Mitogen-activated Protein Kinase Phosphatase Research Articles

A novel regulatory mechanism of the MAPK signaling pathway: the negative regulatory role of MKP-1 and its pathophysiological significance

he increasing number of diseases associated with aberrant MAPK signaling underscores the need for a deeper understanding of its regulatory mechanisms: the MAPK pathway is closely associated with cellular responses to various stimuli, and its dysregulation can cause pathologies. This study focuses on the regulatory mechanisms of the MAPK signaling pathway and the role of MAPK phosphatase (MKP) in regulating its activity. The goal of this study is to elucidate the specific function of MKP in the regulation of MAPK signaling and to identify potential therapeutic targets for diseases associated with MAPK dysregulation. In this study, the interaction between MKP and MAPK signaling was investigated using cellular models and biochemical experiments. The primary targets were cells expressing MAPK and MKP, and protein-protein interactions and signaling dynamics were analyzed using methods such as co-immunoprecipitation, confocal microscopy, and flow cytometry. According to this study, MKP is crucial for the exact temporal and spatial modulation of MAPK signaling through its negative regulation. An imbalance in this balance can lead to pathological conditions. This study highlights the importance of MKP in cellular homeostasis and highlights its potential as a therapeutic agent.

Retinoic acid ameliorates rheumatoid arthritis by attenuating inflammation and modulating macrophage polarization through MKP-1/MAPK signaling pathway.

Macrophages are innate immune cells connected with the development of inflammation. Retinoic acid has previously been proved to have anti-inflammatory and anti-arthritic properties. However, the exact mechanism through which retinoic acid modulates arthritis remains unclear. This study aimed to investigate whether retinoic acid ameliorates rheumatoid arthritis by modulating macrophage polarization. This study used retinoic acid to treat mice with adjuvant arthritis and evaluated anti-inflammatory effects by arthritis score, thermal nociceptive sensitization test, histopathologic examination and immunofluorescence assays. In addition, its specific anti-arthritic mechanism was investigated by flow cytometry, cell transfection and inflammatory signaling pathway assays in RAW264.7 macrophages in vitro. Retinoic acid significantly relieved joint pain and attenuated inflammatory cell infiltration in mice. Furthermore, this treatment modulated peritoneal macrophage polarization, increased levels of arginase 1, as well as decreased inducible nitric oxide synthase expression. In vitro, we verified that retinoic acid promotes macrophage transition from the M1 to M2 type by upregulating mitogen-activated protein kinase (MAPK) phosphatase 1 (MKP-1) expression and inhibiting P38, JNK and ERK phosphorylation in lipopolysaccharide-stimulated RAW264.7 cells. Notably, the therapeutic effects of retinoic acid were inhibited by MKP-1 knockdown. Retinoic acid exerts a significant therapeutic effect on adjuvant arthritis in mice by regulating macrophage polarization through the MKP-1/MAPK pathway, and play an important role in the treatment of rheumatic diseases.

New Innovations in Seed Morphology, and Plant Physiology for Crop Protection

The seed is a mainly reproductive unit of the spermatophyta and links the subsequent generations. Other essential seed activities include survival in arid, cold, or other adverse conditions and dispersal. There is a lot of variety in the internal and external architecture of seeds. OsMKP1, a mitogen-activated protein kinase phosphatase, is encoded by GSN1. Rice glume cells proliferate when GSN1 expression is suppressed, producing larger but fewer rice grains. The GSN1 directly interacts with OsMAPK6 and dephosphorylates it, rendering it inactive. Thus, precise regulation of OsMAPK6 activity via reversible phosphorylation is essential for regulating the rice grain size. Some genes also negatively control the grain width and weight by inhibiting cell proliferation. The embryo, which is made up of cotyledons, hypocotyl, and radicle; the endosperm, which feeds the growing embryo; and the seed coat, which envelops the embryo and the endosperm, are the three main parts of plant seeds, each of which has unique biological functions and outcome. The growing seed can benefit from the ability to predict when a sugar burst would arrive through the phloem, since this would allow the seed to quickly adapt to its storage product synthesis. However, there are still certain obstacles that prevent the use against a variety of insect species. The adoption of various formulation procedures that improve dsRNA persistence and cellular absorption in insects is expected to overcome these issues, which are mostly related to the varying RNAi sensitivity of oral RNAi in insects. The CRISPR/Cas system has become the most popular being shown to be useful for editing the genome of plants, its uses in plants have grown significantly in comparison to other technologies.

