Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.108 (TAT)
 2,389 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The transcription factor NF-kappaB is regulated by the IkappaB family of proteins. The nonphosphorylatable, nondegradable superrepressor IkappaBalpha (srIkappaBalpha) mutant is a potent inhibitor of NF-kappaB activity when expressed in cells. We generated a form of srIkappaBalpha in which its N terminus is fused to the protein transduction domain of HIV TAT (TAT-srIkappaBalpha). Purified TAT-srIkappaBalpha protein rapidly and efficiently entered HeLa or Jurkat T cells. TAT-srIkappaBalpha, when exogenously added to HeLa cells, inhibited in a dose-dependent manner TNF-alpha- or IL-1beta-induced NF-kappaB activation and binding of NF-kappaB to its consensus DNA sequence. TAT-srIkappaBalpha was coimmunoprecipitated with the p65 subunit of NF-kappaB, and this interaction was resistant to stimulation with IL-1beta. Therefore, TAT-srIkappaBalpha-mediated inhibition could result from its nonreversible binding and sequestration of endogenous NF-kappaB. In contrast, exogenously added TAT-srIkappaBalpha did not inhibit IL-1beta-induced activation of extracellular signal-regulated kinase, c-Jun N-terminal kinase, or p38 mitogen-activated protein kinases or the phosphorylation and degradation of endogenous IkappaBalpha. These results identify a novel way for direct regulation of NF-kappaB activity in diverse cell types that may be useful for therapeutic purposes.
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PMID:Inhibition of NF-kappa B activity by a membrane-transducing mutant of I kappa B alpha. 1219 29

Several related isoforms of p38MAPK have been identified and cloned in many species. Although they all contain the dual phosphorylation motif TGY, the expression of these isoforms is not ubiquitous. p38alpha and -beta2 are ubiquitously expressed, whereas p38gamma and -delta appear to have more restricted expression. Because there is evidence for selective activation by upstream kinases and selective preference for downstream substrates, the functions of these conserved proteins is still incompletely understood. We have demonstrated that the renal mesangial cell expresses the mRNA for all the isoforms of p38MAPK, with p38alpha mRNA expressed at the highest level, followed by p38gamma and the lowest levels of expression by p38beta2 and -delta. To determine the functional effects of these proteins on interleukin (IL)-1beta-induced inducible nitric oxide synthase (iNOS) expression, we transduced TAT-p38 chimeric proteins into renal mesangial cells and assessed the effects of wild-type and mutant p38 isoforms on ligand induced iNOS expression. We show that whereas p38gamma and -delta had minimal effects on iNOS expression, p38alpha and -beta2 significantly altered its expression. p38alpha mutant and p38beta2 wild-type dose dependently inhibited IL-1beta-induced iNOS expression. These data suggest that p38alpha and beta2 have reciprocal effects on iNOS expression in the mesangial cell, and these observations may have important consequences for the development of selective inhibitors targeting the p38MAPK family of proteins.
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PMID:Effects of p38MAPK isoforms on renal mesangial cell inducible nitric oxide synthase expression. 1452 18

Activation of group IV cytosolic phospholipase A(2) (gIV-PLA(2)) is the essential first step in the synthesis of inflammatory eicosanoids and in integrin-mediated adhesion of leukocytes. Prior investigations have demonstrated that phosphorylation of gIV-PLA(2) results from activation of at least two isoforms of mitogen-activated protein kinase (MAPK). We investigated the potential role of phosphoinositide 3-kinase (PI3K) in the activation of gIV-PLA(2) and the hydrolysis of membrane phosphatidylcholine in fMLP-stimulated human blood eosinophils. Transduction into eosinophils of Deltap85, a dominant negative form of class IA PI3K adaptor subunit, fused to an HIV-TAT protein transduction domain (TAT-Deltap85) concentration dependently inhibited fMLP-stimulated phosphorylation of protein kinase B, a downstream target of PI3K. FMLP caused increased arachidonic acid (AA) release and secretion of leukotriene C(4) (LTC(4)). TAT-Deltap85 and LY294002, a PI3K inhibitor, blocked the phosphorylation of gIV-PLA(2) at Ser(505) caused by fMLP, thus inhibiting gIV-PLA(2) hydrolysis and production of AA and LTC(4) in eosinophils. FMLP also caused extracellular signal-related kinases 1 and 2 and p38 MAPK phosphorylation in eosinophils; however, neither phosphorylation of extracellular signal-related kinases 1 and 2 nor p38 was inhibited by TAT-Deltap85 or LY294002. Inhibition of 1) p70 S6 kinase by rapamycin, 2) protein kinase B by Akt inhibitor, or 3) protein kinase C by Ro-31-8220, the potential downstream targets of PI3K for activation of gIV-PLA(2), had no effect on AA release or LTC(4) secretion caused by fMLP. We find that PI3K is required for gIV-PLA(2) activation and hydrolytic production of AA in activated eosinophils. Our data suggest that this essential PI3K independently activates gIV-PLA(2) through a pathway that does not involve MAPK.
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PMID:Activation of group IV cytosolic phospholipase A2 in human eosinophils by phosphoinositide 3-kinase through a mitogen-activated protein kinase-independent pathway. 1453 Mar 66

