Serine/threonine kinase which acts as an essential component of the MAP kinase signal transduction pathway. Plays an important role in the cascades of cellular responses evoked by changes in the environment. Mediates signaling for determination of cell fate such as differentiation and survival. Plays a crucial role in the apoptosis signal transduction pathway through mitochondria-dependent caspase activation. MAP3K5/ASK1 is required for the innate immune response, which is essential for host defense against a wide range of pathogens. Mediates signal transduction of various stressors like oxidative stress as well as by receptor-mediated inflammatory signals, such as the tumor necrosis factor (TNF) or lipopolysaccharide (LPS). Once activated, acts as an upstream activator of the MKK/JNK signal transduction cascade and the p38 MAPK signal transduction cascade through the phosphorylation and activation of several MAP kinase kinases like MAP2K4/SEK1, MAP2K3/MKK3, MAP2K6/MKK6 and MAP2K7/MKK7. These MAP2Ks in turn activate p38 MAPKs and c-jun N-terminal kinases (JNKs). Both p38 MAPK and JNKs control the transcription factors activator protein-1 (AP-1).
Apoptosis signal-regulating kinase (ASK) 1 was recently identified as a mitogen-activated protein (MAP) kinase kinase kinase which activates the c-Jun N-terminal kinase (JNK) and p38 MAP kinase pathways and is required for tumor necrosis factor (TNF)-alpha-induced apoptosis; however, the mechanism regulating ASK1 activity is unknown. Through genetic screening for ASK1-binding proteins, thioredoxin (Trx), a reduction/oxidation (redox)-regulatory protein thought to have anti-apoptotic effects, was identified as an interacting partner of ASK1. Trx associated with the N-terminal portion of ASK1 in vitro and in vivo. Expression of Trx inhibited ASK1 kinase activity and the subsequent ASK1-dependent apoptosis. Treatment of cells with N-acetyl-L-cysteine also inhibited serum withdrawal-, TNF-alpha- and hydrogen peroxide-induced activation of ASK1 as well as apoptosis. The interaction between Trx and ASK1 was found to be highly dependent on the redox status of Trx. Moreover, inhibition of Trx resulted in activation of endogenous ASK1 activity, suggesting that Trx is a physiological inhibitor of ASK1. The evidence that Trx is a negative regulator of ASK1 suggests possible mechanisms for redox regulation of the apoptosis signal transduction pathway as well as the effects of antioxidants against cytokine- and stress-induced apoptosis.
Apoptosis signal-regulating kinase 1 (ASK1) plays a pivotal role in oxidative stress-induced cell death. Reactive oxygen species disrupt the interaction of ASK1 with its cellular inhibitor thioredoxin and thereby activates ASK1. However, the precise mechanism by which ASK1 freed from thioredoxin undergoes oligomerization-dependent activation has not been fully elucidated. Here we show that endogenous ASK1 constitutively forms a high molecular mass complex including Trx ( approximately 1,500-2,000 kDa), which we designate ASK1 signalosome. Upon H(2)O(2) treatment, the ASK1 signalosome forms a higher molecular mass complex at least in part because of the recruitment of tumor necrosis factor receptor-associated factor 2 (TRAF2) and TRAF6. Consistent with our previous findings that TRAF2 and TRAF6 activate ASK1, H(2)O(2)-induced ASK1 activation and cell death were strongly reduced in the cells derived from Traf2-/- and Traf6-/- mice. A novel signaling complex including TRAF2, TRAF6, and ASK1 may thus be the key component in oxidative stress-induced cell death.
Cells differentiate in response to various extracellular stimuli. This cellular response requires intracellular signaling pathways. The mitogen-activated protein (MAP) kinase cascade is a core signal transduction pathway that determines the fate of many kinds of cell. MAP kinase kinase kinase activates MAP kinase kinase, which in turn activates MAP kinase. Apoptosis signal-regulating kinase (ASK1) was identified as a MAP kinase kinase kinase involved in the stress-induced apoptosis-signaling cascade that activates the SEK1-JNK and MKK3/MKK6-p38 MAP kinase cascades. Expression of the constitutively active form of ASK1 (ASK1-DeltaN) in keratinocytes induced significant morphological changes and differentiation markers, transglutaminase-1, loricrin, and involucrin. A transient increase in p21(Cip1/WAF1) reduced DNA synthesis, and cell cycle analysis verified the differentiation. p38 MAP kinase inhibitors, SB202190 and SB203580, abolished the induction of differentiation markers, transglutaminase-1, loricrin, and involucrin. In turn, the induction of differentiation with ceramide in keratinocytes caused an increase in ASK1 expression and activity. Furthermore, normal human skin expresses ASK1 protein in the upper epidermis, implicating ASK1 in in vivo keratinocyte differentiation. We propose that the ASK1-p38 MAP kinase cascade is a new intracellular regulator of keratinocyte differentiation.
J. Biol. Chem. 271, 31607-31611 (1996)[PubMed:8940179]
Mitogen-activated protein kinase (MAPK) signaling cascades include MAPK or extracellular signal-regulated kinase (ERK), MAPK kinase (MKK or MEK), and MAPK kinase kinase (MAPKKK or MEKK). MAPKK kinase/MEKK phosphorylates and activates its downstream protein kinase, MAPK kinase/MEK, which in turn activates MAPK. We report herein the isolation of a cDNA encoding a novel protein kinase designated MAPKKK5 from a human macrophage library. The nucleotide sequence predicts that MAPKKK5 encodes an open reading frame of 1374 amino acids with all 11 kinase subdomains. The putative catalytic domain of MAPKKK5 shows significant sequence homology to the kinase domains of the MAPKKK/MEKK level protein kinases from mouse MEKK2 and -3, Drosophila melanogaster PK92B, Saccharomyces cerevisiae STE11, and Schizosaccharomyces pombe BYR2. Northern blot analysis showed that MAPKKK5 transcript is abundantly expressed in human heart and pancreas. When transiently expressed in COS and 293 cells, MAPKKK5 markedly activated c-Jun N-terminal kinase or stress-activated protein kinase, but not MAPK/ERK. Furthermore, MAPKKK5 that was immunoprecipitated from transfected 293 cells was able to phosphorylate and activate MKK4 in vitro, suggesting that MAPKKK5 may be an upstream activator of MKK4 in the c-Jun N-terminal kinase pathway.
ASK1 activates JNK and p38 mitogen-activated protein kinases and constitutes a pivotal signaling pathway in cytokine- and stress-induced apoptosis. However, little is known about the mechanism of how ASK1 executes apoptosis. Here we investigated the roles of caspases and mitochondria in ASK1-induced apoptosis. We found that benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (zVAD-fmk), a broad-spectrum caspase inhibitor, mostly inhibited ASK1-induced cell death, suggesting that caspases are required for ASK1-induced apoptosis. Overexpression of ASK1DeltaN, a constitutively active mutant of ASK1, induced cytochrome c release from mitochondria and activation of caspase-9 and caspase-3 but not of caspase-8-like proteases. Consistently, caspase-8-deficient (Casp8 (-/-)) cells were sensitive to ASK1-induced caspase-3 activation and apoptosis, suggesting that caspase-8 is dispensable for ASK1-induced apoptosis, whereas ASK1 failed to activate caspase-3 in caspase-9-dificient (Casp9 (-/-)) cells. Moreover, mitochondrial cytochrome c release, which was not inhibited by zVAD-fmk, preceded the onset of caspase-3 activation and cell death induced by ASK1. ASK1 thus appears to execute apoptosis mainly by the mitochondria-dependent caspase activation.
Apoptosis signal-regulating kinase 1 (ASK1) is a critical mediator of apoptotic signaling pathways initiated by a variety of death stimuli. Its activity is tightly controlled by various mechanisms such as covalent modification and protein-protein interaction. One of the proteins that control ASK1 function is 14-3-3zeta, a member of the 14-3-3 protein family. Here, we report that ASK1 is capable of binding to other isoforms of 14-3-3, suggesting that binding ASK1 is a general property of the 14-3-3 family. In support of this notion, mutational analysis revealed that the ASK1/14-3-3 interaction was mediated by the conserved amphipathic groove of 14-3-3 with some residue selectivity. Functionally, expression of various isoforms of 14-3-3 suppressed ASK1-induced apoptosis. To understand how 14-3-3 controls the ASK1 activity, we examined intracellular localization of ASK1 upon 14-3-3 co-expression. We found that 14-3-3 co-expression is correlated with the translocation of ASK1 from the cytoplasm to a perinuclear localization, likely the ER compartment. Consistent with this notion, ASK1(S967A), a 14-3-3 binding defective mutant of ASK, showed no change in intracellular distribution upon 14-3-3 co-expression. These data support a model that 14-3-3 proteins regulate the proapoptotic function of ASK1 in part by controlling its subcellular distribution.
Apoptosis signal-regulating kinase 1 (ASK1) is a MAPKKK family member which activates c-Jun N-terminal kinase (JNK) and p38. In non-stressed cells, ASK1 exists as an inactive complex with the reduced form of thioredoxin. Oxidative stress such as hydrogen peroxide (H2O2) disrupts the ASK1-thioredoxin complex by oxidization of thioredoxin and thereby activates ASK1. The precise mechanism by which ASK1 is activated after its release from thioredoxin is unknown. Here we show that phosphorylation of Thr845 at the activation loop is essential for ASK1 to be activated by H2O2. ASK1 appears to form a silent homo-oligomer through its C-terminal coiled-coil region in non-stressed cells. Following H2O2 treatment, pre-existing ASK1 oligomer undergoes conformational change and creates a new interface within an oligomer, which ultimately leads to trans-autophosphorylation of Thr845. Thus, direct interaction via the coiled-coil region is required for self-scaffolding but not sufficient for activation of ASK1. Importantly, Thr845 of ASK1 can also be trans-phosphorylated by an unidentified Thr845 kinase in response to H2O2 treatment. We propose that this potential Thr845 kinase may be an ignition kinase that triggers Thr845 phosphorylation in oligomerized and activation-competent forms of ASK1.