Co-Stimulation with TWEAK and TGF-β1 Induces Steroid-Insensitive TSLP and CCL5 Production in BEAS-2B Human Bronchial Epithelial Cells.

Steroid-resistant asthma is a common cause of refractory asthma. Type 2 inflammation is the main inflammatory response in asthma, and the mechanism underlying the steroid-resistance of type 2 inflammation has not been completely elucidated. Tumor-necrosis-factor-like apoptosis-inducing factor (TWEAK) and transforming growth factor (TGF)-β1 are involved in epithelial-mesenchymal transition (EMT) and the production of thymic stromal lymphopoietin (TSLP) and C-C motif chemokine ligand 5 (CCL5). We herein hypothesize that the combined exposure to TWEAK and TGF-β1 may result in the development of steroid resistance in bronchial epithelial cells. The bronchial epithelial cell line BEAS-2B was cultured with or without TGF-β1 or TWEAK, in the presence or absence of dexamethasone (DEX). The roles of Smad-independent pathways and MAP kinase phosphatase 1 (MKP-1) were also explored. Co-stimulation of TWEAK and TGF-β1 induced E-cadherin reduction, N-cadherin upregulation, and TSLP and CCL5 production, which were not suppressed by DEX. Inhibition of the nuclear factor kappa beta (NF-κB) and mitogen-activated protein kinase pathways downregulated steroid-unresponsive TSLP and CCL5 production, whereas knockdown of MKP-1 improved steroid-unresponsive TSLP production, induced by co-stimulation with TWEAK and TGF-β1. Therefore, co-stimulation with TWEAK and TGF-β1 can induce the steroid-insensitive production of TSLP and CCL5 in the bronchial epithelium and may contribute to airway inflammation.

Arabidopsis uses a molecular grounding mechanism and a biophysical circuit breaker to limit floral abscission signaling

Abscission is the programmed separation of plant organs. It is widespread in the plant kingdom with important functions in development and environmental response. In Arabidopsis, abscission of floral organs (sepals, petals, and stamens) is controlled by two receptor-like protein kinases HAESA (HAE) and HAESA LIKE-2 (HSL2), which orchestrate the programmed dissolution of the abscission zone connecting floral organs to the developing fruit. In this work, we use single-cell RNA sequencing to characterize the core HAE/HSL2 abscission gene expression program. We identify the MAP KINASE PHOSPHATASE-1/MKP1 gene as a negative regulator of this pathway. MKP1 acts prior to activation of HAE/HSL2 signaling to establish a signaling threshold required for the initiation of abscission. Furthermore, we use single-cell data to identify genes expressed in two subpopulations of abscission zone cells: those proximal and those distal to the plane of separation. We identify INFLORESCENCE DEFICIENT IN ABSCISSION/IDA family genes, encoding activating ligands of HAE/HSL2, as enriched in distal abscission zone cells at the base of the abscising organs. We show how this expression pattern forms a biophysical circuit breaker whereby, when the organ is shed, the source of the IDA peptides is removed, leading to cessation of HAE/HSL2 signaling. Overall, this work provides insight into the multiple control mechanisms acting on the abscission-signaling pathway.

Dynamic and structural insights into allosteric regulation on MKP5 a dual-specificity phosphatase.

Dual-specificity mitogen-activated protein kinase (MAPK) phosphatases (MKPs) directly dephosphorylate and inactivate the MAPKs. Although the catalytic mechanism of dephosphorylation of the MAPKs by the MKPs is established, a complete molecular picture of the regulatory interplay between the MAPKs and MKPs still remains to be fully explored. Here, we sought to define the molecular mechanism of MKP5 regulation through an allosteric site within its catalytic domain. We demonstrate using crystallographic and NMR spectroscopy approaches that residue Y435 is required to maintain the structural integrity of the allosteric pocket. Along with molecular dynamics simulations, these data provide insight into how changes in the allosteric pocket propagate conformational flexibility in the surrounding loops to reorganize catalytically crucial residues in the active site. Furthermore, Y435 contributes to the interaction with p38 MAPK and JNK, thereby promoting dephosphorylation. Collectively, these results highlight the role of Y435 in the allosteric site as a novel mode of MKP5 regulation by p38 MAPK and JNK.

