Sample records for phosphoinositide 3-kinase-dependent mechanism

  1. Phosphoinositide 3-kinase-dependent antagonism in mammalian olfactory receptor neurons.

    PubMed

    Ukhanov, Kirill; Brunert, Daniela; Corey, Elizabeth A; Ache, Barry W

    2011-01-01

    Phosphoinositide signaling, in particular, phosphoinositide 3-kinase (PI3K) signaling, has been implicated in mediating inhibitory odorant input to mammalian olfactory receptor neurons (ORNs). To better understand this phenomenon we investigated PI3K-dependent inhibition between single odorant pairs. The concentration-dependent inhibition of the response of native rat ORNs to octanol by citral is PI3K dependent; blocking PI3K activity with the ? and ? isoform-specific inhibitors AS252424 (5-[5-(4-fluoro-2-hydroxy-phenyl)-furan-2-ylmethylene]-thiazolidine-2,4-dione) and TGX221(7-methyl-2-(4-morpholinyl)-9-[1-(phenylamino)ethyl]-4H-pyrido [1,2-a]pyrimidin-4-one) eliminated or strongly reduced the inhibition. Interestingly, blocking PI3K also changed the apparent agonist strength of the otherwise noncompetitive antagonist citral. The excitation evoked by citral after blocking PI3K, could be suppressed by the adenylate cyclase III (ACIII) blockers MDL12330A (cis-N-(2-phenylcyclopentyl)-azacyclotridec-1-en-2-amine hydrochloride) and SQ22536 [9-(tetrahydro-2-furanyl)-9H-purin-6-amine], indicating that citral could also activate ACIII, presumably through the canonical olfactory receptor (OR). The G-protein G(?)? subunit blockers suramin (8,8'-[carbonylbis[imino-3,1-phenylen ecarbonylimino(4-methyl-3,1-phenylene)carbonylimino

  2. Phosphoinositide 3-kinase-dependent Ras activation by tauroursodesoxycholate in rat liver.

    PubMed Central

    Kurz, A K; Block, C; Graf, D; Dahl, S V; Schliess, F; Häussinger, D

    2000-01-01

    Ursodesoxycholic acid, widely used for the treatment of cholestatic liver disease, causes choleretic, anti-apoptotic and immunomodulatory effects. Here the effects on choleresis of its taurine conjugate tauroursodesoxycholate (TUDC), which is present in the enterohepatic circulation, were correlated with the activation of important elements of intracellular signal transduction in cultured rat hepatocytes and perfused rat liver. TUDC induced a time- and concentration-dependent activation of the small GTP-binding protein Ras and of phosphoinositide 3-kinase (PI 3-kinase) in cultured hepatocytes. Ras activation was dependent on PI 3-kinase activity, without the involvement of protein kinase C- and genistein-sensitive tyrosine kinases. Ras activation by TUDC was followed by an activation of the mitogen-activated protein kinases extracellular-signal-regulated kinase-1 (Erk-1) and Erk-2. In perfused rat liver, PI 3-kinase inhibitors largely abolished the stimulatory effect of TUDC on taurocholate excretion, suggesting an important role for a PI 3-kinase/Ras/Erk pathway in the choleretic effect of TUDC. PMID:10926845

  3. Mouse system-N amino acid transporter, mNAT3, expressed in hepatocytes and regulated by insulin-activated and phosphoinositide 3-kinase-dependent signalling.

