scholarly journals Ubiquitin plays an atypical role in GPCR-induced p38 MAP kinase activation on endosomes

2015 ◽  
Vol 210 (7) ◽  
pp. 1117-1131 ◽  
Author(s):  
Neil J. Grimsey ◽  
Berenice Aguilar ◽  
Thomas H. Smith ◽  
Phillip Le ◽  
Amanda L. Soohoo ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Transforming Growth Factor ◽  
Inflammatory Responses ◽  
Transforming Growth Factor Β ◽  
P38 Map Kinase ◽  
Mitogen Activated Protein Kinase ◽  
Endothelial Barrier ◽  
Barrier Permeability

Protease-activated receptor 1 (PAR1) is a G protein–coupled receptor (GPCR) for thrombin and promotes inflammatory responses through multiple pathways including p38 mitogen-activated protein kinase signaling. The mechanisms that govern PAR1-induced p38 activation remain unclear. Here, we define an atypical ubiquitin-dependent pathway for p38 activation used by PAR1 that regulates endothelial barrier permeability. Activated PAR1 K63-linked ubiquitination is mediated by the NEDD4-2 E3 ubiquitin ligase and initiated recruitment of transforming growth factor-β–activated protein kinase-1 binding protein-2 (TAB2). The ubiquitin-binding domain of TAB2 was essential for recruitment to PAR1-containing endosomes. TAB2 associated with TAB1, which induced p38 activation independent of MKK3 and MKK6. The P2Y1 purinergic GPCR also stimulated p38 activation via NEDD4-2–mediated ubiquitination and TAB1–TAB2. TAB1–TAB2-dependent p38 activation was critical for PAR1-promoted endothelial barrier permeability in vitro, and p38 signaling was required for PAR1-induced vascular leakage in vivo. These studies define an atypical ubiquitin-mediated signaling pathway used by a subset of GPCRs that regulates endosomal p38 signaling and endothelial barrier disruption.

scholarly journals Naringenin: A Promising Therapeutic Agent against Organ Fibrosis

2021 ◽  
Vol 2021 ◽  
pp. 1-13
Author(s):  
Yanfei Du ◽  
Jun Ma ◽  
Yu Fan ◽  
Xinyu Wang ◽  
Shuzhan Zheng ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Transforming Growth Factor ◽  
Developed Countries ◽  
Mitogen Activated Protein Kinase ◽  
Drug Candidate ◽  
Wide Range ◽  
Fibrotic Disorders ◽  
Organ Fibrosis

Fibrosis is the final common pathology of most chronic diseases as seen in the heart, liver, lung, kidney, and skin and contributes to nearly half of death in the developed countries. Fibrosis, or scarring, is mainly characterized by the transdifferentiation of fibroblasts into myofibroblasts and the excessive accumulation of extracellular matrix (ECM) secreted by myofibroblasts. Despite immense efforts made in the field of organ fibrosis over the past decades and considerable understanding of the occurrence and development of fibrosis gained, there is still lack of an effective treatment for fibrotic diseases. Therefore, identifying a new therapeutic strategy against organ fibrosis is an unmet clinical need. Naringenin, a flavonoid that occurs naturally in citrus fruits, has been found to confer a wide range of pharmacological effects including antioxidant, anti-inflammatory, and anticancer benefits and thus potentially exerting preventive and curative effects on numerous diseases. In addition, emerging evidence has revealed that naringenin can prevent the pathogenesis of fibrosis in vivo and in vitro via the regulation of various pathways that involved signaling molecules such as transforming growth factor-β1/small mother against decapentaplegic protein 3 (TGF-β1/Smad3), mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt), sirtuin1 (SIRT1), nuclear factor-kappa B (NF-κB), or reactive oxygen species (ROS). Targeting these profibrotic pathways by naringenin could potentially become a novel therapeutic approach for the management of fibrotic disorders. In this review, we present a comprehensive summary of the antifibrotic roles of naringenin in vivo and in vitro and their underlying mechanisms of action. As a food derived compound, naringenin may serve as a promising drug candidate for the treatment of fibrotic disorders.

scholarly journals High doses of TGF-β potently suppress type I collagen via the transcription factor CUX1

