Oxidative Stress-Induced Diseases via the ASK1 Signaling Pathway

Apoptosis signal-regulating kinase 1 (ASK1) is a mitogen-activated protein kinase (MAPK) kinase kinase that activates the downstream MAPKs, c-Jun N-terminal kinase (JNK) and p38. ASK1 is activated by various types of stress, such as oxidative stress, endoplasmic reticulum stress, and infection, and regulates various cellular functions. Recently, it has been reported that ASK1 is associated with various diseases induced by oxidative stress. In this review, we introduce recent findings of the regulatory mechanisms of ASK1 and the oxidative stress-induced diseases mediated by the ASK1 signaling pathway

The roles of ASK family proteins in stress responses and diseases

Apoptosis signal-regulating kinase 1 (ASK1) is a member of the mitogen-activated protein kinase kinase kinase family, which activates c-Jun N-terminal kinase and p38 in response to a diverse array of stresses such as oxidative stress, endoplasmic reticulum stress and calcium influx. In the past decade, various regulatory mechanisms of ASK1 have been elucidated, including its oxidative stress-dependent activation. Recently, it has emerged that ASK family proteins play key roles in cancer, cardiovascular diseases and neurodegenerative diseases. In this review, we summarize the recent findings on ASK family proteins and their implications in various diseases

Serine 58 of 14-3-3ζ Is a molecular switch regulating ASK1 and oxidant stress-induced cell death

Oxidant stress is a ubiquitous stressor with negative impacts on multiple cell types. ASK1 is a central mediator of oxidant injury, but while mechanisms of its inhibition, such as sequestration by 14-3-3 proteins and thioredoxin, have been identified, mechanisms of activation have remained obscure and the signaling pathways regulating this are not clear. Here, we report that phosphorylation of 14-3-3ζ at serine 58 (S58) is dynamically regulated in the cell and that the phosphorylation status of S58 is a critical factor regulating oxidant stress-induced cell death. Phosphorylation of S58 releases ASK1 from 14-3-3ζ, and ASK1 then activates stress-activated protein kinases, leading to cell death. While several members of the mammalian sterile 20 (Mst) family of kinases can phosphorylate S58 when overexpressed, we identify Ste20/oxidant stress response kinase 1 (SOK-1), an Mst family member known to be activated by oxidant stress, as a central endogenous regulator of S58 phosphorylation and thereby of ASK1-mediated cell death. Our findings identify a novel pathway that regulates ASK1 activation and oxidant stress-induced cell death

ASK1-p38-NR4A2 Axis in H2O2-induced Necrosis

Background: The molecular mechanisms of p38 MAPK-mediated necrosis currently have not been well elucidated. Results: During oxidative stress, NR4A2 is phosphorylated and translocated into the cytosol in an ASK1-p38-dependent manner, which ultimately leads to the promotion of necrosis. Conclusion: ASK1-p38 MAPK pathway-dependent phosphorylation and subsequent cytoplasmic translocation of NR4A2 promote oxidative stress-induced necrosis. Significance: We found a novel intracellular signaling pathway that regulates oxidative stress-induced and p38-mediated necrosis.p38 mitogen-activated protein kinases (MAPKs) play important roles in various cellular stress responses, including cell death, which is roughly categorized into apoptosis and necrosis. Although p38 signaling has been extensively studied, the molecular mechanisms of p38-mediated cell death are unclear. ASK1 is a stress-responsive MAP3K that acts as an upstream kinase of p38 and is activated by various stresses, such as oxidative stress. Here, we show that NR4A2, a member of the NR4A nuclear receptor family, acts as a necrosis promoter downstream of ASK1-p38 pathway during oxidative stress. Although NR4A2 is well known as a nucleus-localized transcription factor, we found that it is translocated into the cytosol after phosphorylation by p38. Because the phosphorylation site mutants of NR4A2 cannot rescue the cell death-promoting activity, ASK1-p38 pathway-dependent phosphorylation and subsequent cytoplasmic translocation of NR4A2 may be required for oxidative stress-induced cell death. In addition, NR4A2-mediated cell death does not depend on caspases and receptor-interacting protein 1 (RIP1)-RIP3 complex, suggesting that NR4A2 promotes an RIP kinase-independent necrotic type of cell death. Our findings may enable a more precise understanding of molecular mechanisms that regulate oxidative stress-induced and p38-mediated necrosis