MAPK Phosphatase-5 is required for TGF-β signaling through a JNK-dependent pathway.

Mitogen-activated protein kinase (MAPK) phosphatases (MKPs) constitute members of the dual-specificity family of protein phosphatases that dephosphorylate the MAPKs. MKP-5 dephosphorylates the stress-responsive MAPKs, p38 MAPK and JNK, and has been shown to promote tissue fibrosis. Here, we provide insight into how MKP-5 regulates the transforming growth factor-β (TGF-β) pathway, a well-established driver of fibrosis. We show that MKP-5-deficient fibroblasts in response to TGF-β are impaired in SMAD2 phosphorylation at canonical and non-canonical sites, nuclear translocation, and transcriptional activation of fibrogenic genes. Consistent with this, pharmacological inhibition of MKP-5 is sufficient to block TGF-β signaling, and that this regulation occurs through a JNK-dependent pathway. By utilizing RNA sequencing and transcriptomic analysis, we identify TGF-β signaling activators regulated by MKP-5 in a JNK-dependent manner, providing mechanistic insight into how MKP-5 promotes TGF-β signaling. This study elucidates a novel mechanism whereby MKP-5-mediated JNK inactivation is required for TGF-β signaling and provides insight into the role of MKP-5 in fibrosis.

Wolbachia endosymbionts in Drosophila regulate the resistance to Zika virus infection in a sex dependent manner.

Drosophila melanogaster has been used extensively for dissecting the genetic and functional bases of host innate antiviral immunity and virus-induced pathology. Previous studies have shown that the presence of Wolbachia endosymbionts in D. melanogaster confers resistance to infection by certain viral pathogens. Zika virus is an important vector-borne pathogen that has recently expanded its range due to the wide geographical distribution of the mosquito vector. Here, we describe the effect of Wolbachia on the immune response of D. melanogaster adult flies following Zika virus infection. First, we show that the presence of Wolbachia endosymbionts promotes the longevity of uninfected D. melanogaster wild type adults and increases the survival response of flies following Zika virus injection. We find that the latter effect is more pronounced in females rather than in males. Then, we show that the presence of Wolbachia regulates Zika virus replication during Zika virus infection of female flies. In addition, we demonstrate that the antimicrobial peptide-encoding gene Drosocin and the sole Jun N-terminal kinase-specific MAPK phosphatase Puckered are upregulated in female adult flies, whereas the immune and stress response gene TotM is upregulated in male individuals. Finally, we find that the activity of RNA interference and Toll signaling remain unaffected in Zika virus-infected female and male adults containing Wolbachia compared to flies lacking the endosymbionts. Our results reveal that Wolbachia endosymbionts in D. melanogaster affect innate immune signaling activity in a sex-specific manner, which in turn influences host resistance to Zika virus infection. This information contributes to a better understanding of the complex interrelationship between insects, their endosymbiotic bacteria, and viral infection. Interpreting these processes will help us design more effective approaches for controlling insect vectors of infectious disease.

Nepetoidin B Alleviates Liver Ischemia/Reperfusion Injury via Regulating MKP5 and JNK/P38 Pathway.

Nepetoidin B (NB) has been reported to possess anti-inflammatory, antibacterial, and antioxidant properties. However, its effects on liver ischemia/reperfusion (I/R) injury remain unclear. In this study, a mouse liver I/R injury model and a mouse AML12 cell hypoxia reoxygenation (H/R) injury model were used to investigate the potential role of NB. Serum transaminase levels, liver necrotic area, cell viability, oxidative stress, inflammatory response, and apoptosis were evaluated to assess the effects of NB on liver I/R and cell H/R injury. Quantitative polymerase chain reaction (qPCR) and Western blotting were used to measure mRNA and protein expression levels, respectively. Molecular docking was used to predict the binding capacity of NB and mitogen-activated protein kinase phosphatase 5 (MKP5). The results showed that NB significantly reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, liver necrosis, oxidative stress, reactive oxygen species (ROS) content, inflammatory cytokine content and expression, inflammatory cell infiltration, and apoptosis after liver I/R and AML12 cells H/R injury. Additionally, NB inhibited the JUN protein amino-terminal kinase (JNK)/P38 pathway. Molecular docking results showed good binding between NB and MKP5 proteins, and Western blotting results showed that NB increased the protein expression of MKP5. MKP5 knockout (KO) significantly diminished the protective effects of NB against liver injury and its inhibitory effects on the JNK/P38 pathway. NB exerts hepatoprotective effects against liver I/R injury by regulating the MKP5-mediated P38/JNK signaling pathway.