    PubMed Central

    Gu, Sumin; Langlais, Paul; Liu, Feng; Jiang, Jean X

    2003-01-01

    Amino acid transporters are essential for normal cell function and physiology. In the present study, we report the identification and functional and regulatory characterization of a mouse system-N amino acid transporter, mNAT3. Expression of mNAT3 in Xenopus oocytes revealed that the strongest transport activities were preferred for L-alanine. In addition, mNAT3 is an Na(+)- and pH-dependent low-affinity transporter and it partially tolerates substitution of Na(+) by Li(+). mNAT3 has been found to be expressed predominantly in the liver, where it is localized to the plasma membrane of hepatocytes, with the strongest expression in those cells adjacent to the central vein, decreasing gradually towards the portal tract. Treatment of mouse hepatocyte-like H2.35 cells with insulin led to a significant increase in the expression of mNAT3, and this stimulation was associated closely with an increase in the uptake of L-alanine. Interestingly, this insulin-induced stimulatory effect on mNAT3 expression was attenuated by the phosphoinositide 3-kinase inhibitor LY294002, but not by the mitogen-activated protein kinase inhibitor PD98059, although both kinases were fully activated by insulin. The results suggest that insulin-mediated regulation of mNAT3 is likely to be mediated through a phosphoinositide 3-kinase-dependent signalling pathway. The unique expression pattern and insulin-mediated regulatory properties of mNAT3 suggest that this transporter may play an important role in liver physiology. PMID:12537539

  4. Ephrin reverse signaling controls palate fusion via a PI3 kinase-dependent mechanism

    PubMed Central

    Miguel, Symone San; Serrano, Maria J.; Sachar, Ashneet; Henkemeyer, Mark; Svoboda, Kathy K. H.; Benson, M. Douglas

    2010-01-01

    Secondary palate fusion requires adhesion and epithelial to mesenchymal transition (EMT) of the epithelial layers on opposing palatal shelves. This EMT requires transforming growth factor ?3 (TGF?3), and its failure results in cleft palate. Ephrins, and their receptors, the Ephs, are responsible for migration, adhesion, and midline closure events throughout development. Ephrins can also act as signal transducing receptors in these processes, with the Ephs serving as ligands (termed "reverse" signaling). We found that activation of ephrin reverse signaling in chicken palates induced fusion in the absence of TGF?3, and that PI3K inhibition abrogated this effect. Further, blockage of reverse signaling inhibited TGF?3-induced fusion in the chicken and natural fusion in the mouse. Thus, ephrin reverse signaling is necessary and sufficient to induce palate fusion independent of TGF?3. These data describe both a novel role for ephrins in palate morphogenesis, and a previously unknown mechanism of ephrin signaling. PMID:21246652

  5. RhoG regulates anoikis through a phosphatidylinositol 3-kinase-dependent mechanism

    SciTech Connect

    Yamaki, Nao [Laboratory of Molecular Neurobiology, Graduate School of Biostudies, Kyoto University, Yoshidakonoe-cho, Sakyo-ku, Kyoto 606-8501 (Japan); Negishi, Manabu [Laboratory of Molecular Neurobiology, Graduate School of Biostudies, Kyoto University, Yoshidakonoe-cho, Sakyo-ku, Kyoto 606-8501 (Japan); Katoh, Hironori [Laboratory of Molecular Neurobiology, Graduate School of Biostudies, Kyoto University, Yoshidakonoe-cho, Sakyo-ku, Kyoto 606-8501 (Japan)]. E-mail: hirokato@pharm.kyoto-u.ac.jp

    2007-08-01

    In normal epithelial cells, cell-matrix interaction is required for cell survival and proliferation, whereas disruption of this interaction causes epithelial cells to undergo apoptosis called anoikis. Here we show that the small GTPase RhoG plays an important role in the regulation of anoikis. HeLa cells are capable of anchorage-independent cell growth and acquire resistance to anoikis. We found that RNA interference-mediated knockdown of RhoG promoted anoikis in HeLa cells. Previous studies have shown that RhoG activates Rac1 and induces several cellular functions including promotion of cell migration through its effector ELMO and the ELMO-binding protein Dock180 that function as a Rac-specific guanine nucleotide exchange factor. However, RhoG-induced suppression of anoikis was independent of the ELMO- and Dock180-mediated activation of Rac1. On the other hand, the regulation of anoikis by RhoG required phosphatidylinositol 3-kinase (PI3K) activity, and constitutively active RhoG bound to the PI3K regulatory subunit p85{alpha} and induced the PI3K-dependent phosphorylation of Akt. Taken together, these results suggest that RhoG protects cells from apoptosis caused by the loss of anchorage through a PI3K-dependent mechanism, independent of its activation of Rac1.