KLHDC10 Activates ASK1 by Suppressing PP5

Reactive oxygen species (ROS)-induced activation of Apoptosis signal-regulating kinase 1 (ASK1) plays crucial roles in oxidative stress-mediated cell death through the activation of the JNK and p38 MAPK pathways. However, the regulatory mechanism of ASK1 in the oxidative stress response remains to be elucidated. Here, we identified the kelch repeat protein, Slim, as an activator of ASK1 through a Drosophila misexpression screen. We also performed a proteomics screen and revealed that Kelch domain containing 10 (KLHDC10), a mammalian ortholog of Slim, interacted with Protein phosphatase 5 (PP5), which has been shown to inactivate ASK1 in response to ROS. KLHDC10 bound to the phosphatase domain of PP5 and suppressed its phosphatase activity. Moreover, KLHDC10 was required for H2O2-induced sustained activation of ASK1 and cell death in Neuro2A cells. These findings suggest that Slim/KLHDC10 is an activator of ASK1, contributing to oxidative stress-induced cell death through the suppression of PP5

ASK1-dependent recruitment and activation of macrophages induce hair growth in skin wounds

Apoptosis signal-regulating kinase 1 (ASK1) is a member of the mitogen-activated protein 3-kinase family that activates both c-Jun NH2-terminal kinase and p38 pathways in response to inflammatory cytokines and physicochemical stress. We report that ASK1 deficiency in mice results in dramatic retardation of wounding-induced hair regrowth in skin. Oligonucleotide microarray analysis revealed that expression of several chemotactic and activating factors for macrophages, as well as several macrophage-specific marker genes, was reduced in the skin wound area of ASK1-deficient mice. Intracutaneous transplantation of cytokine-activated bone marrow-derived macrophages strongly induced hair growth in both wild-type and ASK1-deficient mice. These findings indicate that ASK1 is required for wounding-induced infiltration and activation of macrophages, which play central roles in inflammation-dependent hair regrowth in skin

Functional cooperation between ASK1 and p21Waf1/Cip1 in the balance of cell-cycle arrest, cell death and tumorigenesis of stressed keratinocytes

Both CDKN1A (p21 Waf1/Cip1) and Apoptosis signal-regulating kinase 1 (ASK1) play important roles in tumorigenesis. The role of p21 Waf1/Cip1 in attenuating ASK1-induced apoptosis by various stress conditions is well established. However, how ASK1 and p21 Waf1/Cip1 functionally interact during tumorigenesis is still unclear. To address this aspect, we crossed ASK1 knockout (ASK1KO) mice with p21 Waf1/Cip1 knockout (p21KO) mice to compare single and double-mutant mice. We observed that deletion of p21 Waf1/Cip1 leads to increased keratinocyte proliferation but also increased cell death. This is mechanistically linked to the ASK1 axis-induced apoptosis, including p38 and PARP. Indeed, deletion of ASK1 does not alter the proliferation but decreases the apoptosis of p21KO keratinocytes. To analyze as this interaction might affect skin carcinogenesis, we investigated the response of ASK1KO and p21KO mice to DMBA/TPA-induced tumorigenesis. Here we show that while endogenous ASK1 is dispensable for skin homeostasis, ASK1KO mice are resistant to DMBA/TPA-induced tumorigenesis. However, we found that epidermis lacking both p21 and ASK1 reacquires increased sensitivity to DMBA/TPA-induced tumorigenesis. We demonstrate that apoptosis and cell-cycle progression in p21KO keratinocytes are uncoupled in the absence of ASK1. These data support the model that a critical event ensuring the balance between cell death, cell-cycle arrest, and successful divisions in keratinocytes during stress conditions is the p21-dependent ASK1 inactivation