2011 ◽  
Vol 22 (11) ◽  
pp. 1836-1844 ◽  
Author(s):  
Maria Fragiadaki ◽  
Tetsurou Ikeda ◽  
Abigail Witherden ◽  
Roger M Mason ◽  
David Abraham ◽  
...  
Keyword(s):  
Type I Collagen ◽  
Transforming Growth Factor ◽  
Interstitial Fibrosis ◽  
Transforming Growth Factor Β ◽  
Negative Regulator ◽  
Mitogen Activated Protein Kinase ◽  
Type I ◽  
Aberrant Expression

Transforming growth factor-β (TGF-β) is an inducer of type I collagen, and uncontrolled collagen production leads to tissue scarring and organ failure. Here we hypothesize that uncovering a molecular mechanism that enables us to switch off type I collagen may prove beneficial in treating fibrosis. For the first time, to our knowledge, we provide evidence that CUX1 acts as a negative regulator of TGF-β and potent inhibitor of type I collagen transcription. We show that CUX1, a CCAAT displacement protein, is associated with reduced expression of type I collagen both in vivo and in vitro. We show that enhancing the expression of CUX1 results in effective suppression of type I collagen. We demonstrate that the mechanism by which CUX1 suppresses type I collagen is through interfering with gene transcription. In addition, using an in vivo murine model of aristolochic acid (AA)-induced interstitial fibrosis and human AA nephropathy, we observe that CUX1 expression was significantly reduced in fibrotic tissue when compared to control samples. Moreover, silencing of CUX1 in fibroblasts from kidneys of patients with renal fibrosis resulted in increased type I collagen expression. Furthermore, the abnormal CUX1 expression was restored by addition of TGF-β via the p38 mitogen-activated protein kinase pathway. Collectively, our study demonstrates that modifications of CUX1 expression lead to aberrant expression of type I collagen, which may provide a molecular basis for fibrogenesis.

scholarly journals p38 mitogen-activated protein kinase activation during platelet storage: consequences for platelet recovery and hemostatic function in vivo

Blood ◽  
2010 ◽  
Vol 115 (9) ◽  
pp. 1835-1842 ◽  
Author(s):  
Matthias Canault ◽  
Daniel Duerschmied ◽  
Alexander Brill ◽  
Lucia Stefanini ◽  
Daphne Schatzberg ◽  
...  
Keyword(s):  
Protein Kinase ◽  
P38 Mapk ◽  
P38 Map Kinase ◽  
Mitogen Activated Protein Kinase ◽  
Platelet Recovery ◽  
Mitochondrial Injury ◽  
Platelet Storage ◽  
Receptor Shedding

AbstractPlatelets undergo several modifications during storage that reduce their posttransfusion survival and functionality. One important feature of these changes, which are known as platelet storage lesion, is the shedding of the surface glycoproteins GPIb-α and GPV. We recently demonstrated that tumor necrosis factor-α converting enzyme (TACE/ADAM17) mediates mitochondrial injury-induced shedding of adhesion receptors and that TACE activity correlates with reduced posttransfusion survival of these cells. We now confirm that TACE mediates receptor shedding and clearance of platelets stored for 16 hours at 37°C or 22°C. We further demonstrate that both storage and mitochondrial injury lead to the phosphorylation of p38 mitogen-activated kinase (MAPK) in platelets and that TACE-mediated receptor shedding from mouse and human platelets requires p38 MAP kinase signaling. Protein kinase C, extracellular regulated-signal kinase MAPK, and caspases were not involved in TACE activation. Both inhibition of p38 MAPK and inactivation of TACE during platelet storage led to a markedly improved posttransfusion recovery and hemostatic function of platelets in mice. p38 MAPK inhibitors had only minor effects on the aggregation of fresh platelets under static or flow conditions in vitro. In summary, our data suggest that inhibition of p38 MAPK or TACE during storage may significantly improve the quality of stored platelets.

