TAK1 regulates reactive oxygen species and cell death in keratinocytes, which is essential for skin integrity.
ABSTRACT: Mice with a keratinocyte-specific deletion of Tak1 exhibit severe skin inflammation due to hypersensitivity to tumor necrosis factor (TNF) killing. Here we have examined the mechanisms underlying this hypersensitivity. We found that TAK1 deficiency up-regulates reactive oxygen species (ROS) resulting in cell death upon TNF or oxidative stress challenge. Because blockade of NF-kappaB did not increase ROS or did not sensitize cells to oxidative stress in keratinocytes TAK1 regulates ROS mainly through the mechanisms other than those mediated by NF-kappaB. We found that c-Jun was decreased in TAK1-deficient keratinocytes and that ectopic expression of c-Jun could partially inhibit TNF-induced increase of ROS and cell death. Finally, we show that, in an in vivo setting, the antioxidant treatment could reduce an inflammatory condition in keratinocyte-specific Tak1 deletion mice. Thus, TAK1 regulates ROS partially through c-Jun, which is important for preventing ROS-induced skin inflammation.
Project description:Transforming growth factor beta-activated kinase 1 (TAK1) functions downstream of inflammatory cytokines to activate c-Jun N-terminal kinase (JNK) as well as NF-kappaB in several cell types. However, the functional role of TAK1 in an in vivo setting has not been determined. Here we have demonstrated that TAK1 is the major regulator of skin inflammation as well as keratinocyte death in vivo. Epidermal-specific deletion of TAK1 causes a severe inflammatory skin condition by postnatal day 6-8. The mutant skin also exhibits massive keratinocyte death. Analysis of keratinocytes isolated from the mutant skin revealed that TAK1 deficiency results in a striking increase in apoptosis in response to tumor necrosis factor (TNF). TAK1-deficient keratinocytes cannot activate NF-kappaB or JNK upon TNF treatment. These results suggest that TNF induces TAK1-deficient keratinocyte death because of the lack of NF-kappaB (and possibly JNK)-mediated cell survival signaling. Finally, we have shown that deletion of the TNF receptor can largely rescue keratinocyte death as well as inflammatory skin condition in epidermal-specific TAK1-deficient mice. Our results demonstrate that TAK1 is a master regulator of TNF signaling in skin and regulates skin inflammation and keratinocyte death.
Project description:TNF-related apoptosis-inducing ligand (TRAIL) is a potent inducer of cell death in several cancer cells, but many cells are resistant to TRAIL. The mechanism that determines sensitivity to TRAIL-killing is still elusive. Here we report that deletion of TAK1 kinase greatly increased activation of caspase-3 and cell death after TRAIL stimulation in keratinocytes, fibroblasts and cancer cells. Although TAK1 kinase is involved in NF-kappaB pathway, ablation of NF-kappaB did not alter sensitivity to TRAIL. We found that TRAIL could induce accumulation of reactive oxygen species (ROS) when TAK1 was deleted. Furthermore, we found that TAK1 deletion induced TRAIL-dependent downregulation of cIAP, which enhanced activation of caspase-3. These results show that TAK1 deletion facilitates TRAIL-induced cell death by activating caspase through ROS and downregulation of cIAP. Thus, inhibition of TAK1 can be an effective approach to increase TRAIL sensitivity.
Project description:Dysregulated reactive oxygen species (ROS) generation contributes to many human pathologies, including cancer and diabetes. During normal wound repair, inflammation-induced ROS production must be tightly controlled, but the mechanisms reining their generation remain unclear. Herein, we show that transforming growth factor ?-activated kinase 1 (TAK1) directly regulates stem cell factor (SCF) expression, which activates the protein kinase B (PKB)? pro-survival pathway in a cell-autonomous manner to protect keratinocytes from ROS-mediated cell death. TAK1 is a pivotal inflammatory mediator whose expression was transiently elevated during wound healing, paralleling the ROS production profile. TAK1 deficiency in keratinocytes led to increased apoptosis in response to anoikis and TNF-? treatment and was associated with elevated ROS level as analyzed by FACS. Using organotypic skin co-culture and comparative growth factor array analysis, we revealed a cell-autonomous mechanism that involved the SCF/c-Kit/PKB? signaling cascade. Ectopic expression of TAK1 or treatment with exogenous recombinant SCF restored the increased ROS production and apoptotic cell death in TAK1-deficient keratinocytes. Conversely, normal keratinocytes treated with various inhibitors targeting the SCF/c-Kit/PKB? pathway exhibited increased ROS production and TNF-?- or anoikis-induced apoptosis. Our study reveals a novel anti-apoptotic role for SCF in keratinocytes and identifies TAK1 as a novel player uniting inflammation and ROS regulation in skin redox biology.