MPT0E028, a novel pan-HDAC inhibitor, prevents pulmonary fibrosis through inhibition of TGF-β-induced CTGF expression in human lung fibroblasts: Involvement of MKP-1 activation

Histone deacetylase (HDAC) inhibitors are potential candidates for treating pulmonary fibrosis. MPT0E028, a novel pan-HDAC inhibitor, has been reported to exhibit antitumor activity in several cancer cell lines. In this study, we investigated the mechanism underlying the inhibitory effects of MPT0E028 on the expression of fibrogenic proteins in human lung fibroblasts (WI-38). Our results revealed that MPT0E028 inhibited transforming growth factor-β (TGF-β)-, thrombin-, and endothelin 1-induced connective tissue growth factor (CTGF) expression in a concentration-dependent manner. In addition, MPT0E028 suppressed TGF-β-stimulated expression of fibronectin, collagen I, and α-smooth muscle actin (α-SMA). Furthermore, MPT0E028 inhibited the TGF-β-induced phosphorylation of c-Jun N-terminal kinase (JNK), p38, and extracellular signal-regulated kinase (ERK). MPT0E028 reduced the increase in SMAD3 and c-Jun phosphorylation, and SMAD3-and activator protein-1 (AP-1)-luciferase activities under TGF-β stimulation. Transfection with mitogen-activated protein kinase phosphatase-1 (MKP-1) siRNA reversed the suppressive effects of MPT0E028 on TGF-β-induced increases in CTGF expression; JNK, p38, and ERK phosphorylation; and SMAD3 and AP-1 activation. Moreover, MPT0E028 increased MKP-1 acetylation and activity in WI-38 cells. Pretreatment with MPT0E028 reduced the fibrosis score and fibronectin, collagen, and α-SMA expression in bleomycin-induced pulmonary fibrosis mice. In conclusion, MPT0E028 induced MKP-1 acetylation and activation, which in turn inhibited TGF-β-stimulated JNK, p38, and ERK phosphorylation; SMAD3 and AP-1 activation; and subsequent CTGF expression in human lung fibroblasts. Thus, MPT0E028 may be a potential drug for treating pulmonary fibrosis.

Mechanistic Understanding of Dexamethasone-Mediated Protection against Remdesivir-Induced Hepatotoxicity.

Remdesivir (RDV), a broad-spectrum antiviral agent, is often used together with dexamethasone (DEX) for hospitalized COVID-19 patients requiring respiratory support. Potential hepatic adverse drug reaction is a safety concern associated with the use of RDV. We previously reported that DEX cotreatment effectively mitigates RDV-induced hepatotoxicity and reduces elevated serum alanine aminotransferase and aspartate aminotransferase levels in cultured human primary hepatocytes (HPH) and hospitalized COVID-19 patients, respectively. Yet, the precise mechanism behind this protective drug-drug interaction remains largely unknown. Here, we show that through the activation of p38, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinases 1 and 2 (ERK1/2) signaling, RDV induces apoptosis (cleavage of caspases 8, 9, and 3), autophagy (increased autophagosome and LC3-II), and mitochondrial damages (decreased membrane potential, respiration, ATP levels, and increased expression of Bax and the released cytosolic cytochrome C) in HPH. Importantly, cotreatment with DEX partially reversed RDV-induced apoptosis, autophagy, and cell death. Mechanistically, DEX deactivates/dephosphorylates p38, JNK, and ERK1/2 signaling by enhancing the expression of dual specificity protein phosphatase 1 (DUSP1), a mitogen-activated protein kinase (MAPK) phosphatase, in a glucocorticoid receptor (GR)-dependent manner. Knockdown of GR in HPH attenuates DEX-mediated DUSP1 induction, MAPK dephosphorylation, as well as protection against RDV-induced hepatotoxicity. Collectively, our findings suggest a molecular mechanism by which DEX modulates the GR-DUSP1-MAPK regulatory axis to alleviate the adverse actions of RDV in the liver. SIGNIFICANCE STATEMENT: The research uncovers the molecular mechanisms by which dexamethasone safeguards against remdesivir-associated liver damage in the context of COVID-19 treatment.