TRAF2 and ASK1 play essential roles in tumor necrosis factor alpha (TNF-alpha)-induced mitogen-activated protein kinase signaling. Stimulation through TNF receptor 2 (TNFR2) leads to TRAF2 ubiquitination and subsequent proteasomal degradation. Here we show that TNFR2 signaling also leads to selective ASK1 ubiquitination and degradation in proteasomes. c-IAP1 was identified as the ubiquitin protein ligase for ASK1 ubiquitination, and studies with primary B cells from c-IAP1 knock-out animals revealed that c-IAP1 is required for TNFR2-induced TRAF2 and ASK1 degradation. Moreover, in the absence of c-IAP1 TNFR2-mediated p38 and JNK activation was prolonged. Thus, the ubiquitin protein ligase activity of c-IAP1 is responsible for regulating the duration of TNF signaling in primary cells expressing TNFR2.
The stress-activated protein kinases (SAPKs, also called c-Jun NH(2)-terminal kinases) and the p38s, two mitogen-activated protein kinase (MAPK) subgroups activated by cytokines of the tumor necrosis factor (TNF) family, are pivotal to the de novo gene expression elicited as part of the inflammatory response. Apoptosis signal-regulating kinase 1 (ASK1) is a MAPK kinase kinase (MAP3K) that activates both the SAPKs and p38s in vivo. Here we show that TNF receptor (TNFR) associated factor 2 (TRAF2), an adapter protein that couples TNFRs to the SAPKs and p38s, can activate ASK1 in vivo and can interact in vivo with the amino- and carboxyl-terminal noncatalytic domains of the ASK1 polypeptide. Expression of the amino-terminal noncatalytic domain of ASK1 can inhibit TNF and TRAF2 activation of SAPK. TNF can stimulate the production of reactive oxygen species (ROS), and the redox-sensing enzyme thioredoxin (Trx) is an endogenous inhibitor of ASK1. We also show that expression of TRAF2 fosters the production of ROS in transfected cells. We demonstrate that Trx significantly inhibits TRAF2 activation of SAPK and blocks the ASK1-TRAF2 interaction in a reaction reversed by oxidants. Finally, the mechanism of ASK1 activation involves, in part, homo-oligomerization. We show that expression of ASK1 with TRAF2 enhances in vivo ASK1 homo-oligomerization in a manner dependent, in part, upon the TRAF2 RING effector domain and the generation of ROS. Thus, activation of ASK1 by TNF requires the ROS-mediated dissociation of Trx possibly followed by the binding of TRAF2 and consequent ASK1 homo-oligomerization.
Mitogen-activated protein (MAP) kinase cascades are activated in response to various extracellular stimuli, including growth factors and environmental stresses. A MAP kinase kinase kinase (MAPKKK), termed ASK1, was identified that activated two different subgroups of MAP kinase kinases (MAPKK), SEK1 (or MKK4) and MKK3/MAPKK6 (or MKK6), which in turn activated stress-activated protein kinase (SAPK, also known as JNK; c-Jun amino-terminal kinase) and p38 subgroups of MAP kinases, respectively. Overexpression of ASK1 induced apoptotic cell death, and ASK1 was activated in cells treated with tumor necrosis factor-alpha (TNF-alpha). Moreover, TNF-alpha-induced apoptosis was inhibited by a catalytically inactive form of ASK1. ASK1 may be a key element in the mechanism of stress- and cytokine-induced apoptosis.
Tumor necrosis factor (TNF)-induced activation of the c-jun N-terminal kinase (JNK, also known as SAPK; stress-activated protein kinase) requires TNF receptor-associated factor 2 (TRAF2). The apoptosis signal-regulating kinase 1 (ASK1) is activated by TNF and stimulates JNK activation. Here we show that ASK1 interacts with members of the TRAF family and is activated by TRAF2, TRAF5, and TRAF6 overexpression. A truncated derivative of TRAF2, which inhibits JNK activation by TNF, blocks TNF-induced ASK1 activation. A catalytically inactive mutant of ASK1 is a dominant-negative inhibitor of TNF- and TRAF2-induced JNK activation. In untransfected mammalian cells, ASK1 rapidly associates with TRAF2 in a TNF-dependent manner. Thus, ASK1 is a mediator of TRAF2-induced JNK activation.
Cisplatin and its analogues have been widely used for treatment of human cancer. However, most patients eventually develop resistance to treatment through a mechanism that remains obscure. Previously, we found that AKT2 is frequently overexpressed and/or activated in human ovarian and breast cancers. Here we demonstrate that constitutively active AKT2 renders cisplatin-sensitive A2780S ovarian cancer cells resistant to cisplatin, whereas phosphatidylinositol 3-kinase inhibitor or dominant negative AKT2 sensitizes A2780S and cisplatin-resistant A2780CP cells to cisplatin-induced apoptosis through regulation of the ASK1/JNK/p38 pathway. AKT2 interacts with and phosphorylates ASK1 at Ser-83 resulting in inhibition of its kinase activity. Accordingly, activated AKT2 blocked signaling down-stream of ASK1, including activation of JNK and p38 and the conversion of Bax to its active conformation. Expression of nonphosphorylatable ASK1-S83A overrode the AKT2-inhibited JNK/p38 activity and Bax conformational changes, whereas phosphomimic ASK1-S83D inhibited the effects of cisplatin on JNK/p38 and Bax. Cisplatin-induced Bax conformation change was inhibited by inhibitors or dominant negative forms of JNK and p38. In conclusion, our data indicate that AKT2 inhibits cisplatin-induced JNK/p38 and Bax activation through phosphorylation of ASK1 and thus, plays an important role in chemoresistance. Further, regulation of the ASK1/JNK/p38/Bax pathway by AKT2 provides a new mechanism contributing to its antiapoptotic effects.
The mammalian mitogen-activated protein (MAP) kinase kinase kinase apoptosis signal-regulating kinase 1 (ASK1) is a pivotal component in cytokine- and stress-induced apoptosis. It also regulates cell differentiation and survival through p38 MAP kinase activation. Here we show that Ca2+ signalling regulates the ASK1-p38 MAP kinase cascade. Ca2+ influx evoked by membrane depolarization in primary neurons and synaptosomes induced activation of p38, which was impaired in those derived from ASK1-deficient mice. Ca2+/calmodulin-dependent protein kinase type II (CaMKII) activated ASK1 by phosphorylation. Moreover, p38 activation induced by the expression of constitutively active CaMKII required endogenous ASK1. Thus, ASK1 is a critical intermediate of Ca2+ signalling between CaMKII and p38 MAP kinase.
Oxidative stress has been indicated in a variety of pathological processes such as atherosclerosis, diabetes, and neurodegenerative diseases. Understanding how intracellular signaling pathways respond to oxidative insults such as hydrogen peroxide (H(2)O(2)) would have significant therapeutic implications. Recent genetic studies have placed apoptosis signal-regulating kinase 1 (ASK1) in a pivotal position in transmitting H(2)O(2)-initiated signals. How ASK1 is activated by H(2)O(2), though, remains a subject of intense investigation. Here we report a mechanism by which H(2)O(2) induces ASK1 activation through dynamic control of its phosphorylation at serine 967. We found that treatment of COS7 cells with H(2)O(2) triggers dephosphorylation of Ser-967 through an okadaic acid-sensitive phosphatase, resulting in dissociation of the ASK1.14-3-3 complex with concomitant increase of ASK1 catalytic activity and ASK1-mediated activation of JNK and p38 pathways.
Apoptosis signal-regulating kinase 1 (ASK1) is a MAP kinase kinase kinase (MAPKKK) that activates the JNK and p38 MAP kinase cascades and is activated in response to oxidative stress such as hydrogen peroxide (H(2)O(2)). A yeast two-hybrid screening identified a serine/threonine protein phosphatase 5 (PP5) as a binding partner of ASK1. PP5 directly dephosphorylated an essential phospho-threonine residue within the kinase domain of ASK1 and thereby inactivated ASK1 activity in vitro and in vivo. The interaction between PP5 and ASK1 was induced by H(2)O(2) treatment and was followed by the decrease in ASK1 activity. PP5 inhibited not only H(2)O(2)-induced sustained activation of ASK1 but also ASK1-dependent apoptosis. Thus, PP5 appears to act as a physiological inhibitor of ASK1-JNK/p38 pathways by negative feedback.
Proc. Natl. Acad. Sci. U.S.A. 96, 8511-8515 (1999)[PubMed:10411906]
Apoptosis signal-regulating kinase 1 (ASK1) is a pivotal component of a signaling pathway induced by many death stimuli, including tumor necrosis factor alpha, Fas, and the anticancer drugs cisplatin and paclitaxel. Here we report that ASK1 proapoptotic activity is antagonized by association with 14-3-3 proteins. We found that ASK1 specifically bound 14-3-3 proteins via a site involving Ser-967 of ASK1. Interestingly, overexpression of 14-3-3 in HeLa cells blocked ASK1-induced apoptosis whereas disruption of the ASK1/14-3-3 interaction dramatically accelerated ASK1-induced cell death. Targeting of ASK1 by a 14-3-3-mediated survival pathway may provide a novel mechanism for the suppression of apoptosis.