Neutrophil apoptosis occurs both in the bloodstream and in the tissue and is considered essential for the resolution of an inflammatory process. Here, we show that p38-mitogen-activated protein kinase (MAPK) associates to caspase-8 and caspase-3 during neutrophil apoptosis and that p38-MAPK activity, previously shown to be a survival signal in these primary cells, correlates with the levels of caspase-8 and caspase-3 phosphorylation. In in vitro experiments, immunoprecipitated active p38-MAPK phosphorylated and inhibited the activity of the active p20 subunits of caspase-8 and caspase-3. Phosphopeptide mapping revealed that these phosphorylations occurred on serine-364 and serine-150, respectively. Introduction of mutated (S150A), but not wild-type, TAT-tagged caspase-3 into primary neutrophils made the Fas-induced apoptotic response insensitive to p38-MAPK inhibition. Consequently, p38-MAPK can directly phosphorylate and inhibit the activities of caspase-8 and caspase-3 and thereby hinder neutrophil apoptosis, and, in so doing, regulate the inflammatory response.
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PMID:p38-MAPK signals survival by phosphorylation of caspase-8 and caspase-3 in human neutrophils. 1497 Jan 75

Many conventional anticancer treatments kill cells irrespective of whether they are normal or cancerous, so patients suffer from adverse side effects due to the loss of healthy cells. Anticancer insights derived from cell cycle research has given birth to the idea of cell cycle G2 checkpoint abrogation as a cancer cell specific therapy, based on the discovery that many cancer cells have a defective G1 checkpoint resulting in a dependence on the G2 checkpoint during cell replication. Damaged DNA in humans is detected by sensor proteins (such as hHUS1, hRAD1, hRAD9, hRAD17, and hRAD26) that transmit a signal via ATR to CHK1, or by another sensor complex (that may include gammaH2AX, 53BP1, BRCA1, NBS1, hMRE11, and hRAD50), the signal of which is relayed by ATM to CHK2. Most of the damage signals originated by the sensor complexes for the G2 checkpoint are conducted to CDC25C, the activity of which is modulated by 14-3-3. There are also less extensively explored pathways involving p53, p38, PCNA, HDAC, PP2A, PLK1, WEE1, CDC25B, and CDC25A. This review will examine the available inhibitors of CHK1 (Staurosporin, UCN-01, Go6976, SB-218078, ICP-1, and CEP-3891), both CHK1 and CHK2 (TAT-S216A and debromohymenialdisine), CHK2 (CEP-6367), WEE1 (PD0166285), and PP2A (okadaic acid and fostriecin), as well as the unknown checkpoint inhibitors 13-hydroxy-15-ozoapathin and the isogranulatimides. Among these targets, CHK1 seems to be the most suitable target for therapeutic G2 abrogation to date, although an unexplored target such as 14-3-3 or the strategy of targeting multiple proteins at once may be of interest in the future.
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PMID:G2 checkpoint abrogators as anticancer drugs. 1507 95

