Key Spec Table
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H, R, M||WB||M||Purified||Monoclonal Antibody|
|Application||Anti-IKKβ Antibody, clone 10AG2 detects level of IKKβ & has been published & validated for use in WB.|
|Safety Information according to GHS|
|Storage and Shipping Information|
|Storage Conditions||Stable for 1 year at -20ºC from date of receipt.|
|Material Size||100 µg|
|Anti-IKK#946;, clone 10AG2 - DAM1749919||DAM1749919|
|Anti-IKK#946;, clone 10AG2 - 2137999||2137999|
|Anti-IKK, clone 10AG2 - 2064509||2064509|
|Anti-IKK, clone 10AG2 - 2203116||2203116|
|Anti-IKK, clone 10AG2 - 2290926||2290926|
|Anti-IKK, clone 10AG2 - DAM1442517||DAM1442517|
|Anti-IKK, clone 10AG2 - DAM1734752||DAM1734752|
|Anti-IKK, clone 10AG2 - DAM1774677||DAM1774677|
|Anti-IKK, clone 10AG2 - JBC1872234||JBC1872234|
|Anti-IKKBeta, clone 10AG2 - DAM1543628||DAM1543628|
|Reference overview||Application||Species||Pub Med ID|
|IKK is a therapeutic target in KRAS-Induced lung cancer with disrupted p53 activity.|
Bassères, DS; Ebbs, A; Cogswell, PC; Baldwin, AS
Genes & cancer 5 41-55 2014
Activating mutations in KRAS are prevalent in cancer, but therapies targeted to oncogenic RAS have been ineffective to date. These results argue that targeting downstream effectors of RAS will be an alternative route for blocking RAS-driven oncogenic pathways. We and others have shown that oncogenic RAS activates the NF-κB transcription factor pathway and that KRAS-induced lung tumorigenesis is suppressed by expression of a degradation-resistant form of the IκBα inhibitor or by genetic deletion of IKKβ or the RELA/p65 subunit of NF-κB. Here, genetic and pharmacological approaches were utilized to inactivate IKK in human primary lung epithelial cells transformed by KRAS, as well as KRAS mutant lung cancer cell lines. Administration of the highly specific IKKβ inhibitor Compound A (CmpdA) led to NF-κB inhibition in different KRAS mutant lung cells and siRNA-mediated knockdown of IKKα or IKKβ reduced activity of the NF-κB canonical pathway. Next, we determined that both IKKα and IKKβ contribute to oncogenic properties of KRAS mutant lung cells, particularly when p53 activity is disrupted. Based on these results, CmpdA was tested for potential therapeutic intervention in the Kras-induced lung cancer mouse model (LSL-Kras (G12D)) combined with loss of p53 (LSL-Kras (G12D)/p53 (fl/fl)). CmpdA treatment was well tolerated and mice treated with this IKKβ inhibitor presented smaller and lower grade tumors than mice treated with placebo. Additionally, IKKβ inhibition reduced inflammation and angiogenesis. These results support the concept of targeting IKK as a therapeutic approach for oncogenic RAS-driven tumors with altered p53 activity.
|Intestinal tumorigenesis initiated by dedifferentiation and acquisition of stem-cell-like properties.|
Schwitalla, S; Fingerle, AA; Cammareri, P; Nebelsiek, T; Göktuna, SI; Ziegler, PK; Canli, O; Heijmans, J; Huels, DJ; Moreaux, G; Rupec, RA; Gerhard, M; Schmid, R; Barker, N; Clevers, H; Lang, R; Neumann, J; Kirchner, T; Taketo, MM; van den Brink, GR; Sansom, OJ; Arkan, MC; Greten, FR
Cell 152 25-38 2013
Cell-type plasticity within a tumor has recently been suggested to cause a bidirectional conversion between tumor-initiating stem cells and nonstem cells triggered by an inflammatory stroma. NF-κB represents a key transcription factor within the inflammatory tumor microenvironment. However, NF-κB's function in tumor-initiating cells has not been examined yet. Using a genetic model of intestinal epithelial cell (IEC)-restricted constitutive Wnt-activation, which comprises the most common event in the initiation of colon cancer, we demonstrate that NF-κB modulates Wnt signaling and show that IEC-specific ablation of RelA/p65 retards crypt stem cell expansion. In contrast, elevated NF-κB signaling enhances Wnt activation and induces dedifferentiation of nonstem cells that acquire tumor-initiating capacity. Thus, our data support the concept of bidirectional conversion and highlight the importance of inflammatory signaling for dedifferentiation and generation of tumor-initiating cells in vivo.