The Akt family of serine/threonine-directed kinases promotes cellular survival in part by phosphorylating and inhibiting death-inducing proteins. Here we describe a novel functional interaction between Akt and apoptosis signal-regulating kinase 1 (ASK1), a mitogen-activated protein kinase kinase kinase. Akt decreased ASK1 kinase activity stimulated by both oxidative stress and overexpression in 293 cells by phosphorylating a consensus Akt site at serine 83 of ASK1. Activation of the phosphoinositide 3-kinase (PI3-K)/Akt pathway also inhibited the serum deprivation-induced activity of endogenous ASK1 in L929 cells. An association between Akt and ASK1 was detected in cells by coimmunoprecipitation. Phosphorylation by Akt inhibited ASK1-mediated c-Jun N-terminal kinase and activating transcription factor 2 activities in intact cells. Finally, activation of the PI3-K/Akt pathway reduced apoptosis induced by ASK1 in a manner dependent on phosphorylation of serine 83 of ASK1. These results provide the first direct link between Akt and the family of stress-activated kinases.
Apoptosis signal-regulating kinase (ASK) 1 is a mitogen-activated protein kinase kinase kinase (MAP3K) in the c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase pathways that play multiple important roles in cytokine and stress responses. Here we show that ASK2, a highly related serine/threonine kinase to ASK1, also functions as a MAP3K only in a heteromeric complex with ASK1. We found that endogenous ASK2 was constitutively degraded in ASK1-deficient cells, suggesting that ASK1 is required for the stability of ASK2. ASK2 in a heteromeric complex with a kinase-negative mutant of ASK1 (ASK1-KN) effectively activated MAP2K and was more competent to respond to oxidative stress than ASK2 alone. Knockdown of ASK2 revealed that ASK2 was required for oxidative stress-induced JNK activation. These results suggest that ASK2 forms a functional MAP3K complex with ASK1, in which ASK1 supports the stability and the active configuration of ASK2. Moreover, ASK2 was found to activate ASK1 by direct phosphorylation, suggesting that ASK1 and ASK2 in a heteromeric complex facilitate their activities to each other by distinct mechanisms. Such a formation of functional heteromeric complex between different MAP3Ks may be advantageous for cells to cope with a wide variety of stimuli by fine regulation of cellular responses.
Apoptosis signal-regulating kinase 1 (ASK1) is a mitogen-activated protein kinase kinase kinase family member that plays a central role in cytokine- and stress-induced apoptosis by activating c-Jun N-terminal kinase and p38 signaling cascades. ASK1-induced apoptotic activity is up-regulated by two cellular factors, Daxx and TRAF2, through direct protein-protein interactions. Daxx and TRAF2 are death receptor-associated proteins in Fas and tumor necrosis factor-alpha pathways, respectively. Recent studies suggest that calcium signaling may regulate ASK1 pathway. Here we report that human D53L1, a member of the tumor protein D52 family involved in cell proliferation and calcium signaling, up-regulates the ASK1-induced apoptosis. The human D53L1 physically interacts with the C-terminal regulatory domain of ASK1 and promotes ASK1-induced apoptotic activity by activating caspase signaling in mammalian cells. In luciferase reporter assays, hD53L1 activates c-Jun N-terminal kinase-mediated transactivation in the presence of ASK1. Expression of hD53L1 enhances autophosphorylation and kinase activity of ASK1 but has no effect on ASK1 oligomerization that is necessary for kinase activity and on binding of ASK1 to MKK6, a downstream factor of ASK1. Taken together, these results suggest that activation of ASK1 by hD53L1 may provide a novel mechanism for ASK1 regulation.
The Fas death receptor can activate the Jun NH2-terminal kinase (JNK) pathway through the receptor-associated protein Daxx. Daxx was found to activate the JNK kinase kinase ASK1, and overexpression of a kinase-deficient ASK1 mutant inhibited Fas- and Daxx-induced apoptosis and JNK activation. Fas activation induced Daxx to interact with ASK1, which consequently relieved an inhibitory intramolecular interaction between the amino- and carboxyl-termini of ASK1, activating its kinase activity. The Daxx-ASK1 connection completes a signaling pathway from a cell surface death receptor to kinase cascades that modulate nuclear transcription factors.
Evidence
2:
Inferred from Physical InteractionIntAct
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
Evidence
3:
Inferred from Physical InteractionIntAct
Expression and activity of the germinal center kinase, Ste20-like kinase (SLK), are increased during kidney development and recovery from ischemic acute renal failure. In this study, we characterize the activation and functional role of SLK. SLK underwent dimerization via the C-terminal domain, and dimerization enhanced SLK activity. In contrast, the C-terminal domain of SLK did not dimerize with a related kinase, Mst1, and did not affect Mst1 activity. Phosphorylation/dephosphorylation of SLK were not associated with changes in kinase activity. SLK induced phosphorylation of apoptosis signal-regulating kinase-1 (ASK1) and increased ASK1 activity, indicating that ASK1 is a substrate of SLK. Moreover, SLK stimulated phosphorylation of p38 mitogen-activated protein kinase via ASK1, but not c-Jun N-terminal kinase nor extracellular signal-regulated kinase. Chemical anoxia and recovery during re-exposure to glucose (ischemia-reperfusion injury in cell culture) stimulated SLK activity. Overexpression of SLK enhanced anoxia/recovery-induced apoptosis, release of cytochrome c, and activities of caspase-8 and -9, and apoptosis was reduced significantly with p38 and caspase-9 inhibitors. Induction of the endoplasmic reticulum stress response by anoxia/recovery or tunicamycin (monitored by induction of Bip or Grp94 expression, phosphorylation of eukaryotic translation initiation factor 2alpha subunit, expression of CHOP, and activation of caspase-12) was attenuated in cells that overexpress SLK. Thus, SLK is an anoxia/recovery-dependent kinase that is activated via homodimerization and that signals via ASK1 and p38 to promote apoptosis. Attenuation of the protective aspects of the endoplasmic reticulum stress response by SLK may contribute to its proapoptotic effect.
Apoptosis signal-regulating kinase (ASK) 1 is a mitogen-activated protein kinase kinase kinase (MAP3K) in the c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase pathways that play multiple important roles in cytokine and stress responses. Here we show that ASK2, a highly related serine/threonine kinase to ASK1, also functions as a MAP3K only in a heteromeric complex with ASK1. We found that endogenous ASK2 was constitutively degraded in ASK1-deficient cells, suggesting that ASK1 is required for the stability of ASK2. ASK2 in a heteromeric complex with a kinase-negative mutant of ASK1 (ASK1-KN) effectively activated MAP2K and was more competent to respond to oxidative stress than ASK2 alone. Knockdown of ASK2 revealed that ASK2 was required for oxidative stress-induced JNK activation. These results suggest that ASK2 forms a functional MAP3K complex with ASK1, in which ASK1 supports the stability and the active configuration of ASK2. Moreover, ASK2 was found to activate ASK1 by direct phosphorylation, suggesting that ASK1 and ASK2 in a heteromeric complex facilitate their activities to each other by distinct mechanisms. Such a formation of functional heteromeric complex between different MAP3Ks may be advantageous for cells to cope with a wide variety of stimuli by fine regulation of cellular responses.
Catalysis of the phosphorylation and activation of a MAP kinase kinase; each MAP kinase kinase can be phosphorylated by any of several MAP kinase kinase kinases.
Apoptosis signal-regulating kinase (ASK) 1 is a mitogen-activated protein kinase kinase kinase (MAP3K) in the c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase pathways that play multiple important roles in cytokine and stress responses. Here we show that ASK2, a highly related serine/threonine kinase to ASK1, also functions as a MAP3K only in a heteromeric complex with ASK1. We found that endogenous ASK2 was constitutively degraded in ASK1-deficient cells, suggesting that ASK1 is required for the stability of ASK2. ASK2 in a heteromeric complex with a kinase-negative mutant of ASK1 (ASK1-KN) effectively activated MAP2K and was more competent to respond to oxidative stress than ASK2 alone. Knockdown of ASK2 revealed that ASK2 was required for oxidative stress-induced JNK activation. These results suggest that ASK2 forms a functional MAP3K complex with ASK1, in which ASK1 supports the stability and the active configuration of ASK2. Moreover, ASK2 was found to activate ASK1 by direct phosphorylation, suggesting that ASK1 and ASK2 in a heteromeric complex facilitate their activities to each other by distinct mechanisms. Such a formation of functional heteromeric complex between different MAP3Ks may be advantageous for cells to cope with a wide variety of stimuli by fine regulation of cellular responses.
Interacting selectively and non-covalently with any protein or protein complex (a complex of two or more proteins that may include other nonprotein molecules).
Evidence
1:
Inferred from Physical InteractionIntAct
Heat shock protein 27 (HSP27) confers cellular protection against a variety of cytotoxic stresses and also against physiological stresses associated with growth arrest or receptor-mediated apoptosis. Phosphorylation modulates the activity of HSP27 by causing a major change in the supramolecular organization of the protein, which shifts from oligomers to dimers. Here we show that phosphorylated dimers of HSP27 interact with Daxx, a mediator of Fas-induced apoptosis, preventing the interaction of Daxx with both Ask1 and Fas and blocking Daxx-mediated apoptosis. No such inhibition was observed with an HSP27 phosphorylation mutant that is only expressed as oligomers or when apoptosis was induced by transfection of a Daxx mutant lacking its HSP27 binding domain. HSP27 expression had no effect on Fas-induced FADD- and caspase-dependent apoptosis. However, HSP27 blocked Fas-induced translocation of Daxx from the nucleus to the cytoplasm and Fas-induced Daxx- and Ask1-dependent apoptosis. The observations revealed a new level of regulation of the Fas pathway and suggest a mechanism for the phosphorylation-dependent protective function of HSP27 during stress and differentiation.