ASK1 promotes the contact hypersensitivity response through IL-17 production

Contact hypersensitivity (CHS) is a form of delayed-type hypersensitivity triggered by the response to reactive haptens (sensitization) and subsequent challenge (elicitation). Here, we show that ASK1 promotes CHS and that suppression of ASK1 during the elicitation phase is sufficient to attenuate CHS. ASK1 knockout (KO) mice exhibited impaired 2,4-dinitrofluorobenzene (DNFB)-induced CHS. The suppression of ASK1 activity during the elicitation phase through a chemical genetic approach or a specific inhibitory compound significantly reduced the CHS response to a level similar to that observed in ASK1 KO mice. The reduced response was concomitant with the strong inhibition of production of IL-17, a cytokine that plays an important role in CHS and other inflammatory diseases, from sensitized lymph node cells. These results suggest that ASK1 is relevant to the overall CHS response during the elicitation phase and that ASK1 may be a promising therapeutic target for allergic contact dermatitis and other IL-17-related inflammatory diseases

Cellular analysis of SOD1 protein-aggregation propensity and toxicity: a case of ALS with slow progression harboring homozygous SOD1-D92G mutation

Mutations within Superoxide dismutase 1 (SOD1) cause amyotrophic lateral sclerosis (ALS), accounting for approximately 20% of familial cases. The pathological feature is a loss of motor neurons with enhanced formation of intracellular misfolded SOD1. Homozygous SOD1-D90A in familial ALS has been reported to show slow disease progression. Here, we reported a rare case of a slowly progressive ALS patient harboring a novel SOD1 homozygous mutation D92G (homD92G). The neuronal cell line overexpressing SOD1-D92G showed a lower ratio of the insoluble/soluble fraction of SOD1 with fine aggregates of the misfolded SOD1 and lower cellular toxicity than those overexpressing SOD1-G93A, a mutation that generally causes rapid disease progression. Next, we analyzed spinal motor neurons derived from induced pluripotent stem cells (iPSC) of a healthy control subject and ALS patients carrying SOD1-homD92G or heterozygous SOD1-L144FVX mutation. Lower levels of misfolded SOD1 and cell loss were observed in the motor neurons differentiated from patient-derived iPSCs carrying SOD1-homD92G than in those carrying SOD1-L144FVX. Taken together, SOD1-homD92G has a lower propensity to aggregate and induce cellular toxicity than SOD1-G93A or SOD1-L144FVX, and these cellular phenotypes could be associated with the clinical course of slowly progressive ALS

Regulation of NR4A nuclear receptors by p38

In Drosophila, the melanization reaction is an important defense mechanism against injury and invasion of microorganisms. Drosophila tyrosine hydroxylase (TH, also known as Pale) and dopa decarboxylase (Ddc), key enzymes in the dopamine synthesis pathway, underlie the melanin synthesis by providing the melanin precursors dopa and dopamine, respectively. It has been shown that expression of Drosophila TH and Ddc is induced in various physiological and pathological conditions, including bacterial challenge; however, the mechanism involved has not been fully elucidated. Here, we show that ectopic activation of p38 MAPK induces TH and Ddc expression, leading to upregulation of melanization in the Drosophila cuticle. This p38-dependent melanization was attenuated by knockdown of TH and Ddc, as well as by that of Drosophila HR38, a member of the NR4A family of nuclear receptors. In mammalian cells, p38 phosphorylated mammalian NR4As and Drosophila HR38 and potentiated these NR4As to transactivate a promoter containing NR4A-binding elements, with this transactivation being, at least in part, dependent on the phosphorylation. This suggests an evolutionarily conserved role for p38 MAPKs in the regulation of NR4As. Thus, p38-regulated gene induction through NR4As appears to function in the dopamine synthesis pathway and may be involved in immune and stress responses