Treatment of many cancers relies on the combined action of several genotoxins, but the detrimental effect of these drugs on normal cells can cause severe side effects. One major challenge in anticancer therapy is therefore to increase the selectivity of current treatments toward cancer cells in order to spare normal cells. We have recently demonstrated that a RasGAP caspase cleavage fragment is able to sensitize HeLa cells towards cisplatin-induced apoptosis. Here, we extend this observation by showing that this fragment also enhances cell death induced by adriamycin and mitoxantrone, two other widely used genotoxins. Furthermore, we have delineated a short sequence within this fragment that still bears the genotoxin-sensitization property. The peptide encoded by this sequence, when fused to the TAT cell permeation sequence, potently sensitized a number of tumors cells, but not normal cells, towards apoptosis induced by cisplatin, adriamycin and mitoxantrone. This sensitization effect was not mediated through modulation of NFkappaB activity or activation of the JNK and p38 MAPK pathways. Our results demonstrate the feasibility in enhancing the efficacy of currently used drugs to selectively kill cancer cells using peptides derived from pro-apoptotic caspase substrate fragments.
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PMID:A RasGAP-derived cell permeable peptide potently enhances genotoxin-induced cytotoxicity in tumor cells. 1546 50

The effect of fever on neutrophils has not been explored. We tested the hypothesis that fever-like temperature spikes affect neutrophil signaling and function. Prior 60 min, 42 degrees C heat exposure inhibited p38 MAPK, ERK, PI3-Kinase/Akt, and NF-kappaB activation in TNF-alpha-challenged suspended neutrophils. Using pharmacological inhibitors and an inhibitory peptide transduced into neutrophils by a HIV-TAT sequence, we found that p38 MAPK and NF-kappaB mediate TNF-alpha-mediated delayed apoptosis in suspended neutrophils. Heat exposure (39-42 degrees C) did not affect constitutive apoptosis but abrogated TNF-alpha-delayed apoptosis in these suspended cells. In contrast, adhesion-dependent functions were not inhibited. Furthermore, we found that heat exposure neither blocked p38 MAPK, ERK, and NF-kappaB activation in neutrophils on fibronectin nor prevented delayed apoptosis by TNF-alpha when cells interacted with fibronectin. Above and beyond apoptosis, TNF-alpha initiated NF-kappaB-dependent gene transcription. Heat exposure blocked this effect in suspended neutrophils but not in neutrophils on fibronectin. Finally, we show that beta2-integrins, which are not necessary for TNF-alpha-induced NF-kappaB activation at 37 degrees C, transduce costimulatory signals allowing NF-kappaB activation after heat exposure. The effect could protect circulating neutrophils from TNF-alpha activation, while not interfering with activation of adherent neutrophils. Fever could make neutrophils more parsimonious.
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PMID:The effect of fever-like temperatures on neutrophil signaling. 1575 71

In the present study, we analyzed human follicle-stimulating hormone (FSH)-induced cell proliferation and transactivation of estrogen-sensitive reporter genes-in L cells stably expressing the human FSH receptor [L-(hFSHR(+)) cells]. In order to dissect the signaling pathways involved in this process, L-(hFSHR(+)) cells were transiently transfected with either the 3X-ERE-TAT-Luc or the ERE-VitA2-TK-CAT reporter genes and treated with FSH or PKA activators (cholera toxin, forskolin and 8-Br-cAMP) in the presence or absence of various kinase inhibitors. We found that FSH and all PKA activators, specifically induced transactivation of both reporter genes. Transactivation of estrogen-sensitive genes by FSH or PKA activators were blocked (approximately 90%) by H89 (PKA inhibitor) and LY294002 but not by Wortmannin (PI3-K inhibitors), 4-OH-tamoxifen, ICI182,780 or SB203580 (p38 MAPK inhibitor); PD98059 (ERK1/2 inhibitor) partially (approximately 30%) blocked the FSH-mediated effect. The combination of FSH and estradiol resulted in a synergistic effect on transactivation as well as on cell proliferation, and this enhancement was attenuated by antiestrogens. We additionally analyzed the participation of the coactivators SRC-1 and cAMP response element binding protein (CREB)-binding protein (CBP) in FSH-evoked estrogen receptor (ER)-dependent transactivation; we found that CBP but not SRC-1 potentiated FSH-induced transcriptional activation of both ER-sensitive reporters, being this effect stronger on the ERE-VitA2-TK-CAT than on the 3X-ERE-TAT-Luc reporter. Thus, in L-(hFSHR(+)) cells FSH induces transcriptional activation of estrogen-sensitive genes through an A-kinase-triggered signaling pathway, using also to a lesser extent the ERK1/2 and p38 pathways. PI3-K is not apparently involved in this FSH-mediated process since LY294002, but not Wortmannin, specifically binds ERs and completely blocks estrogen action. Presumably, CBP cooperates with the ER on genes that contain estrogen responsive elements through mechanisms involving the participation of other proteins and/or basal transcription factors (e.g. CREB), which in turn mediate the transcriptional response of estrogen-sensitive reporter genes to FSH stimulation.
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PMID:Effects of FSH and 17beta-estradiol on the transactivation of estrogen-regulated promoters and cell proliferation in L cells. 1585 48