Evidence
2:
Inferred from Physical InteractionIntAct
The Akt family of serine/threonine-directed kinases promotes cellular survival in part by phosphorylating and inhibiting death-inducing proteins. Here we describe a novel functional interaction between Akt and apoptosis signal-regulating kinase 1 (ASK1), a mitogen-activated protein kinase kinase kinase. Akt decreased ASK1 kinase activity stimulated by both oxidative stress and overexpression in 293 cells by phosphorylating a consensus Akt site at serine 83 of ASK1. Activation of the phosphoinositide 3-kinase (PI3-K)/Akt pathway also inhibited the serum deprivation-induced activity of endogenous ASK1 in L929 cells. An association between Akt and ASK1 was detected in cells by coimmunoprecipitation. Phosphorylation by Akt inhibited ASK1-mediated c-Jun N-terminal kinase and activating transcription factor 2 activities in intact cells. Finally, activation of the PI3-K/Akt pathway reduced apoptosis induced by ASK1 in a manner dependent on phosphorylation of serine 83 of ASK1. These results provide the first direct link between Akt and the family of stress-activated kinases.
Evidence
3:
Inferred from Physical InteractionBHF-UCL
TNF-alpha activates ASK1 in part by dissociating 14-3-3 from apoptosis signal-regulating kinase 1 (ASK1). In the present study, we identified a novel Ras GTPase-activating protein (Ras-GAP) as an ASK1-interacting protein (AIP1). AIP1 binds to the C-terminal domain of ASK1 via a lysine-rich cluster within the N-terminal C2 domain. AIP1 exists in a closed form through an intramolecular interaction between the N-terminus and the C-terminus, and TNF-alpha induces unfolding of AIP1 leading to association of AIP1 with ASK1. Thus, the N-terminus of AIP1 containing the C2 and GAP domains constitutively binds to ASK1 and facilitates the release of 14-3-3 from ASK1. In contrast to 14-3-3, AIP1 binds preferentially to dephosphorylated ASK1. Recruited AIP1 enhances ASK1-induced JNK activation, and the ASK1 binding and the GAP activity of AIP1 are critical for AIP1-enhanced ASK1 activation. Furthermore, TNF-induced ASK1/JNK activation is significantly blunted in cells where AIP1 is knocked down by RNA interference. These data suggest that AIP1 mediates TNF-alpha-induced ASK1 activation by facilitating dissociation of inhibitor 14-3-3 from ASK1, a novel mechanism by which TNF-alpha activates ASK1.
Evidence
4:
Inferred from Physical InteractionIntAct
Apoptosis signal-regulating kinase (ASK) 1 is a mitogen-activated protein kinase kinase kinase (MAP3K) in the c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase pathways that play multiple important roles in cytokine and stress responses. Here we show that ASK2, a highly related serine/threonine kinase to ASK1, also functions as a MAP3K only in a heteromeric complex with ASK1. We found that endogenous ASK2 was constitutively degraded in ASK1-deficient cells, suggesting that ASK1 is required for the stability of ASK2. ASK2 in a heteromeric complex with a kinase-negative mutant of ASK1 (ASK1-KN) effectively activated MAP2K and was more competent to respond to oxidative stress than ASK2 alone. Knockdown of ASK2 revealed that ASK2 was required for oxidative stress-induced JNK activation. These results suggest that ASK2 forms a functional MAP3K complex with ASK1, in which ASK1 supports the stability and the active configuration of ASK2. Moreover, ASK2 was found to activate ASK1 by direct phosphorylation, suggesting that ASK1 and ASK2 in a heteromeric complex facilitate their activities to each other by distinct mechanisms. Such a formation of functional heteromeric complex between different MAP3Ks may be advantageous for cells to cope with a wide variety of stimuli by fine regulation of cellular responses.
Evidence
5:
Inferred from Physical InteractionIntAct
The Fas death receptor can activate the Jun NH2-terminal kinase (JNK) pathway through the receptor-associated protein Daxx. Daxx was found to activate the JNK kinase kinase ASK1, and overexpression of a kinase-deficient ASK1 mutant inhibited Fas- and Daxx-induced apoptosis and JNK activation. Fas activation induced Daxx to interact with ASK1, which consequently relieved an inhibitory intramolecular interaction between the amino- and carboxyl-termini of ASK1, activating its kinase activity. The Daxx-ASK1 connection completes a signaling pathway from a cell surface death receptor to kinase cascades that modulate nuclear transcription factors.
Evidence
6:
Inferred from Physical InteractionIntAct
Apoptosis signal-regulating kinase 1 (ASK1) is a critical mediator of apoptotic signaling pathways initiated by a variety of death stimuli. Its activity is tightly controlled by various mechanisms such as covalent modification and protein-protein interaction. One of the proteins that control ASK1 function is 14-3-3zeta, a member of the 14-3-3 protein family. Here, we report that ASK1 is capable of binding to other isoforms of 14-3-3, suggesting that binding ASK1 is a general property of the 14-3-3 family. In support of this notion, mutational analysis revealed that the ASK1/14-3-3 interaction was mediated by the conserved amphipathic groove of 14-3-3 with some residue selectivity. Functionally, expression of various isoforms of 14-3-3 suppressed ASK1-induced apoptosis. To understand how 14-3-3 controls the ASK1 activity, we examined intracellular localization of ASK1 upon 14-3-3 co-expression. We found that 14-3-3 co-expression is correlated with the translocation of ASK1 from the cytoplasm to a perinuclear localization, likely the ER compartment. Consistent with this notion, ASK1(S967A), a 14-3-3 binding defective mutant of ASK, showed no change in intracellular distribution upon 14-3-3 co-expression. These data support a model that 14-3-3 proteins regulate the proapoptotic function of ASK1 in part by controlling its subcellular distribution.
Evidence
7:
Inferred from Physical InteractionUniProtKB
In vivo infection of lymphatic tissues by the human immunodeficiency virus type 1 (HIV-1) leads to enhanced apoptosis, which prominently involves uninfected bystander cells. Increased killing of such bystander cells is mediated in part through Nef induction of Fas ligand (FasL) expression on the surface of the virally infected T cells. The subsequent interaction of FasL with Fas (CD95) displayed on neighbouring cells, including HIV-1-specific cytotoxic T lymphocytes, may lead to bystander cell killing and thus forms an important mechanism of immune evasion. As HIV-1 also enhances Fas expression on virally infected cells, it is unclear how these hosts avoid rapid cell-autonomous apoptosis mediated through cis ligation of Fas by FasL. Here we show that HIV-1 Nef associates with and inhibits apoptosis signal-regulating kinase 1 (ASK1), a serine/threonine kinase that forms a common and key signalling intermediate in the Fas and tumour-necrosis factor-alpha (TNFalpha) death-signalling pathways. The interaction of Nef with ASK1 inhibits both Fas- and TNFalpha-mediated apoptosis, as well as the activation of the downstream c-Jun amino-terminal kinase. Our findings reveal a strategy by which HIV-1 Nef promotes the killing of bystander cells through the induction of FasL, while simultaneously protecting the HIV-1-infected host cell from these same pro-apoptotic signals through its interference with ASK1 function.
Evidence
8:
Inferred from Physical InteractionUniProtKB
The type 1 insulin-like growth factor receptor (IGF-IR) is a receptor-tyrosine kinase that plays a critical role in signaling cell survival and proliferation. IGF-IR binding to its ligand, insulin-like growth factor (IGF-I) activates phosphoinositide 3-kinase (PI3K), promotes cell proliferation by activating the mitogen-activated protein kinase (MAPK) cascade, and blocks apoptosis by inducing the phosphorylation and inhibition of proapoptotic proteins such as BAD. Apoptosis signal-regulating kinase 1 (ASK1) is a MAP kinase kinase kinase (MAPKKK) that is required for c-Jun N-terminal kinase (JNK) and p38 activation in response to Fas and tumor necrosis factor (TNF) receptor stimulation, and for oxidative stress- and TNFalpha-induced apoptosis. The results presented here indicate that ASK1 forms a complex with the IGF-IR and becomes phosphorylated on tyrosine residue(s) in a manner dependent on IGF-IR activity. IGF-IR signaling inhibited ASK1 irrespective of TNFalpha-induced ASK1 activation and resulted in decreased ASK1-dependent JNK1 stimulation. Signaling through IGF-IR rescued cells from ASK1-induced apoptotic cell death in a manner independent of PI3K activity. These results indicate that IGF-IR signaling suppresses the ASK-1-mediated stimulation of JNK/p38 and the induction of programmed cell death. The simultaneous activation of MAP kinases and the inhibition of the stress-activated arm of the cascade by IGF-IR may constitute a potent proliferative signaling system and is possibly a mechanism by which IGF-I can stimulate growth and inhibit cell death in a wide variety of cell types and biological settings.