Project description:The cytokines IL-1 and TNF induce expression of a series of genes that regulate inflammation through activation of NF-kappaB signal transduction pathways. TAK1, a MAPKKK, is critical for both IL-1- and TNF-induced activation of the NF-kappaB pathway. TAB2, a TAK1-binding protein, is involved in IL-1-induced NF-kappaB activation by physically linking TAK1 to TRAF6. However, IL-1-induced activation of NF-kappaB is not impaired in TAB2-deficient embryonic fibroblasts. Here we report the identification and characterization of a novel protein designated TAB3, a TAB2-like molecule that associates with TAK1 and can activate NF-kappaB similar to TAB2. Endogenous TAB3 interacts with TRAF6 and TRAF2 in an IL-1- and a TNF-dependent manner, respectively. Further more, IL-1 signaling leads to the ubiquitination of TAB2 and TAB3 through TRAF6. Cotransfection of siRNAs directed against both TAB2 and TAB3 inhibit both IL-1- and TNF-induced activation of TAK1 and NF-kappaB. These results suggest that TAB2 and TAB3 function redundantly as mediators of TAK1 activation in IL-1 and TNF signal transduction.
Project description:Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is known as a "death ligand"-a member of the TNF superfamily that binds to receptors bearing death domains. As well as causing apoptosis of certain types of tumor cells, TRAIL can activate both NF-kappaB and JNK signalling pathways. To determine the role of TGF-beta-Activated Kinase-1 (TAK1) in TRAIL signalling, we analyzed the effects of adding TRAIL to mouse embryonic fibroblasts (MEFs) derived from TAK1 conditional knockout mice. TAK1-/- MEFs were significantly more sensitive to killing by TRAIL than wild-type MEFs, and failed to activate NF-kappaB or JNK. Overexpression of IKK2-EE, a constitutive activator of NF-kappaB, protected TAK1-/- MEFs against TRAIL killing, suggesting that TAK1 activation of NF-kappaB is critical for the viability of cells treated with TRAIL. Consistent with this model, TRAIL failed to induce the survival genes cIAP2 and cFlipL in the absence of TAK1, whereas activation of NF-kappaB by IKK2-EE restored the levels of both proteins. Moreover, ectopic expression of cFlipL, but not cIAP2, in TAK1-/- MEFs strongly inhibited TRAIL-induced cell death. These results indicate that cells that survive TRAIL treatment may do so by activation of a TAK1-NF-kappaB pathway that drives expression of cFlipL, and suggest that TAK1 may be a good target for overcoming TRAIL resistance.
Project description:The intestinal epithelium is constantly exposed to inducers of reactive oxygen species (ROS), such as commensal microorganisms. Levels of ROS are normally maintained at nontoxic levels, but dysregulation of ROS is involved in intestinal inflammatory diseases. In this article, we report that TGF-?-activated kinase 1 (TAK1) is a key regulator of ROS in the intestinal epithelium. tak1 gene deletion in the mouse intestinal epithelium caused tissue damage involving enterocyte apoptosis, disruption of tight junctions, and inflammation. Disruption of TNF signaling, which is a major intestinal damage inducer, rescued the inflammatory conditions but not apoptosis or disruption of tight junctions in the TAK1-deficient intestinal epithelium, suggesting that TNF is not a primary inducer of the damage noted in TAK1-deficient intestinal epithelium. We found that TAK1 deficiency resulted in reduced expression of several antioxidant-responsive genes and reduced the protein level of a key antioxidant transcription factor NF-E2-related factor 2, which resulted in accumulation of ROS. Exogenous antioxidant treatment reduced apoptosis and disruption of tight junctions in the TAK1-deficient intestinal epithelium. Thus, TAK1 signaling regulates ROS through transcription factor NF-E2-related factor 2, which is important for intestinal epithelial integrity.