  6. Insulin and insulin-like growth factor I up-regulate GLUT4 gene expression in fetal brown adipocytes, in a phosphoinositide 3-kinase-dependent manner.

    PubMed Central

    Valverde, A M; Navarro, P; Teruel, T; Conejo, R; Benito, M; Lorenzo, M

    1999-01-01

    Fetal brown adipocytes cultured in a serum-free medium, containing 5 mM glucose, expressed both GLUT4 and GLUT1 glucose transporters at the mRNA and protein level. Treatment with either insulin or insulin-like growth factor (IGF)-I at physiological concentrations up-regulates the expression of the GLUT4 gene, producing a time-dependent mRNA accumulation (7-fold increase at 24 h) and a 2.5-fold increase in the amount of protein in the total membrane fraction. However, insulin treatment down-regulates GLUT1 mRNA and protein expression. Moreover, either insulin or IGF-I transactivates a full-promoter GLUT4-chloramphenicol acetyltransferase gene (CAT) construct transiently transfected to the cells, without affecting GLUT1-CAT activity. In consequence, insulin treatment for 24 h increased by 3-fold the basal glucose uptake. Inhibition of phosphoinositide (PI) 3-kinase activity with chemical agents such as wortmannin or LY294002 partially blocked insulin-induced GLUT4 mRNA accumulation, insulin-induced GLUT4 protein content, GLUT4-CAT transactivation and glucose uptake. Furthermore, co-transfection of brown adipocytes with a dominant-negative form of PI 3-kinase precluded the transactivation of the GLUT4 promoter by insulin. However, inhibition of p70S6 kinase (p70(s6k)) with rapamycin or of mitogen-activated protein kinase (MAPK) with PD098059 does not preclude insulin effects on GLUT4 gene expression or glucose uptake. Our results show for the first time a positive effect of insulin on GLUT4 gene expression in fetal brown adipocytes, suggesting the existence of insulin response element(s) in its promoter. Moreover, PI 3-kinase, but not p70(s6k) or MAPK, is an essential requirement for insulin regulation of GLUT4 gene expression. PMID:9895282

  7. Enteropathogenic Escherichia coli mediates antiphagocytosis through the inhibition of PI 3-kinase-dependent pathways

    PubMed Central

    Celli, Jean; Olivier, Martin; Finlay, B.Brett

    2001-01-01

    The extracellular pathogen enteropathogenic Escherichia coli (EPEC) uses a type III secretion system to inhibit its uptake by macrophages. We show that EPEC antiphagocytosis is independent of the translocated intimin receptor Tir and occurs by preventing F-actin polymerization required for bacterial uptake. EPEC–macrophage contact triggered activation of phosphatidylinositol (PI) 3-kinase, which was subsequently inhibited in a type III secretion-dependent manner. Inhibition of PI 3-kinase significantly reduced uptake of a secretion-deficient mutant, without affecting antiphagocytosis by the wild type, suggesting that EPEC blocks a PI 3-kinase-dependent phagocytic pathway. EPEC specifically inhibited Fc? receptor- but not CR3-receptor mediated phagocytosis of opsonized zymosan. We showed that EPEC inhibits PI 3-kinase activity rather than its recruitment to the site of bacterial contact. Phago cytosis of a secretion mutant correlated with the association of PI 3-kinase with tyrosine-phosphorylated proteins, which wild-type EPEC prevented. These results show that EPEC blocks its uptake by inhibiting a PI 3-kinase-mediated pathway, and translocates effectors other than Tir to interfere with actin-driven host cell processes. This constitutes a novel mechanism of phagocytosis avoidance by an extracellular pathogen. PMID:11250891

  8. Deletion of the phosphoinositide 3-Kinase p110(gamma) gene attenuates murine atherosclerosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Inflammatory cell activation by chemokines requires intracellular signaling through phosphoinositide 3-kinase (PI3-kinase) and the PI3-kinase-dependent protein serine/threonine kinase Akt. Atherosclerosis is a chronic inflammatory process driven by oxidatively modified (atherogenic) lipoproteins, ch...