The E3 Ubiquitin Protein Ligase LINCR Amplifies the TLR-Mediated Signals through Direct Degradation of MKP1.

Toll-like receptors (TLRs) induce innate immune responses through activation of intracellular signaling pathways, such as MAP kinase and NF-κB signaling pathways, and play an important role in host defense against bacterial or viral infections. Meanwhile, excessive activation of TLR signaling leads to a variety of inflammatory disorders, including autoimmune diseases. TLR signaling is therefore strictly controlled to balance optimal immune response and inflammation. However, its balancing mechanisms are not fully understood. In this study, we identified the E3 ubiquitin ligase LINCR/ NEURL3 as a critical regulator of TLR signaling. In LINCR-deficient cells, the sustained activation of JNK and p38 MAPKs induced by the agonists for TLR3, TLR4, and TLR5, was clearly attenuated. Consistent with these observations, TLR-induced production of a series of inflammatory cytokines was significantly attenuated, suggesting that LINCR positively regulates innate immune responses by promoting the activation of JNK and p38. Interestingly, our further mechanistic study identified MAPK phosphatase-1 (MKP1), a negative regulator of MAP kinases, as a ubiquitination target of LINCR. Thus, our results demonstrate that TLRs fine-tune the activation of MAP kinase pathways by balancing LINCR (the positive regulator) and MKP1 (the negative regulator), which may contribute to the induction of optimal immune responses.

6-Gingerol attenuates hepatic ischemia/reperfusion injury through regulating MKP5-mediated P38/JNK pathway

6-Gingerol, the main bioactive compound of ginger, has antioxidant, anti-inflammatory, anti-cancer and neuroprotective effects. However, it is unclear whether 6-Gingerol has protective effects against hepatic ischemia/reperfusion (I/R) injury. In this study, the mouse liver I/R injury model and the mouse AML12 cell hypoxia/reoxygenation (H/R) model were established by pretreatment with 6-Gingerol at different concentrations to explore the potential effects of 6-Gingerol. Serum transaminase levels, liver necrotic area, cell viability, inflammatory response, and cell apoptosis were used to assess the effect of 6-Gingerol on hepatic I/R or cell H/R injury. Quantitative polymerase chain reaction (qPCR) and Western blotting were used to detect the mRNA and protein expression. The results show that 6-Gingerol decreased serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) levels, liver necrosis, inflammatory cytokines IL-1β, IL-6, MCP-1, TNF-α expression, Ly6g+ inflammatory cell infiltration, protein phosphorylation of NF-κB signaling pathway, Terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) positive cells, cell apoptosis rate, the protein expression of pro-apoptotic protein BAX and C-Caspase3, increased cell viability, and expression of anti-apoptotic protein BCL-2. Moreover, 6-Gingerol could increase the mRNA and protein expression of mitogen activated protein kinase phosphatase 5 (MKP5) and inhibit the activation of P38/JNK signaling pathway. In MKP5 knockout (KO) mice, the protective effect of 6-gingerol and the inhibition of P38/JNK pathway were significantly weakened. Therefore, our results suggest that 6-Gingerol exerts anti-inflammatory and anti-apoptotic effects to attenuate hepatic I/R injury by regulating the MKP5-mediated P38/JNK signaling pathway.