1. Myristoylated pseudosubstrate of PKCzeta (mPS) - a synthetic myristoylated peptide with a sequence (13 amino acids) mimicking the endogenous PKCzeta pseudosubstrate region -- is considered a selective cell-permeable inhibitor of PKCzeta. We present strong evidence that in endothelial cells the action of mPS is not limited to inhibition of PKC activity and that myristoylation of certain peptides can activate eNOS (endothelial nitric oxide synthase) through Akt phosphorylation. 2. mPS at micromolar concentrations (1-10 microM) induced profound phosphorylation of eNOS, Akt, ERK 1/2, and p38 MAPK in cultured pulmonary artery endothelial cells (PAEC). The same changes were observed after treatment of PAEC with a myristoylated scrambled version of mPS (mScr), whereas a cell-permeable version of PKCzeta pseudosubstrate fused to the HIV-TAT membrane-translocating peptide did not induce analogous changes, suggesting that myristoylation confers new properties on the peptides consisting of activation of different signaling pathways in endothelial cells. 3. In addition to mPS and mScr, a number of other myristoylated peptides induced phosphorylation of eNOS suggesting that myristoylation of peptides can activate eNOS by mechanisms unrelated to inhibition of PKC. All active myristoylated peptides contained basic amino acids motif and were longer than six amino acids. 4. Activation of eNOS by myristoylated peptides was dependent on the PI3K/Akt pathway and the rise of intracellular calcium and was associated with an elevation of cGMP levels in PAEC and with relaxation of precontracted isolated pulmonary artery segments. 5. Myristoylated peptides can be considered a new class of activators of NO production in endothelial cells and that using mPS as a specific inhibitor of PKC should be done with caution, especially in endothelial cells.
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PMID:Peptides modified by myristoylation activate eNOS in endothelial cells through Akt phosphorylation. 1671 18

Neutrophil NADPH oxidase plays a key role in host defense and in inflammation by releasing large amounts of superoxide and other ROSs. Proinflammatory cytokines such as GM-CSF and TNF-alpha prime ROS production by neutrophils through unknown mechanisms. Here we used peptide sequencing by tandem mass spectrometry to show that GM-CSF and TNF-alpha induce phosphorylation of Ser345 on p47phox, a cytosolic component of NADPH oxidase, in human neutrophils. As Ser345 is located in the MAPK consensus sequence, we tested the effects of MAPK inhibitors. Inhibitors of the ERK1/2 pathway abrogated GM-CSF-induced phosphorylation of Ser345, while p38 MAPK inhibitor abrogated TNF-alpha-induced phosphorylation of Ser345. Transfection of HL-60 cells with a mutated p47phox (S345A) inhibited GM-CSF- and TNF-alpha-induced priming of ROS production. This event was also inhibited in neutrophils by a cell-permeable peptide containing a TAT-p47phox-Ser345 sequence. Furthermore, ROS generation, p47phox-Ser345 phosphorylation, and ERK1/2 and p38 MAPK phosphorylation were increased in synovial neutrophils from rheumatoid arthritis (RA) patients, and TAT-Ser345 peptide inhibited ROS production by these primed neutrophils. This study therefore identifies convergent MAPK pathways on Ser345 that are involved in GM-CSF- and TNF-alpha-induced priming of neutrophils and are activated in RA. Inhibition of the point of convergence of these pathways might serve as a novel antiinflammatory strategy.
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PMID:A specific p47phox -serine phosphorylated by convergent MAPKs mediates neutrophil NADPH oxidase priming at inflammatory sites. 1677 89


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