Project description:TGF-beta-activated kinase 1 (TAK1) is a MAP3K family member that activates NF-kappaB and JNK via Toll-like receptors and the receptors for IL-1, TNF-alpha, and TGF-beta. Because the TAK1 downstream molecules NF-kappaB and JNK have opposite effects on cell death and carcinogenesis, the role of TAK1 in the liver is unpredictable. To address this issue, we generated hepatocyte-specific Tak1-deficient (Tak1DeltaHEP) mice. The Tak1DeltaHEP mice displayed spontaneous hepatocyte death, compensatory proliferation, inflammatory cell infiltration, and perisinusoidal fibrosis at age 1 month. Older Tak1DeltaHEP mice developed multiple cancer nodules characterized by increased expression of fetal liver genes including alpha-fetoprotein. Cultures of primary hepatocytes deficient in Tak1 exhibited spontaneous cell death that was further increased in response to TNF-alpha. TNF-alpha increased caspase-3 activity but activated neither NF-kappaB nor JNK in Tak1-deficient hepatocytes. Genetic abrogation of TNF receptor type I (TNFRI) in Tak1DeltaHEP mice reduced liver damage, inflammation, and fibrosis compared with unmodified Tak1DeltaHEP mice. In conclusion, hepatocyte-specific deletion of TAK1 in mice resulted in spontaneous hepatocyte death, inflammation, fibrosis, and carcinogenesis that was partially mediated by TNFR signaling, indicating that TAK1 is an essential component for cellular homeostasis in the liver.
Project description:CC2D1A is an evolutionarily conserved protein that contains four DM14 domains at the N terminus and a C2 domain at the C terminus. Loss-of-function mutations in CC2D1A have been linked to mental retardation in human, but the biochemical function of this protein is largely unknown. Here, we show that CC2D1A is a potent activator of NF-kappaB. The activation of NF-kappaB by CC2D1A requires its C2 domain. CC2D1A activates NF-kappaB in a manner that depends on the ubiquitin-conjugating enzyme Ubc13, TNF receptor-associated factor TRAF2, the protein kinase TAK1, and the IkappaB kinase (IKK) complex. In addition, the deubiquitination enzyme Cylindromatosis (CYLD) negatively regulates the activity of CC2D1A. These results suggest that CC2D1A activates NF-kappaB through the canonical IKK pathway.
Project description:Helicobacter pylori-initiated chronic gastritis is characterized by the cag pathogenicity island-dependent upregulation of proinflammatory cytokines, which is largely mediated by the transcription factor nuclear factor (NF)-kappaB. However, the cag pathogenicity island-encoded proteins and cellular signalling molecules that are involved in H. pylori-induced NF-kappaB activation and inflammatory response remain unclear. Here, we show that H. pylori virulence factor CagA and host protein transforming growth factor-beta-activated kinase 1 (TAK1) are essential for H. pylori-induced activation of NF-kappaB. CagA physically associates with TAK1 and enhances its activity and TAK1-induced NF-kappaB activation through the tumour necrosis factor receptor-associated factor 6-mediated, Lys 63-linked ubiquitination of TAK1. These findings show that polyubiquitination of TAK1 regulates the activation of NF-kappaB, which in turn is used by H. pylori CagA for the H. pylori-induced inflammatory response.
Project description:Transforming growth factor beta-activated kinase 1 (TAK1), a member of the MAPKKK family, is a key mediator of proinflammatory and stress signals. Activation of TAK1 by proinflammatory cytokines and T and B cell receptors induces the nuclear localization of nuclear factor kappaB (NF-kappaB) and the activation of c-Jun N-terminal kinase (JNK)/AP1 and P38, which play important roles in mediating inflammation, immune responses, T and B cell activation, and epithelial cell survival. Here, we report that TAK1 is critical for the survival of both hematopoietic cells and hepatocytes. Deletion of TAK1 results in bone marrow (BM) and liver failure in mice due to the massive apoptotic death of hematopoietic cells and hepatocytes. Hematopoietic stem cells and progenitors were among those hematopoietic cells affected by TAK1 deletion-induced cell death. This apoptotic cell death is autonomous, as demonstrated by reciprocal BM transplantation. Deletion of TAK1 resulted in the inactivation of both JNK and NF-kappaB signaling, as well as the down-regulation of expression of prosurvival genes.