  9. Resolvin D1 Reduces Infarct Size Through a Phosphoinositide 3-Kinase/Protein Kinase B Mechanism.

    PubMed

    Gilbert, Kim; Bernier, Judith; Bourque-Riel, Valérie; Malick, Mandy; Rousseau, Guy

    2015-07-01

    This study was designed to determine if Resolvin D1 (RvD1), a pro-resolution metabolite of the omega-3 polyunsaturated fatty acid docosahexaenoic acid, could decrease myocardial infarct size with delivered at the onset of ischemia. Male Sprague Dawley rats underwent 40 minutes of myocardial ischemia followed by reperfusion. These animals received 1 intraventricular injection of RvD1 (0.01, 0.1, or 0.3 ?g RvD1) or vehicle (saline) before coronary occlusion. Infarct size and neutrophil accumulation were evaluated 24 hours after the onset of reperfusion. Caspase-3, caspase-8, protein kinase B (Akt) activities were evaluated 30 minutes after the reperfusion. Rats receiving 0.1 or 0.3 ?g RvD1 showed a significant decrease of infarct size and caspase-3 and caspase-8 activities compared with the vehicle controls. Neutrophil accumulations were reduced in rats administered RvD1 compared with vehicle, independently of dose level. Akt activation was increased only in animals receiving 0.1 or 0.3 ?g, whereas no change was observed in the 0.01 ?g group. When they were treated with LY-294002, a phosphoinositide 3-kinase (PI3K)/Akt inhibitor, cardioprotection by RvD1 was abrogated. RvD1 treatment at the onset of ischemia decreases infarct size by a mechanism involving the PI3K/Akt pathway. PMID:25806690

  10. Small GTPases and phosphoinositides in the regulatory mechanisms of macropinosome formation and maturation

    PubMed Central

    Egami, Youhei; Taguchi, Tomohiko; Maekawa, Masashi; Arai, Hiroyuki; Araki, Nobukazu

    2014-01-01

    Macropinosome formation requires the sequential activation of numerous signaling pathways that coordinate the actin-driven formation of plasma membrane protrusions (ruffles) and circular ruffles (macropinocytic cups), followed by the closure of these macropinocytic cups into macropinosomes. In the process of macropinosome formation, localized productions of phosphoinositides such as PI(4,5)P2 and PI(3,4,5)P3 spatiotemporally orchestrate actin polymerization and rearrangement through recruiting and activating a variety of actin-associated proteins. In addition, the sequential activation of small GTPases, which are known to be master regulators of the actin cytoskeleton, plays a pivotal role in parallel with phosphoinositides. To complete macropinosome formation, phosphoinositide breakdown and Rho GTPase deactivation must occur in appropriate timings. After the nascent macropinosomes are formed, phosphoinositides and several Rab GTPases control macropinosome maturation by regulating vesicle trafficking and membrane fusion. In this review, we summarize recent advances in our understanding of the critical functions of phosphoinositide metabolism and small GTPases in association with their downstream effectors in macropinocytosis. PMID:25324782

  11. Inhibition of excitatory amino acid-induced phosphoinositide hydrolysis as a possible mechanism of nitroprusside neurotoxicity.

