaB is activated by various stimuli including inflammatory cytokines and stresses. A key step in the activation of NF-kappaB is the phosphorylation of its inhibitors, IkappaBs, by an IkappaB kinase (IKK) complex. Recently, two closely related kinases, designated IKKalpha and IKKbeta, have been identified to be the components of the IKK complex that phosphorylate critical serine residues of IkappaBs for degradation. A previously identified NF-kappaB-inducing kinase (NIK), which mediates NF-kappaB activation by TNFalpha and IL-1, has been demonstrated to activate IKKalpha. Previous studies showed that mitogen-activated protein kinase/ERK kinase kinase-1 (MEKK1), which constitutes the c-Jun N-terminal kinase/stress-activated protein kinase pathway, also activates NF-kappaB by an undefined mechanism. Here, we show that overexpression of MEKK1 preferentially stimulates the kinase activity of IKKbeta, which resulted in phosphorylation of IkappaBs. Moreover, a catalytically inactive mutant of IKKbeta blocked the MEKK1-induced NF-kappaB activation. By contrast, overexpression of NIK stimulates kinase activities of both IKKalpha and IKKbeta comparably, suggesting a qualitative difference between NIK- and MEKK1-mediated NF-kappaB activation pathways. Collectively, these results indicate that NIK and MEKK1 independently activate the IKK complex and that the kinase activities of IKKalpha and IKKbeta are differentially regulated by two upstream kinases, NIK and MEKK1, which are responsive to distinct stimuli.     

Interleukin-4 down-regulates both forms of tumor necrosis factor receptor and receptor-mediated apoptosis, NF-kappaB, AP-1, and c-Jun N-terminal kinase. Comparison with interleukin-13

The activity of tumor necrosis factor (TNF), a proinflammatory cytokine, is regulated by a number of other cytokines, including interleukin (IL)-4. How IL-4 regulates various activities of TNF is not fully understood. In the present report, we investigated the effect of IL-4 on the cell surface TNF receptors in human histiocytic lymphoma U-937 cells. Pretreatment of cells with IL-4 down-regulated TNF receptors in a dose- and time-dependent manner; an almost 90% decrease occurred with 10 ng/ml IL-4 treatment for 24 h. Scatchard analysis revealed that the decrease was due to receptor number and not affinity. IL-13, which shares a common receptor subunit and various biological activities with IL-4, had no effect on TNF receptors. IL-4's effect on TNF receptors was not cell type-specific, since decreases also occurred on various epithelial and T cells. Both the p60 and p80 forms of the TNF receptor were down-regulated to the same extent. Western blot showed that IL-4 induced shedding of the TNF receptors. The decrease of TNF receptors by IL-4 was accompanied by down-regulation of TNF-induced activities, including cytotoxicity, caspase-3 activation, NF-kappaB and AP-1 activation, and c-Jun N-terminal kinase induction. Wortmannin reversed the IL-4-induced TNF receptor down-regulation and all other measured cellular responses, indicating a critical role of phosphatidylinositol 3-kinase. Rapamycin also blocked the effect of IL-4-induced regulation, thus suggesting the role of p70 S6 kinase. Overall, our results suggest that TNF receptor down-regulation by IL-4 plays a critical role in the antagonistic effects of IL-4 on TNF-induced cellular responses and that this mechanism differs from that of IL-13.     

RIP2 is a novel NF-kappaB-activating and cell death-inducing kinase

Through specific interactions with members of the tumor necrosis receptor (TNFR) family, adapter molecules such as the serine/threonine (Ser/Thr) kinase RIP mediate divergent signaling pathways including NF-kappaB activation and cell death. In this study, we have identified and characterized a novel 61-kDa protein kinase related to RIP that is a component of both the TNFR-1 and the CD40 signaling complexes. Receptor interacting protein-2 (RIP2) contains an N-terminal domain with homology to Ser/Thr kinases and a C-terminal caspase activation and recruitment domain (CARD), a homophilic interaction motif that mediates the recruitment of caspase death proteases. Overexpression of RIP2 signaled both NF-kappaB activation and cell death. Mutational analysis revealed the pro-apoptotic function of RIP2 to be restricted to its C-terminal CARD domain, whereas the intact molecule was necessary for NF-kappaB activation. RIP2 interacted with other members of the TNFR-1 signaling complex, including inhibitor of apoptosis protein cIAP1 and with members of the TNFR-associated factor (TRAF) family, specifically TRAF1, TRAF5, and TRAF6, but not with TRAF2, TRAF3, or TRAF4. These TRAF interactions mediate the recruitment of RIP2 to receptor signaling complexes.     

CD27, a member of the tumor necrosis factor receptor superfamily, activates NF-kappaB and stress-activated protein kinase/c-Jun N-terminal kinase via TRAF2, TRAF5, and NF-kappaB-inducing kinase

CD27 is a member of the tumor necrosis factor (TNF) receptor superfamily and is expressed on T, B, and NK cells. The signal via CD27 plays pivotal roles in T-T and T-B cell interactions. Here we demonstrate that overexpression of CD27 activates NF-kappaB and stress-activated protein kinase (SAPK)/c-Jun N-terminal kinase (JNK). Deletion analysis of the cytoplasmic domain of CD27 revealed that the C-terminal PIQEDYR motif was indispensable for both NF-kappaB and SAPK/JNK activation and was also required for the interaction with TNF receptor-associated factor (TRAF) 2 and TRAF5, both of which have been implicated in NF-kappaB activation by members of the TNF-R superfamily. Co-transfection of a dominant negative TRAF2 or TRAF5 blocked NF-kappaB and SAPK/JNK activation induced by CD27. Recently, a TRAF2-interacting kinase has been identified, termed NF-kappaB-inducing kinase (NIK). A kinase-inactive mutant NIK blocked CD27-, TRAF2-, and TRAF5-mediated NF-kappaB and SAPK/JNK activation. These results indicate that TRAF2 and TRAF5 are involved in NF-kappaB and SAPK/JNK activation by CD27, and NIK is a common downstream kinase of TRAF2 and TRAF5 for NF-kappaB and SAPK/JNK activation.