The lincRNA JUNI regulates the stress-dependent induction of c-Jun, cellular migration and survival through the modulation of the DUSP14-JNK axis

Cancer cells employ adaptive mechanisms to survive various stressors, including genotoxic drugs. Understanding the factors promoting survival is crucial for developing effective treatments. In this study, we unveil a previously unexplored long non-coding RNA, JUNI (JUN-DT, LINC01135), which is upregulated by genotoxic drugs through the activation of stress-activated MAPKs, JNK, and p38 and consequently exerts positive control over the expression of its adjacent gene product c-Jun, a well-known oncoprotein, which transduces signals to multiple transcriptional outputs. JUNI regulates cellular migration and has a crucial role in conferring cellular resistance to chemotherapeutic drugs or UV radiation. Depletion of JUNI markedly increases the sensitivity of cultured cells and spheroids to chemotherapeutic agents. We identified 57 proteins interacting with JUNI. The activity of one of them the MAPK phosphatase and inhibitor, DUSP14, is counteracted by JUNI, thereby, facilitating efficient JNK phosphorylation and c-Jun induction when cells are exposed to UV radiation. The antagonistic interplay with DUSP14 contributes not only to c-Jun induction but also augments the survival of UV-exposed cells. In summary, we introduce JUNI as a novel stress-inducible regulator of c-Jun, positioning it as a potential target for enhancing the sensitivity of cancer cells to chemotherapy.

Sanguinarine highly sensitises breast cancer cells to doxorubicin-induced apoptosis

Breast cancer is the most commonly diagnosed cancer and the second most common cause of cancer death in women. The anthracycline, doxorubicin, is a well-known and highly effective treatment for breast cancer patients; however, many patients present with resistance to chemotherapeutic drugs, which ultimately results in treatment failure and contributes to high mortality rates. It is well established that the mitogen-activated protein kinase phosphatase 1 (MKP-1) mediates the response to chemotherapy, where upregulated MKP-1 is associated with chemoresistance. We investigated whether MKP-1 inhibition or silencing can sensitise triple-negative MDA-MB-231 breast cancer cells to doxorubicin therapy. We found that MKP-1 inhibition and silencing sensitises breast cancer cells to doxorubicin-induced apoptosis. Additionally, the inhibition of MKP-1 in combination with doxorubicin treatment promotes autophagy induction, while doxorubicin and not MKP-1 modulation increased lysosomal acidic compartments. As such, this study demonstrated that MKP-1 inhibition has a potential therapeutic benefit for breast cancer patients by increasing the efficacy of conventional chemotherapy. Therefore, MKP-1 inhibition should be developed as a clinically relevant adjuvant therapy, which could provide a novel avenue for therapeutic intervention in combination with chemotherapy in breast cancer patients.

Abstract 5953: DUSP6 is a novel target in neuroendocrine tumors

Abstract Background: Neuroendocrine tumors (NETs) are rare malignancies derived from enterochromaffin cells of the diffuse neuroendocrine system and most frequently observed in the gastrointestinal tract and bronchopulmonary system. Surgery is the major treatment for localized NETs. In high-grade NETs, treatment opportunities include chemotherapeutic drugs such as temozolomide (TEM), and capecitabine and small-molecule targeted therapies such as everolimus or sunitinib. Despite promising initial responses, most patients develop therapy resistance by unknown mechanisms. Therefore, there is a pressing need to apprehend the mechanisms of resistance to improve the therapeutic efficacy of the current treatments. Phosphatases are emerging and novel targets in human malignancies and their modulation yields significant anti-tumor activities. Dual specificity phosphatase 6 (DUSP6) is a MAPK phosphatase and mainly binds to and dephosphorylates extracellular signal-regulated kinase (ERK)1/2. DUSP6 expression is increased in several malignancies, correlating with distant metastases and a worse clinical outcome. Moreover, DUSP6 mediates resistance to certain chemotherapeutics and small molecular tyrosine kinase inhibitors and its blockade increases therapeutic sensitivity. The current study aimed to investigate the contribution of DUSP6 to therapy resistance in the NET models. Material and Methods: Using Western blot analysis, the expression of DUSP6 was measured in 8 clinical samples of small bowel neuroendocrine tumors, the NET cell lines BON-1, H69, IMR-32, QGP and H524 and two PDX models, 1452 (rectal neuroendocrine) and 913 (pancreatic neuroendocrine). The effects of DUSP6 depletion on cell viability were explored using a pharmacological inhibitor, BCI, and its siRNA-mediated knockdown. Moreover, BCI-mediated apoptotic cell death was measured by an annexin V/PI staining assay. Results: DUSP6 expression was positive in 5 out of 8 clinical samples, 4 NET cell lines and both the PDX models. BCI treatment induced an apoptotic response in the NET models and synergistically increased sensitivity to TMZ and cisplatin in 1452 cells., RNAi-mediated depletion of DUSP6 also increased sensitivity to sunitinib and triggered TMZ-induced apoptotic cell death in 1452 cells, which was associated with changes in (O6-Methylguanine-DNA-methyltransferase) MGMT expression. These findings suggest that DUSP6 is an important therapeutic target in neuroendocrine tumors and its blockade is a promising therapeutic approach to enhance the therapeutic index of current treatments. Citation Format: Majid Momeny, Vahid Khalaj, Solmaz AghaAmiri, Po Hien Ear, Servando Hernandez Vargas, Sukhen Ghosh, Ali Azhdarinia. DUSP6 is a novel target in neuroendocrine tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5953.