    PubMed

    Yu, O; Chuang, D M

    1996-01-01

    Inclusion of sodium nitroprusside (Na2[Fe(2+)-(CN)5NO]) into the culture medium is toxic to cultured rat cerebellar granule neurons. A possible underlying mechanism may be the inhibition of phosphoinositide (PI) response to excitatory amino acids (EAAs) because activation of glutamate receptors can be neuroprotective and neurotrophic in differentiating neurons. Sodium nitroprusside selectively inhibited the PI response to EAAs (NMDA > glutamate = quisqualate > kainate) without affecting that to carbachol or KCl. In contrast, S-nitroso-N-acetyl-penicillamine (SNAP), another nitric oxide (NO) donor, potentiated NMDA-induced PI hydrolysis. Hemoglobin reversed the effects of nitroprusside and SNAP. However, NO may not be involved because NO solution was without effect and N-acetylpenicillamine, a SNAP analogue that does not contain a NO moiety, also potentiated NMDA-induced PI hydrolysis in a hemoglobin-sensitive manner. Furthermore, the metabolites of NO (nitrate and nitrite), L-arginine, reduced glutathione, 8-bromo-cyclic guanosine 3':5'-cyclic monophosphate (8-Br-cGMP), and atrial natriuretic peptide, which accelerates the production of cGMP independent of NO, were ineffective as modulators. However, potassium ferrocyanide (K4[Fe2+(CN)6]), but not potassium ferricyanide (K3[Fe3+(CN)6]), inhibited NMDA-induced PI hydrolysis as effectively as nitroprusside, but this inhibition was not reversed by hemoglobin. Cyanide, a product from the disintegration of nitroprusside, potentiated rather than inhibited NMDA-induced PI hydrolysis. Taken together, these results suggest that the parent molecule itself, nitroprusside, contributes primarily in inhibiting EAA-induced PI hydrolysis. Inhibition of EAA-induced PI hydrolysis may in part mediate the mechanisms of nitroprusside toxicity in primary cultures of differentiating cerebellar granule neurons. PMID:8522973

  12. Mechanism of the regulation of type IB phosphoinositide 3OH-kinase byG-protein betagamma subunits.

    PubMed Central

    Krugmann, Sonja; Cooper, Matthew A; Williams, Dudley H; Hawkins, Phillip T; Stephens, Len R

    2002-01-01

    Type IB phosphoinositide 3OH-kinase (PI3K) is activated by G-protein betagamma subunits (Gbetagammas). The enzyme is soluble and largely cytosolic in vivo. Its substrate, PtdIns(4,5)P(2), and the Gbetagammas are localized at the plasma membrane. We have addressed the mechanism by which Gbetagammas regulate the PI3K using an in vitro approach. We used sedimentation assays and surface plasmon resonance to determine association of type IB PI3K with lipid monolayers and vesicles of varying compositions, some of which had Gbetagammas incorporated. Association and dissociation rate constants were determined. Our results indicated that in an assay situation in vitro the majority of PI3K will be associated with lipid vesicles, irrespective of the presence or absence of Gbetagammas. In line with this, a constitutively active membrane-targeted PI3K construct could still be activated substantially by Gbetagammas in vitro. We conclude that Gbetagammas activate type IB PI3K by a mechanism other than translocation to the plasma membrane. PMID:11879201

  13. Structural Basis and Mechanism of Autoregulation in 3-Phosphoinositide-Dependent Grp1 Family Arf GTPase Exchange Factors

    Microsoft Academic Search

    Jonathan P. DiNitto; Anna M. Delprato; Meng-Tse Gabe Lee; Thomas Charles Cronin; Shaohui Huang; Adilson L. Guilherme; Michael P. Czech; David G. Lambright

    2007-01-01

    Arf GTPases regulate membrane trafficking and actin dynamics. Grp1, ARNO, and Cytohesin-1 comprise a family of phosphoinositide-dependent Arf GTPase exchange factors with a Sec7-pleckstrin homology (PH) domain tandem. Here, we report that the exchange activity of the Sec7 domain is potently autoinhibited by conserved elements proximal to the PH domain. The crystal structure of the Grp1 Sec7-PH tandem reveals a