Evidence
9:
Inferred from Physical InteractionIntAct
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
Evidence
10:
Inferred from Physical InteractionIntAct
Although Daxx (death-associated protein) was first reported to mediate the apoptotic signal from Fas to JNK in the cytoplasm, other data suggested that Daxx is mainly located in the nucleus as a transcriptional regulator. Here, we demonstrated that cellular localization of Daxx could be determined by the relative concentration of a proapoptotic kinase, apoptosis signal-regulating kinase 1 (ASK1) by using immunofluorescence and transcriptional reporter assay. ASK1 sequestered Daxx in the cytoplasm and inhibited the repressive activity of Daxx in transcription. In addition, Daxx was bound to the activated Fas only in the presence of ASK1, accelerating the Fas-mediated apoptosis. These results suggest that Daxx requires ASK1 for its cytoplasmic localization and Fas-mediated signaling. Taken together, we could conclude that ASK1 controls the dual function of Daxx as a transcriptional repressor in the nucleus and as a proapoptotic signal mediator in the cytoplasm.
Evidence
11:
Inferred from Physical InteractionUniProtKB
Apoptosis signal-regulating kinase 1 (ASK1) is a mitogen-activated protein kinase kinase kinase family member that plays a central role in cytokine- and stress-induced apoptosis by activating c-Jun N-terminal kinase and p38 signaling cascades. ASK1-induced apoptotic activity is up-regulated by two cellular factors, Daxx and TRAF2, through direct protein-protein interactions. Daxx and TRAF2 are death receptor-associated proteins in Fas and tumor necrosis factor-alpha pathways, respectively. Recent studies suggest that calcium signaling may regulate ASK1 pathway. Here we report that human D53L1, a member of the tumor protein D52 family involved in cell proliferation and calcium signaling, up-regulates the ASK1-induced apoptosis. The human D53L1 physically interacts with the C-terminal regulatory domain of ASK1 and promotes ASK1-induced apoptotic activity by activating caspase signaling in mammalian cells. In luciferase reporter assays, hD53L1 activates c-Jun N-terminal kinase-mediated transactivation in the presence of ASK1. Expression of hD53L1 enhances autophosphorylation and kinase activity of ASK1 but has no effect on ASK1 oligomerization that is necessary for kinase activity and on binding of ASK1 to MKK6, a downstream factor of ASK1. Taken together, these results suggest that activation of ASK1 by hD53L1 may provide a novel mechanism for ASK1 regulation.
Apoptosis signal-regulating kinase 1 (ASK1) is a MAPKKK family member which activates c-Jun N-terminal kinase (JNK) and p38. In non-stressed cells, ASK1 exists as an inactive complex with the reduced form of thioredoxin. Oxidative stress such as hydrogen peroxide (H2O2) disrupts the ASK1-thioredoxin complex by oxidization of thioredoxin and thereby activates ASK1. The precise mechanism by which ASK1 is activated after its release from thioredoxin is unknown. Here we show that phosphorylation of Thr845 at the activation loop is essential for ASK1 to be activated by H2O2. ASK1 appears to form a silent homo-oligomer through its C-terminal coiled-coil region in non-stressed cells. Following H2O2 treatment, pre-existing ASK1 oligomer undergoes conformational change and creates a new interface within an oligomer, which ultimately leads to trans-autophosphorylation of Thr845. Thus, direct interaction via the coiled-coil region is required for self-scaffolding but not sufficient for activation of ASK1. Importantly, Thr845 of ASK1 can also be trans-phosphorylated by an unidentified Thr845 kinase in response to H2O2 treatment. We propose that this potential Thr845 kinase may be an ignition kinase that triggers Thr845 phosphorylation in oligomerized and activation-competent forms of ASK1.
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
Interacting selectively and non-covalently with a protein kinase, any enzyme that catalyzes the transfer of a phosphate group, usually from ATP, to a protein substrate.
Stress-activated protein kinase (SAPK) pathway-regulating phosphatase 1 (SKRP1) has been identified as a member of the mitogen-activated protein kinase (MAPK) phosphatase (MKP) family that interacts physically with the MAPK kinase (MAPKK) MKK7, a c-Jun N-terminal kinase (JNK) activator, and inactivates the MAPK JNK pathway. Although these findings indicated that SKRP1 contributes to the precise regulation of JNK signaling, it remains to be elucidated how SKRP1 is integrated into this pathway. We report that SKRP1 also plays a scaffold role for the JNK signaling, judged by the following observations. SKRP1 selectively formed the stable complexes with MKK7 but not with MKK4 and biphasically regulated the MKK7 activity and MKK7-induced gene transcription in vivo. Co-precipitation analysis between SKRP1 and MKK7-activating MAPKK kinases (MAPKKKs) revealed that SKRP1 also interacted with the MAPKKK, apoptosis signal-regulating kinase 1 (ASK1), but not with MAP kinase kinase kinase 1 (MEKK1). Consistent with these findings, SKRP1 expression increased the ASK1-MKK7 complexes in a dose-dependent manner and specifically enhanced the activation of MKK7 by ASK1. Thus, our findings are, to our knowledge, the first evidence to show that an MKP also functions as a scaffold protein for the particular MAPK signaling.
Evidence
2:
Inferred from Physical InteractionBHF-UCL
The function of protein phosphatases with EF-hand domains (PPEF) in mammals is not known. Large-scale expression profiling experiments suggest that PPEF expression may correlate with stress protective responses, cell survival, growth, proliferation, or neoplastic transformation. Apoptosis signal regulating kinase-1 (ASK1) is a MAP kinase kinase kinase implicated in cancer, cardiovascular and neurodegenerative diseases. ASK1 is activated by oxidative stress and induces pro-apoptotic or inflammatory signalling, largely via sustained activation of MAP kinases p38 and/or JNK. We identify human PPEF2 as a novel interacting partner and a negative regulator of ASK1. In COS-7 or HEK 293A cells treated with H(2)O(2), expression of PPEF2 abrogated sustained activation of p38 and one of the JNK p46 isoforms, and prevented ASK1-dependent caspase-3 cleavage and activation. PPEF2 efficiently suppressed H(2)O(2)-induced activation of ASK1. Overexpessed as well as endogenous ASK1 co-immunoprecipitated with PPEF2. PPEF2 was considerably more potent both as a suppressor of ASK1 activation and as its interacting partner as compared to protein phosphatase 5 (PP5), a well-known negative regulator of ASK1. PPEF2 was found to form complexes with endogenous Hsp70 and to a lesser extent Hsp90, which are also known interacting partners of PP5. These data identify, for the first time, a possible downstream signalling partner of a mammalian PPEF phosphatase, and suggest that, despite structural divergence, PPEF and PP5 phosphatases may share common interacting partners and functions.
Apoptosis signal-regulating kinase 1 (ASK1) is a mitogen-activated protein kinase kinase kinase family member that plays a central role in cytokine- and stress-induced apoptosis by activating c-Jun N-terminal kinase and p38 signaling cascades. ASK1-induced apoptotic activity is up-regulated by two cellular factors, Daxx and TRAF2, through direct protein-protein interactions. Daxx and TRAF2 are death receptor-associated proteins in Fas and tumor necrosis factor-alpha pathways, respectively. Recent studies suggest that calcium signaling may regulate ASK1 pathway. Here we report that human D53L1, a member of the tumor protein D52 family involved in cell proliferation and calcium signaling, up-regulates the ASK1-induced apoptosis. The human D53L1 physically interacts with the C-terminal regulatory domain of ASK1 and promotes ASK1-induced apoptotic activity by activating caspase signaling in mammalian cells. In luciferase reporter assays, hD53L1 activates c-Jun N-terminal kinase-mediated transactivation in the presence of ASK1. Expression of hD53L1 enhances autophosphorylation and kinase activity of ASK1 but has no effect on ASK1 oligomerization that is necessary for kinase activity and on binding of ASK1 to MKK6, a downstream factor of ASK1. Taken together, these results suggest that activation of ASK1 by hD53L1 may provide a novel mechanism for ASK1 regulation.
Mitogen-activated protein (MAP) kinase cascades are activated in response to various extracellular stimuli, including growth factors and environmental stresses. A MAP kinase kinase kinase (MAPKKK), termed ASK1, was identified that activated two different subgroups of MAP kinase kinases (MAPKK), SEK1 (or MKK4) and MKK3/MAPKK6 (or MKK6), which in turn activated stress-activated protein kinase (SAPK, also known as JNK; c-Jun amino-terminal kinase) and p38 subgroups of MAP kinases, respectively. Overexpression of ASK1 induced apoptotic cell death, and ASK1 was activated in cells treated with tumor necrosis factor-alpha (TNF-alpha). Moreover, TNF-alpha-induced apoptosis was inhibited by a catalytically inactive form of ASK1. ASK1 may be a key element in the mechanism of stress- and cytokine-induced apoptosis.
Stress-activated protein kinase (SAPK) pathway-regulating phosphatase 1 (SKRP1) has been identified as a member of the mitogen-activated protein kinase (MAPK) phosphatase (MKP) family that interacts physically with the MAPK kinase (MAPKK) MKK7, a c-Jun N-terminal kinase (JNK) activator, and inactivates the MAPK JNK pathway. Although these findings indicated that SKRP1 contributes to the precise regulation of JNK signaling, it remains to be elucidated how SKRP1 is integrated into this pathway. We report that SKRP1 also plays a scaffold role for the JNK signaling, judged by the following observations. SKRP1 selectively formed the stable complexes with MKK7 but not with MKK4 and biphasically regulated the MKK7 activity and MKK7-induced gene transcription in vivo. Co-precipitation analysis between SKRP1 and MKK7-activating MAPKK kinases (MAPKKKs) revealed that SKRP1 also interacted with the MAPKKK, apoptosis signal-regulating kinase 1 (ASK1), but not with MAP kinase kinase kinase 1 (MEKK1). Consistent with these findings, SKRP1 expression increased the ASK1-MKK7 complexes in a dose-dependent manner and specifically enhanced the activation of MKK7 by ASK1. Thus, our findings are, to our knowledge, the first evidence to show that an MKP also functions as a scaffold protein for the particular MAPK signaling.