|Transcription of Tnfaip3 is regulated by NF-κB and p38 via C/EBPβ in activated macrophages.|
Lai, TY; Wu, SD; Tsai, MH; Chuang, EY; Chuang, LL; Hsu, LC; Lai, LC
PloS one 8 e73153 2013
Macrophages play a pivotal role in the immune system through recognition and elimination of microbial pathogens. Toll-like receptors (TLRs) on macrophages interact with microbial substances and initiate signal transduction through intracellular adapters. TLR4, which recognizes the lipopolysaccharides (LPS) on Gram-positive and Gram-negative bacteria, triggers downstream signaling mediators and eventually activates IκB kinase (IKK) complex and mitogen-activated protein kinases (MAPKs) such as p38. Previous reports revealed that, in addition to NF-κB, a core transcription factor of the innate immune response, the induction of some LPS-induced genes in macrophages required another transcription factor whose activity depends on p38. However, these additional transcription factors remain to be identified. In order to identify p38-activated transcription factors that cooperate with NF-κB in response to LPS stimulation, microarrays were used to identify genes regulated by both NF-κB and p38 using wild-type, IKK-depleted, and p38 inhibitor-treated mouse bone marrow-derived macrophages (BMDMs). In silico analysis of transcription factor binding sites was used to predict the potential synergistic transcription factors from the co-expressed genes. Among these genes, NF-κB and C/EBPβ, a p38 downstream transcription factor, were predicted to co-regulate genes in LPS-stimulated BMDMs. Based on the subsequent results of a chromatin immunoprecipitation assay and TNFAIP3 expression in C/EBPβ-ablated macrophages, we demonstrated that Tnfaip3 is regulated by both NF-κB and p38-dependent C/EBPβ. These results identify a novel regulatory mechanism in TLR4-mediated innate immunity.
|IκB kinase β is required for activation of NF-κB and AP-1 in CD3/CD28-stimulated primary CD4(+) T cells.|
Lupino, E; Ramondetti, C; Piccinini, M
Journal of immunology (Baltimore, Md. : 1950) 188 2545-55 2012
Engagement of the TCR and CD28 coreceptor by their respective ligands activates signal transduction cascades that ultimately lead to the activation of the transcription factors NFAT, AP-1, and NF-κB, which are required for the expression of cytokines and T cell clonal expansion. Previous studies have demonstrated that in mature T cells, activation of AP-1 and NF-κB is dependent on protein kinase C θ, suggesting the existence of a common signaling pathway. In this study, we show that in human primary CD4(+) T cells, exposure to the cell-permeable IKKβ inhibitor PS-1145 or genetic ablation of IKKβ abrogates cell proliferation and impairs the activation of NF-κB and AP-1 transcription factors in response to engagement of CD3 and CD28 coreceptor. In addition, we show that stimulation of T cells in the absence of IKKβ activity promotes the time-dependent and cyclosporine-sensitive expression of negative regulators of T cell signaling leading to a hyporesponsive state of T cells.
|zVAD-induced necroptosis in L929 cells depends on autocrine production of TNFα mediated by the PKC-MAPKs-AP-1 pathway.|
Wu, YT; Tan, HL; Huang, Q; Sun, XJ; Zhu, X; Shen, HM
Cell death and differentiation 18 26-37 2011
It is intriguing that some pan-caspase inhibitors such as zVAD-fmk (zVAD) are capable of inducing necrotic cell death in a selected group of cells. As earlier reports from our laboratory have ruled out the original notion that zVAD-induced necrosis in mouse fibrosarcoma L929 cells was autophagic cell death, the main objective of this study was thus to determine the underlying mechanism of this form of cell death. In this study, we provided clear evidence that zVAD-induced necroptosis in L929 cells and such cell death is dependent on autocrine production of tumor necrosis factor-α (TNFα) at the transcriptional level. More importantly, we identified that activating protein-1 (AP-1), but not nuclear factor κ-B, is the transcription factor controlling zVAD-induced TNFα transcription. Moreover, zVAD is able to activate AP-1 through activation of two upstream mitogen-activated kinases (MAPKs), c-Jun N-terminal kinase and extracellular signal-regulated kinase. Finally, we found that protein kinase C is the important upstream signaling molecule in mediating zVAD-induced activation of MAPKs and AP-1, and subsequent autocrine production of TNFα and cell death. Data from this study reveal the molecular mechanisms underlying zVAD-induced necroptosis, an important form of programmed necrotic cell death with increasing understanding of its biological significance in health and diseases.