  14. Targeting the phosphoinositide 3-kinase pathway in hematologic malignancies

    PubMed Central

    Jabbour, Elias; Ottmann, Oliver G.; Deininger, Michael; Hochhaus, Andreas

    2014-01-01

    The phosphoinositide 3-kinase pathway represents an important anticancer target because it has been implicated in cancer cell growth, survival, and motility. Recent studies show that PI3K may also play a role in the development of resistance to currently available therapies. In a broad range of cancers, various components of the phosphoinositide 3-kinase signaling axis are genetically modified, and the pathway can be activated through many different mechanisms. The frequency of genetic alterations in the phosphoinositide 3-kinase pathway, coupled with the impact in oncogenesis and disease progression, make this signaling axis an attractive target in anticancer therapy. A better understanding of the critical function of the phosphoinositide 3-kinase pathway in leukemias and lymphomas has led to the clinical evaluation of novel rationally designed inhibitors in this setting. Three main categories of phosphoinositide 3-kinase inhibitors have been developed so far: agents that target phosphoinositide 3-kinase and mammalian target of rapamycin (dual inhibitors), pan-phosphoinositide 3-kinase inhibitors that target all class I isoforms, and isoform-specific inhibitors that selectively target the ?, -?, -?, or -? isoforms. Emerging data highlight the promise of phosphoinositide 3-kinase inhibitors in combination with other therapies for the treatment of patients with hematologic malignancies. Further evaluation of phosphoinositide 3-kinase inhibitors in first-line or subsequent regimens may improve clinical outcomes. This article reviews the role of phosphoinositide 3-kinase signaling in hematologic malignancies and the potential clinical utility of inhibitors that target this pathway. PMID:24425689

  15. Regulation of AMP deaminase by phosphoinositides.

    PubMed

    Sims, B; Mahnke-Zizelman, D K; Profit, A A; Prestwich, G D; Sabina, R L; Theibert, A B

    1999-09-01

    AMP deaminase (AMPD) converts AMP to IMP and is a diverse and highly regulated enzyme that is a key component of the adenylate catabolic pathway. In this report, we identify the high affinity interaction between AMPD and phosphoinositides as a mechanism for regulation of this enzyme. We demonstrate that endogenous rat brain AMPD and the human AMPD3 recombinant enzymes specifically bind inositide-based affinity probes and to mixed lipid micelles that contain phosphatidylinositol 4,5-bisphosphate. Moreover, we show that phosphoinositides specifically inhibit AMPD catalytic activity. Phosphatidylinositol 4,5-bisphosphate is the most potent inhibitor, effecting pure noncompetitive inhibition of the wild type human AMPD3 recombinant enzyme with a K(i) of 110 nM. AMPD activity can be released from membrane fractions by in vitro treatment with neomycin, a phosphoinositide-binding drug. In addition, in vivo modulation of phosphoinositide levels leads to a change in the soluble and membrane-associated pools of AMPD activity. The predicted human AMPD3 sequence contains pleckstrin homology domains and (R/K)X(n)(R/K)XKK sequences, both of which are characterized phosphoinositide-binding motifs. The interaction between AMPD and phosphoinositides may mediate membrane localization of the enzyme and function to modulate catalytic activity in vivo. PMID:10464307

  16. Channelopathies linked to plasma membrane phosphoinositides

    PubMed Central

    Logothetis, Diomedes E.; Petrou, Vasileios I.; Adney, Scott K.; Mahajan, Rahul

    2014-01-01

    The plasma membrane phosphoinositide phosphatidylinositol 4,5-bisphosphate (PIP2) controls the activity of most ion channels tested thus far through direct electrostatic interactions. Mutations in channel proteins that change their apparent affinity to PIP2 can lead to channelopathies. Given the fundamental role that membrane phosphoinositides play in regulating channel activity, it is surprising that only a small number of channelopathies have been linked to phosphoinositides. This review proposes that for channels whose activity is PIP2-dependent and for which mutations can lead to channelopathies, the possibility that the mutations alter channel-PIP2 interactions ought to be tested. Similarly, diseases that are linked to disorders of the phosphoinositide pathway result in altered PIP2 levels. In such cases, it is proposed that the possibility for a concomitant dysregulation of channel activity also ought to be tested. The ever-growing list of ion channels whose activity depends on interactions with PIP2 promises to provide a mechanism by which defects on either the channel protein or the phosphoinositide levels can lead to disease. PMID:20396900