A series of molecular signals which triggers the apoptotic death of a cell. The pathway starts with reception of a signal, and ends when the execution phase of apoptosis is triggered.
Mitogen-activated protein (MAP) kinase cascades are activated in response to various extracellular stimuli, including growth factors and environmental stresses. A MAP kinase kinase kinase (MAPKKK), termed ASK1, was identified that activated two different subgroups of MAP kinase kinases (MAPKK), SEK1 (or MKK4) and MKK3/MAPKK6 (or MKK6), which in turn activated stress-activated protein kinase (SAPK, also known as JNK; c-Jun amino-terminal kinase) and p38 subgroups of MAP kinases, respectively. Overexpression of ASK1 induced apoptotic cell death, and ASK1 was activated in cells treated with tumor necrosis factor-alpha (TNF-alpha). Moreover, TNF-alpha-induced apoptosis was inhibited by a catalytically inactive form of ASK1. ASK1 may be a key element in the mechanism of stress- and cytokine-induced apoptosis.
Any process that results in a change in state or activity of a cell (in terms of movement, secretion, enzyme production, gene expression, etc.) as a result of a hydrogen peroxide (H2O2) stimulus.
The function of protein phosphatases with EF-hand domains (PPEF) in mammals is not known. Large-scale expression profiling experiments suggest that PPEF expression may correlate with stress protective responses, cell survival, growth, proliferation, or neoplastic transformation. Apoptosis signal regulating kinase-1 (ASK1) is a MAP kinase kinase kinase implicated in cancer, cardiovascular and neurodegenerative diseases. ASK1 is activated by oxidative stress and induces pro-apoptotic or inflammatory signalling, largely via sustained activation of MAP kinases p38 and/or JNK. We identify human PPEF2 as a novel interacting partner and a negative regulator of ASK1. In COS-7 or HEK 293A cells treated with H(2)O(2), expression of PPEF2 abrogated sustained activation of p38 and one of the JNK p46 isoforms, and prevented ASK1-dependent caspase-3 cleavage and activation. PPEF2 efficiently suppressed H(2)O(2)-induced activation of ASK1. Overexpessed as well as endogenous ASK1 co-immunoprecipitated with PPEF2. PPEF2 was considerably more potent both as a suppressor of ASK1 activation and as its interacting partner as compared to protein phosphatase 5 (PP5), a well-known negative regulator of ASK1. PPEF2 was found to form complexes with endogenous Hsp70 and to a lesser extent Hsp90, which are also known interacting partners of PP5. These data identify, for the first time, a possible downstream signalling partner of a mammalian PPEF phosphatase, and suggest that, despite structural divergence, PPEF and PP5 phosphatases may share common interacting partners and functions.
Programmed cell death induced by hydrogen peroxide. Programmed cell death is the cell death resulting from activation of endogenous cellular processes.
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
Innate immune responses are defense responses mediated by germline encoded components that directly recognize components of potential pathogens.
IEAUniProtKB KW
Intrinsic apoptotic signaling pathway in response to oxidative stressdefinition[GO:0008631]
A series of molecular signals in which an intracellular signal is conveyed to trigger the apoptotic death of a cell. The pathway is induced in response to oxidative stress, a state often resulting from exposure to high levels of reactive oxygen species, and ends when the execution phase of apoptosis is triggered.
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
An intracellular protein kinase cascade containing at least a JNK (a MAPK), a JNKK (a MAPKK) and a JUN3K (a MAP3K). The cascade can also contain two additional tiers: the upstream MAP4K and the downstream MAP Kinase-activated kinase (MAPKAPK). The kinases in each tier phosphorylate and activate the kinases in the downstream tier to transmit a signal within a cell.
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
An intracellular protein kinase cascade containing at least a MAPK, a MAPKK and a MAP3K. The cascade can also contain two additional tiers: the upstream MAP4K and the downstream MAP Kinase-activated kinase (MAPKAPK). The kinases in each tier phosphorylate and activate the kinases in the downstream tier to transmit a signal within a cell.
Apoptosis signal-regulating kinase (ASK) 1 is a mitogen-activated protein kinase kinase kinase (MAP3K) in the c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase pathways that play multiple important roles in cytokine and stress responses. Here we show that ASK2, a highly related serine/threonine kinase to ASK1, also functions as a MAP3K only in a heteromeric complex with ASK1. We found that endogenous ASK2 was constitutively degraded in ASK1-deficient cells, suggesting that ASK1 is required for the stability of ASK2. ASK2 in a heteromeric complex with a kinase-negative mutant of ASK1 (ASK1-KN) effectively activated MAP2K and was more competent to respond to oxidative stress than ASK2 alone. Knockdown of ASK2 revealed that ASK2 was required for oxidative stress-induced JNK activation. These results suggest that ASK2 forms a functional MAP3K complex with ASK1, in which ASK1 supports the stability and the active configuration of ASK2. Moreover, ASK2 was found to activate ASK1 by direct phosphorylation, suggesting that ASK1 and ASK2 in a heteromeric complex facilitate their activities to each other by distinct mechanisms. Such a formation of functional heteromeric complex between different MAP3Ks may be advantageous for cells to cope with a wide variety of stimuli by fine regulation of cellular responses.
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
A zinc finger protein, ZPR9, has been identified as a physiological substrate of murine protein serine/threonine kinase 38 (MPK38), which is involved in various cellular responses, including the cell cycle, apoptosis, embryonic development, and oncogenesis. Here, ZPR9 was found to physically interact with apoptosis signal-regulating kinase 1 (ASK1) through a disulfide linkage involving Cys(1351) and Cys(1360) of ASK1 and Cys(305) and Cys(308) of ZPR9. ASK1 directly phosphorylated ZPR9 at Ser(314) and Thr(318), suggesting that ZPR9 can act as an ASK1 substrate. Ectopic expression of wild-type ZPR9, but not an S314A/T318A mutant, stimulated ASK1 kinase activity and positively regulated ASK1-mediated signaling to both JNK and p38 kinases by destabilizing complex formation between ASK1 and its negative regulators, Trx and 14-3-3, or by increasing complex formation between ASK1 and its substrate MKK3. ZPR9 functionally stimulated ASK1-induced AP-1 transcriptional activity as well as H(2)O(2)-mediated apoptosis in a phosphorylation-dependent manner. ASK1-mediated phosphorylation of ZPR9 at Ser(314) and Thr(318) was also responsible for ZPR9-induced apoptosis. Moreover, ZPR9 inhibited PDK1-mediated signaling through ASK1 activation. These results suggest that ZPR9 functions as a novel positive regulator of ASK1.
The function of protein phosphatases with EF-hand domains (PPEF) in mammals is not known. Large-scale expression profiling experiments suggest that PPEF expression may correlate with stress protective responses, cell survival, growth, proliferation, or neoplastic transformation. Apoptosis signal regulating kinase-1 (ASK1) is a MAP kinase kinase kinase implicated in cancer, cardiovascular and neurodegenerative diseases. ASK1 is activated by oxidative stress and induces pro-apoptotic or inflammatory signalling, largely via sustained activation of MAP kinases p38 and/or JNK. We identify human PPEF2 as a novel interacting partner and a negative regulator of ASK1. In COS-7 or HEK 293A cells treated with H(2)O(2), expression of PPEF2 abrogated sustained activation of p38 and one of the JNK p46 isoforms, and prevented ASK1-dependent caspase-3 cleavage and activation. PPEF2 efficiently suppressed H(2)O(2)-induced activation of ASK1. Overexpessed as well as endogenous ASK1 co-immunoprecipitated with PPEF2. PPEF2 was considerably more potent both as a suppressor of ASK1 activation and as its interacting partner as compared to protein phosphatase 5 (PP5), a well-known negative regulator of ASK1. PPEF2 was found to form complexes with endogenous Hsp70 and to a lesser extent Hsp90, which are also known interacting partners of PP5. These data identify, for the first time, a possible downstream signalling partner of a mammalian PPEF phosphatase, and suggest that, despite structural divergence, PPEF and PP5 phosphatases may share common interacting partners and functions.
Activated by various stressors, including oxidative stress, endoplasmic reticulum stress, and calcium overload, as well as by receptor-mediated inflammatory signals, such as the tumor necrosis factor (TNF) and lipopolysaccharide (LPS). Homophilic association of MAP3K5/ASK1 through the C-terminal coiled-coil domains and the heteromeric complex formation of MAP3K5/ASK1 with the reduced form of thioredoxin (TXN), constitutes an inactive form of the kinase. Upon ROS-induced dissociation of TXN from MAP3K5/ASK1, TRAF2 and TRAF6 are reciprocally recruited to MAP3K5/ASK1 and form the active MAP3K5/ASK1 signalosome, in which TRAF2 and TRAF6 appear to facilitate the active configuration of MAP3K5/ASK1. MAP3K5/ASK1 activity is also regulated through several phosphorylation and dephosphorylation events. Thr-838 is an activating phosphorylation site that is autophosphorylated and phosphorylated by MAP3K6/ASK2 and dephosphorylated by PPP5C. Ser-83 and Ser-1033 are inactivating phosphorylation sites, the former of which is phosphorylated by AKT1 and AKT2. Phosphorylation of Ser-966 induces association of MAP3K5/ASK1 with the 14-3-3 family proteins, which suppresses MAP3K5/ASK1 activity. Calcium/calmodulin-activated protein phosphatase calcineurin (PPP3CA) has been shown to directly dephosphorylate this site. SOCS1 binds to ASK1 by recognizing phosphorylation of Tyr-718 and induces MAP3K5/ASK1 degradation in endothelial cells. Also dephosphorylated and activated by PGAM5. Contains an N-terminal autoinhibitory domain.