|Crystal structure of inhibitor of κB kinase β.|
Xu, G; Lo, YC; Li, Q; Napolitano, G; Wu, X; Jiang, X; Dreano, M; Karin, M; Wu, H
Nature 472 325-30 2011
Inhibitor of κB (IκB) kinase (IKK) phosphorylates IκB proteins, leading to their degradation and the liberation of nuclear factor κB for gene transcription. Here we report the crystal structure of IKKβ in complex with an inhibitor, at a resolution of 3.6 Å. The structure reveals a trimodular architecture comprising the kinase domain, a ubiquitin-like domain (ULD) and an elongated, α-helical scaffold/dimerization domain (SDD). Unexpectedly, the predicted leucine zipper and helix-loop-helix motifs do not form these structures but are part of the SDD. The ULD and SDD mediate a critical interaction with IκBα that restricts substrate specificity, and the ULD is also required for catalytic activity. The SDD mediates IKKβ dimerization, but dimerization per se is not important for maintaining IKKβ activity and instead is required for IKKβ activation. Other IKK family members, IKKα, TBK1 and IKK-i, may have a similar trimodular architecture and function.
|NF-κB essential modulator (NEMO) interaction with linear and lys-63 ubiquitin chains contributes to NF-κB activation.|
Hadian, K; Griesbach, RA; Dornauer, S; Wanger, TM; Nagel, D; Metlitzky, M; Beisker, W; Schmidt-Supprian, M; Krappmann, D
The Journal of biological chemistry 286 26107-17 2011
The IκB kinase (IKK) complex acts as a gatekeeper of canonical NF-κB signaling in response to upstream stimulation. IKK activation requires sensing of ubiquitin chains by the essential IKK regulatory subunit IKKγ/NEMO. However, it has remained enigmatic whether NEMO binding to Lys-63-linked or linear ubiquitin chains is critical for triggering IKK activation. We show here that the NEMO C terminus, comprising the ubiquitin binding region and a zinc finger, has a high preference for binding to linear ubiquitin chains. However, immobilization of NEMO, which may be reminiscent of cellular oligomerization, facilitates the interaction with Lys-63 ubiquitin chains. Moreover, selective mutations in NEMO that abolish association with linear ubiquitin but do not affect binding to Lys-63 ubiquitin are only partially compromising NF-κB signaling in response to TNFα stimulation in fibroblasts and T cells. In line with this, TNFα-triggered expression of NF-κB target genes and induction of apoptosis was partially compromised by NEMO mutations that selectively impair the binding to linear ubiquitin chains. Thus, in vivo NEMO interaction with linear and Lys-63 ubiquitin chains is required for optimal IKK activation, suggesting that both type of chains are cooperating in triggering canonical NF-κB signaling.