Blocking p38 signalling inhibits chondrogenesis in vitro but not ankylosis in a model of ankylosing spondylitis in vivo

2011 ◽  
Vol 71 (5) ◽  
pp. 722-728 ◽  
Author(s):  
Kirsten Braem ◽  
Frank P Luyten ◽  
Rik J U Lories
Keyword(s):  
Proinflammatory Cytokines ◽  
Chondrogenic Differentiation ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Inhibitory Effect ◽  
Mitogen Activated Protein Kinase ◽  
Alternative Activation ◽  
Beneficial Effects

ObjectivesTo investigate p38 mitogen activated protein kinase (MAPK) signalling in an in vitro model of bone morphogenetic protein (BMP) and transforming growth factor β (TGFβ)-induced chondrogenesis and in vivo, with specific attention to its potential role in ankylosing enthesitis.MethodsHuman periosteum-derived cells (hPDCs) were cultured in pellets and stimulated with BMP2 or TGFβ1 in the presence or absence of a p38 inhibitor SB203580 or proinflammatory cytokines. Chondrogenic differentiation was evaluated using quantitative PCR. Male DBA/1 mice from different litters were caged together at the age of 8 weeks and treated with SB203580 in both a preventive and therapeutic strategy. The mice were evaluated for prospective signs of arthritis and the toe joints were analysed histologically to assess disease severity.Resultsp38 inhibition by SB203580 and proinflammatory cytokines downregulated chondrogenic markers in pellet cultures stimulated by BMP2 or TGFβ1. In contrast, the in vivo experiments resulted in an increased clinical incidence of arthritis and pathology severity score, reflecting progression towards ankylosis in mice given SB203580.ConclusionInhibition of p38 inhibited chondrogenic differentiation of progenitor cells, showing that not only the SMAD signalling pathways and also alternative activation of MAPKs including p38 contribute to chondrogenesis. Such an inhibitory effect is not found in an in vivo model of joint ankylosis and spondyloarthritis. Increased incidence and severity of disease in preventive experiments and shifts in disease stages in a therapeutic experimental set-up suggest that specific inhibition of p38 may have deleterious rather than beneficial effects.

scholarly journals In Vivo and In Vitro Characterization of a Novel Neuroprotective Strategy for Stroke: Ischemic Postconditioning

2007 ◽  
Vol 28 (2) ◽  
pp. 232-241 ◽  
Author(s):  
Giuseppe Pignataro ◽  
Robert Meller ◽  
Koichi Inoue ◽  
Andrea N Ordonez ◽  
Michelle D Ashley ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Infarct Volume ◽  
Ischemia Reperfusion ◽  
In Vitro Models ◽  
P38 Map Kinase ◽  
Mitogen Activated Protein Kinase ◽  
Neuroprotective Strategy ◽  
The Brain

As clinical trials of pharmacological neuroprotective strategies in stroke have been disappointing, attention has turned to the brain's own endogenous strategies for neuroprotection. Recently, a hypothesis has been offered that modified reperfusion subsequent to a prolonged ischemic episode may also confer ischemic neuroprotection, a phenomenon termed ‘postconditioning’. Here we characterize both in vivo and in vitro models of postconditioning in the brain and offer data suggesting a biological mechanism for protection. Postconditioning treatment reduced infarct volume by up to 50% in vivo and by ∼30% in vitro. A duration of 10 mins of postconditioning ischemia after 10 mins of reperfusion produced the most effective postconditioning condition both in vivo and in vitro. The degree of neuroprotection after postconditioning was equivalent to that observed in models of ischemic preconditioning. However, subjecting the brain to both preconditioning as well as postconditioning did not cause greater protection than each treatment alone. The prosurvival protein kinases extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (MAPK), and Akt show prolonged phosphorylation in the cortex of postconditioned rats. Neuroprotection after postconditioning was inhibited only in the presence of LY294002, which blocks Akt activation, but not U0126 or SB203580, which block ERK and P38 MAP kinase activity. In contrast, preconditioning-induced protection was blocked by LY294002, U0126, and SB203580. Our data suggest that postconditioning may represent a novel neuroprotective approach for focal ischemia/reperfusion, and one that is mediated, at least in part, by the activation of the protein kinase Akt.

scholarly journals Induction of Nitric Oxide Synthase and Activation of Signaling Proteins in Anopheles Mosquitoes by the Malaria Pigment, Hemozoin

2007 ◽  
Vol 75 (8) ◽  
pp. 4012-4019 ◽  
Author(s):  
Leyla Akman-Anderson ◽  
Martin Olivier ◽  
Shirley Luckhart
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Mammalian Cells ◽  
Transforming Growth Factor ◽  
Mitogen Activated Protein Kinase ◽  
Parasite Development ◽  
Mosquito Cells ◽  
Malaria Pigment