  17. Regulation of transient receptor potential (TRP) channels by phosphoinositides

    Microsoft Academic Search

    Tibor Rohacs; Bernd Nilius

    2007-01-01

    This review summarizes the modulation of transient receptor potential (TRP) channels, by phosphoinositides. TRP channels are\\u000a characterized by polymodal activation and a surprising complexity of regulation mechanisms. Possibly, most if not all TRP\\u000a channels are modulated by phosphoinositides. Modulation by phosphatidylinositol 4,5-biphosphate (PIP2) has been shown in detail for TRP vanilloid (TRPV) 1, TRPV5, TRP melastatin (TRPM) 4, TRPM5, TRPM7,

  18. Phosphoinositides alter lipid bilayer properties

    PubMed Central

    Hobart, E. Ashley; Koeppe, Roger E.; Andersen, Olaf S.

    2013-01-01

    Phosphatidylinositol-4,5-bisphosphate (PIP2), which constitutes ?1% of the plasma membrane phospholipid, plays a key role in membrane-delimited signaling. PIP2 regulates structurally and functionally diverse membrane proteins, including voltage- and ligand-gated ion channels, inwardly rectifying ion channels, transporters, and receptors. In some cases, the regulation is known to involve specific lipid–protein interactions, but the mechanisms by which PIP2 regulates many of its various targets remain to be fully elucidated. Because many PIP2 targets are membrane-spanning proteins, we explored whether the phosphoinositides might alter bilayer physical properties such as curvature and elasticity, which would alter the equilibrium between membrane protein conformational states—and thereby protein function. Taking advantage of the gramicidin A (gA) channels’ sensitivity to changes in lipid bilayer properties, we used gA-based fluorescence quenching and single-channel assays to examine the effects of long-chain PIP2s (brain PIP2, which is predominantly 1-stearyl-2-arachidonyl-PIP2, and dioleoyl-PIP2) on bilayer properties. When premixed with dioleoyl-phosphocholine at 2 mol %, both long-chain PIP2s produced similar changes in gA channel function (bilayer properties); when applied through the aqueous solution, however, brain PIP2 was a more potent modifier than dioleoyl-PIP2. Given the widespread use of short-chain dioctanoyl-phosphoinositides, we also examined the effects of diC8-phosphoinositol (PI), PI(4,5)P2, PI(3,5)P2, PI(3,4)P2, and PI(3,4,5)P3. The diC8 phosphoinositides, except for PI(3,5)P2, altered bilayer properties with potencies that decreased with increasing head group charge. Nonphosphoinositide diC8 phospholipids generally were more potent bilayer modifiers than the polyphosphoinositides. These results show that physiological increases or decreases in plasma membrane PIP2 levels, as a result of activation of PI kinases or phosphatases, are likely to alter lipid bilayer properties, in addition to any other effects they may have. The results further show that exogenous PIP2, as well as structural analogues that differ in acyl chain length or phosphorylation state, alters lipid bilayer properties at the concentrations used in many cell physiological experiments. PMID:23712549

  19. Phosphoinositides and Engulfment

    PubMed Central

    Swanson, Joel A.