Mitogen-activated protein kinase phosphatase 1 controls broad spectrum disease resistance in Arabidopsis thaliana through diverse mechanisms of immune activation.

Arabidopsis thaliana Mitogen-activated protein Kinase Phosphatase 1 (MKP1) negatively balances production of reactive oxygen species (ROS) triggered by Microbe-Associated Molecular Patterns (MAMPs) through uncharacterized mechanisms. Accordingly, ROS production is enhanced in mkp1 mutant after MAMP treatment. Moreover, mkp1 plants show a constitutive activation of immune responses and enhanced disease resistance to pathogens with distinct colonization styles, like the bacterium Pseudomonas syringae pv. tomato DC3000, the oomycete Hyaloperonospora arabidopsidis Noco2 and the necrotrophic fungus Plectosphaerella cucumerina BMM. The molecular basis of this ROS production and broad-spectrum disease resistance controlled by MKP1 have not been determined. Here, we show that the enhanced ROS production in mkp1 is not due to a direct interaction of MKP1 with the NADPH oxidase RBOHD, nor is it the result of the catalytic activity of MKP1 on RBHOD phosphorylation sites targeted by BOTRYTIS INDUCED KINASE 1 (BIK1) protein, a positive regulator of RBOHD-dependent ROS production. The analysis of bik1 mkp1 double mutant phenotypes suggested that MKP1 and BIK1 targets are different. Additionally, we showed that phosphorylation residues stabilizing MKP1 are essential for its functionality in immunity. To further decipher the molecular basis of disease resistance responses controlled by MKP1, we generated combinatory lines of mkp1-1 with plants impaired in defensive pathways required for disease resistance to pathogen: cyp79B2 cyp79B3 double mutant defective in synthesis of tryptophan-derived metabolites, NahG transgenic plant that does not accumulate salicylic acid, aba1-6 mutant impaired in abscisic acid (ABA) biosynthesis, and abi1 abi2 hab1 triple mutant impaired in proteins described as ROS sensors and that is hypersensitive to ABA. The analysis of these lines revealed that the enhanced resistance displayed by mkp1-1 is altered in distinct mutant combinations: mkp1-1 cyp79B2 cyp79B3 fully blocked mkp1-1 resistance to P. cucumerina, whereas mkp1-1 NahG displays partial susceptibility to H. arabidopsidis, and mkp1-1 NahG, mkp1-1 aba1-6 and mkp1-1 cyp79B2 cyp79B3 showed compromised resistance to P. syringae. These results suggest that MKP1 is a component of immune responses that does not directly interact with RBOHD but rather regulates the status of distinct defensive pathways required for disease resistance to pathogens with different lifestyles.

MAPK phosphatase 1 inhibition of p38α within lung myofibroblasts is essential for spontaneous fibrosis resolution.