ABSTRACT Anopheles stephensi, a major vector for malaria parasite transmission, responds to Plasmodium infection by synthesis of inflammatory levels of nitric oxide (NO), which can limit parasite development in the midgut. We have previously shown that Plasmodium falciparum glycosylphosphatidylinositols (PfGPIs) can induce A. stephensi NO synthase (AsNOS) expression in the midgut epithelium in vivo in a manner similar to the manner in which cytokines and NO are induced by PfGPIs in mammalian cells. In mosquito cells, signaling by PfGPIs and P. falciparum merozoites is mediated through Akt/protein kinase B (Akt/PKB), the mitogen-activated protein kinase kinase DSOR1, and extracellular signal-regulated kinase (ERK). In mammalian cells, a second parasite factor, malaria pigment or hemozoin (Hz), signals NOS induction through ERK- and nuclear factor kappa B-dependent pathways and has been demonstrated to be a novel proinflammatory ligand for Toll-like receptor 9. In this study, we demonstrate that Hz can also induce AsNOS gene expression in immortalized A. stephensi and Anopheles gambiae cell lines in vitro and in A. stephensi midgut tissue in vivo. In mosquito cells, Hz signaling is mediated through transforming growth factor β-associated kinase 1, Akt/PKB, ERK, and atypical protein kinase C zeta/lambda. Our results show that Hz is a prominent parasite-derived signal for Anopheles and that signaling pathways activated by PfGPIs and Hz have both unique and shared components. Together with our previous findings, our data indicate that parasite signaling of innate immunity is conserved in mosquito and mammalian cells.

In vivo RNAi-mediated silencing of TAK1 decreases inflammatory Th1 and Th17 cells through targeting of myeloid cells

Blood ◽  
2010 ◽  
Vol 116 (18) ◽  
pp. 3505-3516 ◽  
Author(s):  
Gabriel Courties ◽  
Virginia Seiffart ◽  
Jessy Presumey ◽  
Virginie Escriou ◽  
Daniel Scherman ◽  
...  
Keyword(s):  
Th17 Cells ◽  
Transforming Growth Factor ◽  
Myeloid Cells ◽  
Transforming Growth Factor Β ◽  
Mitogen Activated Protein Kinase ◽  
Inflammatory Disorder ◽  
T Effector Cells ◽  
Proinflammatory Mediators

Abstract Cells from the mononuclear phagocyte system (MPS) act as systemic and local amplifiers that contribute to the progression of chronic inflammatory disorders. Transforming growth factor-β–activated kinase 1 (TAK1) is a pivotal upstream mitogen-activated protein kinase-kinase-kinase acting as a mediator of cytokine expression. It remains critical to determine in vivo the implication of TAK1 in controlling the innate immune system. Here, we describe a vehicle tailored to selectively deliver siRNAs into MPS cells after intravenous administration, and validate in vivo the potential of the RNAi-mediated TAK1 knock down for immunomodulation. In a mouse model of immune-mediated inflammatory disorder, we show that anti-TAK1 siRNA lipoplexes efficiently alleviate inflammation, severely impair the downstream c-Jun N-terminal kinase and nuclear factor-κB signaling pathways, and decrease the expression of proinflammatory mediators. Importantly, the systemic TAK1 gene silencing decreases the frequency of Th1 and Th17 cells, both mediating autoimmunity in experimental arthritis, demonstrating the immunomodulatory potential of TAK1. Finally, in vitro inhibition of TAK1 in myeloid cells decreases interferon-γ–producing T cells, suggesting that a delivery sys-tem able to target MPS cells and to silence TAK1 impacts on pathogenic T effector cells in autoimmunity.