The type 1 insulin-like growth factor receptor (IGF-IR) is a receptor-tyrosine kinase that plays a critical role in signaling cell survival and proliferation. IGF-IR binding to its ligand, insulin-like growth factor (IGF-I) activates phosphoinositide 3-kinase (PI3K), promotes cell proliferation by activating the mitogen-activated protein kinase (MAPK) cascade, and blocks apoptosis by inducing the phosphorylation and inhibition of proapoptotic proteins such as BAD. Apoptosis signal-regulating kinase 1 (ASK1) is a MAP kinase kinase kinase (MAPKKK) that is required for c-Jun N-terminal kinase (JNK) and p38 activation in response to Fas and tumor necrosis factor (TNF) receptor stimulation, and for oxidative stress- and TNFalpha-induced apoptosis. The results presented here indicate that ASK1 forms a complex with the IGF-IR and becomes phosphorylated on tyrosine residue(s) in a manner dependent on IGF-IR activity. IGF-IR signaling inhibited ASK1 irrespective of TNFalpha-induced ASK1 activation and resulted in decreased ASK1-dependent JNK1 stimulation. Signaling through IGF-IR rescued cells from ASK1-induced apoptotic cell death in a manner independent of PI3K activity. These results indicate that IGF-IR signaling suppresses the ASK-1-mediated stimulation of JNK/p38 and the induction of programmed cell death. The simultaneous activation of MAP kinases and the inhibition of the stress-activated arm of the cascade by IGF-IR may constitute a potent proliferative signaling system and is possibly a mechanism by which IGF-I can stimulate growth and inhibit cell death in a wide variety of cell types and biological settings.
Tumor necrosis factor (TNF)-induced activation of the c-jun N-terminal kinase (JNK, also known as SAPK; stress-activated protein kinase) requires TNF receptor-associated factor 2 (TRAF2). The apoptosis signal-regulating kinase 1 (ASK1) is activated by TNF and stimulates JNK activation. Here we show that ASK1 interacts with members of the TRAF family and is activated by TRAF2, TRAF5, and TRAF6 overexpression. A truncated derivative of TRAF2, which inhibits JNK activation by TNF, blocks TNF-induced ASK1 activation. A catalytically inactive mutant of ASK1 is a dominant-negative inhibitor of TNF- and TRAF2-induced JNK activation. In untransfected mammalian cells, ASK1 rapidly associates with TRAF2 in a TNF-dependent manner. Thus, ASK1 is a mediator of TRAF2-induced JNK activation.
Cisplatin and its analogues have been widely used for treatment of human cancer. However, most patients eventually develop resistance to treatment through a mechanism that remains obscure. Previously, we found that AKT2 is frequently overexpressed and/or activated in human ovarian and breast cancers. Here we demonstrate that constitutively active AKT2 renders cisplatin-sensitive A2780S ovarian cancer cells resistant to cisplatin, whereas phosphatidylinositol 3-kinase inhibitor or dominant negative AKT2 sensitizes A2780S and cisplatin-resistant A2780CP cells to cisplatin-induced apoptosis through regulation of the ASK1/JNK/p38 pathway. AKT2 interacts with and phosphorylates ASK1 at Ser-83 resulting in inhibition of its kinase activity. Accordingly, activated AKT2 blocked signaling down-stream of ASK1, including activation of JNK and p38 and the conversion of Bax to its active conformation. Expression of nonphosphorylatable ASK1-S83A overrode the AKT2-inhibited JNK/p38 activity and Bax conformational changes, whereas phosphomimic ASK1-S83D inhibited the effects of cisplatin on JNK/p38 and Bax. Cisplatin-induced Bax conformation change was inhibited by inhibitors or dominant negative forms of JNK and p38. In conclusion, our data indicate that AKT2 inhibits cisplatin-induced JNK/p38 and Bax activation through phosphorylation of ASK1 and thus, plays an important role in chemoresistance. Further, regulation of the ASK1/JNK/p38/Bax pathway by AKT2 provides a new mechanism contributing to its antiapoptotic effects.
The mammalian mitogen-activated protein (MAP) kinase kinase kinase apoptosis signal-regulating kinase 1 (ASK1) is a pivotal component in cytokine- and stress-induced apoptosis. It also regulates cell differentiation and survival through p38 MAP kinase activation. Here we show that Ca2+ signalling regulates the ASK1-p38 MAP kinase cascade. Ca2+ influx evoked by membrane depolarization in primary neurons and synaptosomes induced activation of p38, which was impaired in those derived from ASK1-deficient mice. Ca2+/calmodulin-dependent protein kinase type II (CaMKII) activated ASK1 by phosphorylation. Moreover, p38 activation induced by the expression of constitutively active CaMKII required endogenous ASK1. Thus, ASK1 is a critical intermediate of Ca2+ signalling between CaMKII and p38 MAP kinase.
Apoptosis signal-regulating kinase 1 (ASK1) is a MAP kinase kinase kinase (MAPKKK) that activates the JNK and p38 MAP kinase cascades and is activated in response to oxidative stress such as hydrogen peroxide (H(2)O(2)). A yeast two-hybrid screening identified a serine/threonine protein phosphatase 5 (PP5) as a binding partner of ASK1. PP5 directly dephosphorylated an essential phospho-threonine residue within the kinase domain of ASK1 and thereby inactivated ASK1 activity in vitro and in vivo. The interaction between PP5 and ASK1 was induced by H(2)O(2) treatment and was followed by the decrease in ASK1 activity. PP5 inhibited not only H(2)O(2)-induced sustained activation of ASK1 but also ASK1-dependent apoptosis. Thus, PP5 appears to act as a physiological inhibitor of ASK1-JNK/p38 pathways by negative feedback.
Apoptosis signal-regulating kinase 1 (ASK1) is a serine/threonine kinase that mediates cell stress signaling initiated by diverse stimuli, such as H(2)O(2) and TNF alpha. Owing to its critical role in promoting apoptosis, ASK1 activity is highly controlled in cells. Phosphorylation of ASK1 at Thr-845 has been correlated with its activation, while phosphorylation at Ser-967 negatively controls its death promoting activity. Here, we report the identification of a novel phosphorylation site at Ser-1034 in the C-terminal regulatory domain of ASK1. Mutating Ser-1034 to an unphosphorylatable Ala led to increased catalytic activity of ASK1 and enhanced proapoptotic function of ASK1. Thus, the proapoptotic function of ASK1 is suppressed in part by phosphorylation at its C-terminal regulatory domain, which may couple upstream survival kinases to the death regulatory machinery.
Apoptosis signal-regulating kinase (ASK) 1 is a mitogen-activated protein kinase kinase kinase (MAP3K) in the c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase pathways that play multiple important roles in cytokine and stress responses. Here we show that ASK2, a highly related serine/threonine kinase to ASK1, also functions as a MAP3K only in a heteromeric complex with ASK1. We found that endogenous ASK2 was constitutively degraded in ASK1-deficient cells, suggesting that ASK1 is required for the stability of ASK2. ASK2 in a heteromeric complex with a kinase-negative mutant of ASK1 (ASK1-KN) effectively activated MAP2K and was more competent to respond to oxidative stress than ASK2 alone. Knockdown of ASK2 revealed that ASK2 was required for oxidative stress-induced JNK activation. These results suggest that ASK2 forms a functional MAP3K complex with ASK1, in which ASK1 supports the stability and the active configuration of ASK2. Moreover, ASK2 was found to activate ASK1 by direct phosphorylation, suggesting that ASK1 and ASK2 in a heteromeric complex facilitate their activities to each other by distinct mechanisms. Such a formation of functional heteromeric complex between different MAP3Ks may be advantageous for cells to cope with a wide variety of stimuli by fine regulation of cellular responses.
Apoptosis signal-regulating kinase (ASK) 1 was recently identified as a mitogen-activated protein (MAP) kinase kinase kinase which activates the c-Jun N-terminal kinase (JNK) and p38 MAP kinase pathways and is required for tumor necrosis factor (TNF)-alpha-induced apoptosis; however, the mechanism regulating ASK1 activity is unknown. Through genetic screening for ASK1-binding proteins, thioredoxin (Trx), a reduction/oxidation (redox)-regulatory protein thought to have anti-apoptotic effects, was identified as an interacting partner of ASK1. Trx associated with the N-terminal portion of ASK1 in vitro and in vivo. Expression of Trx inhibited ASK1 kinase activity and the subsequent ASK1-dependent apoptosis. Treatment of cells with N-acetyl-L-cysteine also inhibited serum withdrawal-, TNF-alpha- and hydrogen peroxide-induced activation of ASK1 as well as apoptosis. The interaction between Trx and ASK1 was found to be highly dependent on the redox status of Trx. Moreover, inhibition of Trx resulted in activation of endogenous ASK1 activity, suggesting that Trx is a physiological inhibitor of ASK1. The evidence that Trx is a negative regulator of ASK1 suggests possible mechanisms for redox regulation of the apoptosis signal transduction pathway as well as the effects of antioxidants against cytokine- and stress-induced apoptosis.