|Requirement of the NF-kappaB subunit p65/RelA for K-Ras-induced lung tumorigenesis.|
Bassères, DS; Ebbs, A; Levantini, E; Baldwin, AS
Cancer research 70 3537-46 2010
K-Ras-induced lung cancer is a very common disease, for which there are currently no effective therapies. Because therapy directly targeting the activity of oncogenic Ras has been unsuccessful, a different approach for novel therapy design is to identify critical Ras downstream oncogenic targets. Given that oncogenic Ras proteins activate the transcription factor NF-kappaB, and the importance of NF-kappaB in oncogenesis, we hypothesized that NF-kappaB would be an important K-Ras target in lung cancer. To address this hypothesis, we generated a NF-kappaB-EGFP reporter mouse model of K-Ras-induced lung cancer and determined that K-Ras activates NF-kappaB in lung tumors in situ. Furthermore, a mouse model was generated where activation of oncogenic K-Ras in lung cells was coupled with inactivation of the NF-kappaB subunit p65/RelA. In this model, deletion of p65/RelA reduces the number of K-Ras-induced lung tumors both in the presence and in the absence of the tumor suppressor p53. Lung tumors with loss of p65/RelA have higher numbers of apoptotic cells, reduced spread, and lower grade. Using lung cell lines expressing oncogenic K-Ras, we show that NF-kappaB is activated in these cells in a K-Ras-dependent manner and that NF-kappaB activation by K-Ras requires inhibitor of kappaB kinase beta (IKKbeta) kinase activity. Taken together, these results show the importance of the NF-kappaB subunit p65/RelA in K-Ras-induced lung transformation and identify IKKbeta as a potential therapeutic target for K-Ras-induced lung cancer.
|LPS induces KH-type splicing regulatory protein-dependent processing of microRNA-155 precursors in macrophages.|
Tina Ruggiero, Michele Trabucchi, Francesca De Santa, Simona Zupo, Brian D Harfe, Michael T McManus, M Geoff Rosenfeld, Paola Briata, Roberto Gherzi
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology 23 2898-908 2009
The importance of post-transcriptional mechanisms for the regulation of the homoeostasis of the immune system and the response to challenge by microorganisms is becoming increasingly appreciated. We investigated the contribution of microRNAs (miRNAs) to macrophage activation induced by lipopolysaccharide (LPS). We first observed that Dicer knockout in bone marrow-derived macrophages (BMDMs) increases the LPS-induced expression of some inflammation mediators. miRNA microarray analysis in BMDMs revealed that LPS significantly induces the expression of a single miRNA, miR-155, and this induction depends on enhanced miR-155 maturation from its precursors. The single-strand RNA-binding protein KH-type splicing regulatory protein (KSRP) binds to the terminal loop of miR-155 precursors and promotes their maturation. Both inhibition of miR-155 and KSRP knockdown enhance the LPS-induced expression of select inflammation mediators, and the effect of KSRP knockdown is reverted by mature miR-155. Our studies unveil the existence of an LPS-dependent post-transcriptional regulation of miR-155 biogenesis. Once induced, miR-155 finely tunes the expression of select inflammation mediators in response to LPS.
|Inactivation of NF-kappaB p50 leads to insulin sensitization in liver through post-translational inhibition of p70S6K.|
Zhanguo Gao,Jun Yin,Jin Zhang,Qing He,Owen P McGuinness,Jianping Ye
The Journal of biological chemistry 284 2009
In this study, we investigated the metabolic phenotype of the NF-kappaB p50 knock-out (p50-KO) mice. Compared with wild type mice, the p50-KO mice had an increase in food intake, but a decrease in body fat content. On chow diet, their blood glucose dropped much more than the wild type (WT) mice in the insulin tolerance test. Their glucose infusion rate was 30% higher than that of the WT mice in the hyperinsulinemic-euglycemic clamp. Their hepatic glucose production was suppressed more actively by insulin, and their insulin-induced glucose uptake was not altered in skeletal muscle or adipose tissue. In the liver, their p70S6K (S6K1) protein was significantly lower, and tumor necrosis factor-alpha (TNF-alpha) expression was much higher. Their S6K1 protein was reduced by TNF-alpha treatment in the primary culture of hepatocytes. S6K1 reduction was blocked by the proteasome inhibitor MG132. In their livers, IKK2 (IKKbeta) activity was reduced together with IKKgamma. Their S6K1 degradation was dependent on IKK2 deficiency. Reconstitution of the S6K1 protein in their liver blocked the increase in insulin sensitivity. S6K1 degradation was not observed in hepatocytes of the WT mice. The data suggest that inactivation of NF-kappaB p50 leads to suppression of IKK2 activity in the liver. IKK2 deficiency leads to S6K1 inhibition through TNF-induced protein degradation. The S6K1 reduction may contribute to insulin sensitivity in p50-KO mice. This study suggests that hepatic S6K1 may be a drug target in the treatment of insulin resistance.Full Text Article
|Pathways and Biomarkers of Toll-like Receptor (TLR) Signaling|