Fibrosis following tissue injury is distinguished from normal repair by the accumulation of pathogenic and apoptosis-resistant myofibroblasts (MFs), which arise primarily by differentiation from resident fibroblasts. Endogenous molecular brakes that promote MF dedifferentiation and clearance during spontaneous resolution of experimental lung fibrosis may provide insights that could inform and improve the treatment of progressive pulmonary fibrosis in patients. MAPK phosphatase 1 (MKP1) influences the cellular phenotype and fate through precise and timely regulation of MAPK activity within various cell types and tissues, yet its role in lung fibroblasts and pulmonary fibrosis has not been explored. Using gain- and loss-of-function studies, we found that MKP1 promoted lung MF dedifferentiation and restored the sensitivity of these cells to apoptosis - effects determined to be mainly dependent on MKP1's dephosphorylation of p38α MAPK (p38α). Fibroblast-specific deletion of MKP1 following peak bleomycin-induced lung fibrosis largely abrogated its subsequent spontaneous resolution. Such resolution was restored by treating these transgenic mice with the p38α inhibitor VX-702. We conclude that MKP1 is a critical antifibrotic brake whose inhibition of pathogenic p38α in lung fibroblasts is necessary for fibrosis resolution following lung injury.

Ovomucin Hydrolysates Reduce Bacterial Adhesion and Inflammation in Enterotoxigenic Escherichia coli (ETEC) K88-Challenged Intestinal Epithelial Cells.

Enterotoxigenic Escherichia coli (ETEC) K88 is the most common cause of diarrhea in neonatal and postweaning pigs. After adhering to small intestinal epithelial cells via glycoprotein receptor recognition, the pathogen can produce enterotoxins, impair intestinal integrity, trigger watery diarrhea, and induce inflammation via nuclear factor κB (NF-κB) and mitogen-activated protein kinase phosphatase (MAPK) pathways. Inhibiting ETEC K88 adhesion to cell surfaces by interfering with the receptor-fimbriae recognition provides a promising strategy to prevent the initiation and progression of infection. Ovomucin is a highly glycosylated protein in chicken egg white with diverse bioactivities. Ovomucin hydrolysates prepared by the enzymes Protex 26L (OP) and pepsin/pancreatin (OPP) were previously revealed to prevent adhesion of ETEC K88 to IPEC-J2 cells. Herein, we investigated the protective effects of ovomucin hydrolysates on ETEC K88-induced barrier integrity damage and inflammation in IPEC-J2 and Caco-2 cells. Both hydrolysates inhibited ETEC K88 adhesion to cells and protected epithelial cell integrity by restoring transepithelial electronic resistance (TEER) values. Removing sialic acids in the hydrolysates reduced their antiadhesive capacities. Ovomucin hydrolysates suppressed ETEC-induced activation of NF-κB and MAPK signaling pathways in both cell lines. The ability of ETEC K88 in activating calcium/calmodulin-dependent protein kinase 2 (CaMK II), elevating intracellular Ca2+ concentration, and inducing oxidative stress was attenuated by both hydrolysates. In conclusion, this study demonstrated the potential of ovomucin hydrolysates to prevent ETEC K88 adhesion and alleviate inflammation and oxidative stress in intestinal epithelial cells.

A major gene for chilling tolerance variation in Indica rice codes for a kinase OsCTK1 that phosphorylates multiple substrates under cold.

Rice is susceptible to chilling stress. Identifying chilling tolerance genes and their mechanisms are key to improve rice performance. Here, we performed a genome-wide association study to identify regulatory genes for chilling tolerance in rice. One major gene for chilling tolerance variation in Indica rice was identified as a casein kinase gene OsCTK1. Its function and natural variation are investigated at the physiological and molecular level by its mutants and transgenic plants. Potential substrates of OsCTK1 were identified by phosphoproteomic analysis, protein-protein interaction assay, in vitro kinase assay, and mutant characterization. OsCTK1 positively regulates rice chilling tolerance. Three of its putative substrates, acidic ribosomal protein OsP3B, cyclic nucleotide-gated ion channel OsCNGC9, and dual-specific mitogen-activated protein kinase phosphatase OsMKP1, are each involved in chilling tolerance. In addition, a natural OsCTK1 chilling-tolerant (CT) variant exhibited a higher kinase activity and conferred greater chilling tolerance compared with a chilling-sensitive (CS) variant. The CT variant is more prevalent in CT accessions and is distributed more frequently in higher latitude compared with the CS variant. This study thus enables a better understanding of chilling tolerance mechanisms and provides gene variants for genetic improvement of chilling tolerance in rice.