We have previously shown that ASK1 undergoes ubiquitination and degradation in resting endothelial cells (EC) and that proinflammatory cytokine tumor necrosis factor (TNF) induces deubiquitination and stabilization, leading to ASK1 activation. However, the mechanism for the regulation of ASK1 stability is not known. In the present study, we have shown that SOCS1, a member of suppressor of cytokine signaling, induces ASK1 degradation. SOCS1 was constitutively expressed in EC and formed a labile complex with ASK1 that can be stabilized by proteasomal inhibitors. The phosphotyrosine-binding SH2 domain of SOCS1 was critical for its association with ASK1. Thus a SOCS1 mutant defective in phosphotyrosine binding failed to bind to and induce ASK1 degradation. Phosphotyrosine of ASK1 was induced in response to growth factors, and TNF induced dephosphorylation and dissociation of ASK1 from SOCS1. ASK1 with a mutation at Tyr-718 diminished the binding to SOCS1, suggesting that the phosphotyrosine-718 of ASK1 is critical for SOCS1 binding. Moreover, ASK1 expression and activity were up-regulated in SOCS1-deficient mice and derived EC, resulting in enhanced TNF-induced activation of JNK, expression of proinflammatory molecules, and apoptotic responses. We concluded that SOCS1 functions as a negative regulator in TNF-induced inflammation in EC, in part, by inducing ASK1 degradation.
The Akt family of serine/threonine-directed kinases promotes cellular survival in part by phosphorylating and inhibiting death-inducing proteins. Here we describe a novel functional interaction between Akt and apoptosis signal-regulating kinase 1 (ASK1), a mitogen-activated protein kinase kinase kinase. Akt decreased ASK1 kinase activity stimulated by both oxidative stress and overexpression in 293 cells by phosphorylating a consensus Akt site at serine 83 of ASK1. Activation of the phosphoinositide 3-kinase (PI3-K)/Akt pathway also inhibited the serum deprivation-induced activity of endogenous ASK1 in L929 cells. An association between Akt and ASK1 was detected in cells by coimmunoprecipitation. Phosphorylation by Akt inhibited ASK1-mediated c-Jun N-terminal kinase and activating transcription factor 2 activities in intact cells. Finally, activation of the PI3-K/Akt pathway reduced apoptosis induced by ASK1 in a manner dependent on phosphorylation of serine 83 of ASK1. These results provide the first direct link between Akt and the family of stress-activated kinases.
Proc. Natl. Acad. Sci. U.S.A. 96, 8511-8515 (1999)[PubMed:10411906]
Apoptosis signal-regulating kinase 1 (ASK1) is a pivotal component of a signaling pathway induced by many death stimuli, including tumor necrosis factor alpha, Fas, and the anticancer drugs cisplatin and paclitaxel. Here we report that ASK1 proapoptotic activity is antagonized by association with 14-3-3 proteins. We found that ASK1 specifically bound 14-3-3 proteins via a site involving Ser-967 of ASK1. Interestingly, overexpression of 14-3-3 in HeLa cells blocked ASK1-induced apoptosis whereas disruption of the ASK1/14-3-3 interaction dramatically accelerated ASK1-induced cell death. Targeting of ASK1 by a 14-3-3-mediated survival pathway may provide a novel mechanism for the suppression of apoptosis.
The stress-activated protein kinases (SAPKs, also called c-Jun NH(2)-terminal kinases) and the p38s, two mitogen-activated protein kinase (MAPK) subgroups activated by cytokines of the tumor necrosis factor (TNF) family, are pivotal to the de novo gene expression elicited as part of the inflammatory response. Apoptosis signal-regulating kinase 1 (ASK1) is a MAPK kinase kinase (MAP3K) that activates both the SAPKs and p38s in vivo. Here we show that TNF receptor (TNFR) associated factor 2 (TRAF2), an adapter protein that couples TNFRs to the SAPKs and p38s, can activate ASK1 in vivo and can interact in vivo with the amino- and carboxyl-terminal noncatalytic domains of the ASK1 polypeptide. Expression of the amino-terminal noncatalytic domain of ASK1 can inhibit TNF and TRAF2 activation of SAPK. TNF can stimulate the production of reactive oxygen species (ROS), and the redox-sensing enzyme thioredoxin (Trx) is an endogenous inhibitor of ASK1. We also show that expression of TRAF2 fosters the production of ROS in transfected cells. We demonstrate that Trx significantly inhibits TRAF2 activation of SAPK and blocks the ASK1-TRAF2 interaction in a reaction reversed by oxidants. Finally, the mechanism of ASK1 activation involves, in part, homo-oligomerization. We show that expression of ASK1 with TRAF2 enhances in vivo ASK1 homo-oligomerization in a manner dependent, in part, upon the TRAF2 RING effector domain and the generation of ROS. Thus, activation of ASK1 by TNF requires the ROS-mediated dissociation of Trx possibly followed by the binding of TRAF2 and consequent ASK1 homo-oligomerization.
Apoptosis signal-regulating kinase 1 (ASK1) is a MAPKKK family member which activates c-Jun N-terminal kinase (JNK) and p38. In non-stressed cells, ASK1 exists as an inactive complex with the reduced form of thioredoxin. Oxidative stress such as hydrogen peroxide (H2O2) disrupts the ASK1-thioredoxin complex by oxidization of thioredoxin and thereby activates ASK1. The precise mechanism by which ASK1 is activated after its release from thioredoxin is unknown. Here we show that phosphorylation of Thr845 at the activation loop is essential for ASK1 to be activated by H2O2. ASK1 appears to form a silent homo-oligomer through its C-terminal coiled-coil region in non-stressed cells. Following H2O2 treatment, pre-existing ASK1 oligomer undergoes conformational change and creates a new interface within an oligomer, which ultimately leads to trans-autophosphorylation of Thr845. Thus, direct interaction via the coiled-coil region is required for self-scaffolding but not sufficient for activation of ASK1. Importantly, Thr845 of ASK1 can also be trans-phosphorylated by an unidentified Thr845 kinase in response to H2O2 treatment. We propose that this potential Thr845 kinase may be an ignition kinase that triggers Thr845 phosphorylation in oligomerized and activation-competent forms of ASK1.
Oxidative stress has been indicated in a variety of pathological processes such as atherosclerosis, diabetes, and neurodegenerative diseases. Understanding how intracellular signaling pathways respond to oxidative insults such as hydrogen peroxide (H(2)O(2)) would have significant therapeutic implications. Recent genetic studies have placed apoptosis signal-regulating kinase 1 (ASK1) in a pivotal position in transmitting H(2)O(2)-initiated signals. How ASK1 is activated by H(2)O(2), though, remains a subject of intense investigation. Here we report a mechanism by which H(2)O(2) induces ASK1 activation through dynamic control of its phosphorylation at serine 967. We found that treatment of COS7 cells with H(2)O(2) triggers dephosphorylation of Ser-967 through an okadaic acid-sensitive phosphatase, resulting in dissociation of the ASK1.14-3-3 complex with concomitant increase of ASK1 catalytic activity and ASK1-mediated activation of JNK and p38 pathways.
Protein involved in apoptotic programmed cell death. Apoptosis is characterized by cell morphological changes, including blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation and chromosomal DNA fragmentation, and eventually death. Unlike necrosis, apoptosis produces cell fragments, called apoptotic bodies, that phagocytic cells are able to engulf and quickly remove before the contents of the cell can spill out onto surrounding cells and cause damage. In general, apoptosis confers advantages during an organism's life cycle.
Viral protein involved in a direct and specific interaction with a host macromolecule. Viruses interact with many cellular pathways to achieve their replication cycle. Entry into the host cell, transport to the viral replication sites or viral budding are all steps that require interaction between the host and the virus. Additionally, the evasion from the host immune response requires a lot of viral proteins to associate with and inhibit cellular proteins with antiviral functions.
Protein involved in immunity, any immune system process that functions in the response of an organism to a potential internal or invasive threat. The vertebrate immune system is formed by the innate immune system (composed of phagocytes, complement, antimicrobial peptides, etc) and by the adaptive immune system which consists of T- and B- lymphocytes.
Protein involved in innate immunity, an inborn defense mechanism used by organisms to defend themselves against invasion by pathogens (bacteria, fungi, viruses, etc.). Initially discovered in insects which are devoid of an adaptive immune system and rely only on innate immune reactions for their defense, this immediate response accomplishes many activities including recognition and effector functions. Recognition is mediated by broad specificity, pattern recognition, receptors which recognize many related molecular structures (e.g. polysaccharides, polynucleotides) present in microorganisms but not found in the host. The innate responses include the release of antimicrobial peptides, production of cytokines, acute- phase proteins, complement. Although many different innate immune mechanisms are deployed for host defence, a unifying theme of innate immunity is the use of germline-encoded pattern recognition receptors for pathogens or damaged self components, such as the Toll-like receptors, nucleotide-binding domain leucine-rich repeat (LRR)- containing receptors, retinoic acid-inducible gene I-like RNA helicases and C-type lectin receptors.
Protein involved in the response to stress, a change in state or activity of a cell or an organism (in terms of movement, secretion, enzyme production, gene expression, etc.) as a result of some stressful conditions. The stress is usually, but not necessarily, exogenous (e.g. temperature, humidity, ionizing radiation, hypertonicity, amino acid deprivation).
Protein which catalyzes the phosphorylation of serine or threonine residues on target proteins by using ATP as phosphate donor. Such phosphorylation may cause changes in the function of the target protein. Protein kinases share a conserved catalytic core common to both serine/ threonine and tyrosine protein kinases.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
My favorites 
My labels 
Login