Abstract 32: Investigating the Autoactivation of p38α Mitogen-Activated Protein Kinase Mediated by Transforming Growth Factor-β-Activated Protein Kinase Binding Protein 1

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Gian Felice De Nicola ◽  
E D Martin ◽  
Rekha Bassi ◽  
Sharwari Verma ◽  
Maria Conte ◽  
...  
Keyword(s):  
Growth Factor ◽  
Protein Kinase ◽  
Transforming Growth Factor ◽  
Binding Protein ◽  
Transforming Growth Factor Β ◽  
Mitogen Activated Protein Kinase ◽  
Scaffolding Protein ◽  
Titration Calorimetry ◽  
Activation Loop

Activation of p38α MAPK (p38α), by phosphorylation of two residues in the TGY motif of the activation loop, can occur independently of upstream kinases. One such mechanism involves the scaffolding protein Transforming Growth Factor-β-activated protein kinase binding protein 1 (TAB1). Under certain circumstances, such as myocardial ischemia, this activation can aggravate lethal injury. It is one of a few examples of activating autophosphorylation and poses a conundrum. How does an inactive kinase, and therefore with low affinity for ATP, phosphorylate its own activation loop when ATP binding is a prerequisite step for phosphotransfer? The aim of this study was to characterize the TAB1 binding of p38α. The binding characteristics of p38α and TAB1 were determined by Isothermal Titration Calorimetry, followed by the binding of p38α and ATPγS, a slowly hydrolysable form of ATP, in the presence and absence of TAB1. The binding of TAB1 to p38α increased significantly the affinity of p38α for ATP. Following the identification of a key region in TAB1 responsible for p38α binding, a synthetic peptide encompassing this region was used to analyze the biophysical and biological consequences of TAB1 binding. In vitro kinase assays were used to test the biochemical characteristics using a combination of wildtype kinase, kinase dead (K53M) or both in the absence or presence of TAB1(371-416). Using an antibody specific to the dual phosphorylation of the TGY motif as a readout, TAB1 binding to p38α increased p38α autophosphorylation in cis . NMR was employed to map the interaction surfaces between of p38α and TAB1 and to analyze the effect of TAB1-binding on p38α. The residues identified as important for the interaction between TAB1 and p38α were mutated and tested in cell free and biological systems to confirm their role as critical determinants for binding. In conclusion, we have further elucidated a mechanism whereby TAB1 binding to p38α alters the conformation of p38α, increasing its affinity for ATP and thereby facilitating autophosphorylation. We have identified the binding contacts of TAB1 and p38α that may be important in the design of therapeutics enabling selective and circumstance-specific inhibition of p38α activation.

scholarly journals Determinants That Control the Specific Interactions between TAB1 and p38α

2006 ◽  
Vol 26 (10) ◽  
pp. 3824-3834 ◽  
Author(s):  
Huamin Zhou ◽  
Min Zheng ◽  
Jianming Chen ◽  
Changchuan Xie ◽  
Anand R. Kolatkar ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Conformational Changes ◽  
Transforming Growth Factor ◽  
Mutational Analysis ◽  
Specific Binding ◽  
Point Mutations ◽  
Transforming Growth Factor Β ◽  
Mitogen Activated Protein Kinase ◽  
Docking Site ◽  
C Terminus

ABSTRACT Previous studies have revealed that transforming growth factor-β-activated protein kinase 1 (TAB1) interacts with p38α and induces p38α autophosphorylation. Here, we examine the sequence requirements in TAB1 and p38α that drive their interaction. Deletion and point mutations in TAB1 reveal that a proline residue in the C terminus of TAB1 (Pro412) is necessary for its interaction with p38α. Furthermore, a cryptic D-domain-like docking site was identified adjacent to the N terminus of Pro412, putting Pro412 in the φB+3 position of the docking site. Through mutational analysis, we found that the previously identified hydrophobic docking groove in p38α is involved in this interaction, whereas the CD domain and ED domain are not. Furthermore, chimeric analysis with p38β (which does not bind to TAB1) revealed a previously unidentified locus of p38α comprising Thr218 and Ile275 that is essential for specific binding of p38α to TAB1. Converting either of these residues to the corresponding amino acid of p38β abolishes p38α interaction with TAB1. These p38α mutants still can be fully activated by p38α upstream activating kinase mitogen-activated protein kinase kinase 6, but their basal activity and activation in response to some extracellular stimuli are reduced. Adjacent to Thr218 and Ile275 is a site where large conformational changes occur in the presence of docking-site peptides derived from p38α substrates and activators. This suggests that TAB1-induced autophosphorylation of p38α results from conformational changes that are similar but unique to those seen in p38α interactions with its substrates and activating kinases.

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