    2015-01-01

    Cellular engulfment of particles, cells or solutes displaces large domains of plasma membrane into intracellular membranous vacuoles. This transfer of membrane is accompanied by major transitions of the phosphoinositide (PI) species that comprise the cytoplasmic face of membrane bilayers. Mapping of membrane PIs during engulfment reveals distinct patterns of protein and PI distributions associated with each stage of engulfment, which correspond with activities that regulate the actin cytoskeleton, membrane movements and vesicle secretion. Experimental manipulation of PI chemistry during engulfment indicates that PIs integrate organelle identity and orient signal transduction cascades within confined subdomains of membrane. These pathways are exploited by microbial pathogens to direct or redirect the engulfment process. PMID:25073505

  20. Rho GTPases, phosphoinositides, and actin

    PubMed Central

    Croisé, Pauline; Estay-Ahumada, Catherine; Gasman, Stéphane; Ory, Stéphane

    2014-01-01

    Rho GTPases are well known regulators of the actin cytoskeleton that act by binding and activating actin nucleators. They are therefore involved in many actin-based processes, including cell migration, cell polarity, and membrane trafficking. With the identification of phosphoinositide kinases and phosphatases as potential binding partners or effectors, Rho GTPases also appear to participate in the regulation of phosphoinositide metabolism. Since both actin dynamics and phosphoinositide turnover affect the efficiency and the fidelity of vesicle transport between cell compartments, Rho GTPases have emerged as critical players in membrane trafficking. Rho GTPase activity, actin remodeling, and phosphoinositide metabolism need to be coordinated in both space and time to ensure the progression of vesicles along membrane trafficking pathways. Although most molecular pathways are still unclear, in this review, we will highlight recent advances made in our understanding of how Rho-dependent signaling pathways organize actin dynamics and phosphoinositides and how phosphoinositides potentially provide negative feedback to Rho GTPases during endocytosis, exocytosis and membrane exchange between intracellular compartments. PMID:24914539

  1. Endosomal phosphoinositides and human diseases.

    PubMed

    Nicot, Anne-Sophie; Laporte, Jocelyn

    2008-08-01

    Phosphoinositides (PIs) are lipid second messengers implicated in signal transduction and membrane trafficking. Seven distinct PIs can be synthesized by phosphorylation of the inositol ring of phosphatidylinositol (PtdIns), and their metabolism is accurately regulated by PI kinases and phosphatases. Two of the PIs, PtdIns3P and PtdIns(3,5)P(2), are present on intracellular endosomal compartments, and several studies suggest that they have a role in membrane remodeling and trafficking. We refer to them as 'endosomal PIs'. An increasing number of human genetic diseases including myopathy and neuropathies are associated to mutations in enzymes regulating the turnover of these endosomal PIs. The PtdIns3P and PtdIns(3,5)P(2) 3-phosphatase myotubularin gene is mutated in X-linked centronuclear myopathy, whereas its homologs MTMR2 and MTMR13 and the PtdIns(3,5)P(2) 5-phosphatase SAC3/FIG4 are implicated in Charcot-Marie-Tooth peripheral neuropathies. Mutations in the gene encoding the PtdIns3P 5-kinase PIP5K3/PIKfyve have been found in patients affected with François-Neetens fleck corneal dystrophy. This review presents the roles of the endosomal PIs and their regulators and proposes defects of membrane remodeling as a common pathological mechanism for the corresponding diseases. PMID:18429927

  2. IL-21 promotes CD4 T cell responses by phosphatidylinositol 3-kinase-dependent upregulation of CD86 on B cells.

    PubMed

    Attridge, Kesley; Kenefeck, Rupert; Wardzinski, Lukasz; Qureshi, Omar S; Wang, Chun Jing; Manzotti, Claire; Okkenhaug, Klaus; Walker, Lucy S K

    2014-03-01

    The cytokine IL-21 is a potent immune modulator with diverse mechanisms of action on multiple cell types. IL-21 is in clinical use to promote tumor rejection and is an emerging target for neutralization in the setting of autoimmunity. Despite its clinical potential, the biological actions of IL-21 are not yet fully understood and the full range of effects of this pleiotropic cytokine are still being uncovered. In this study, we identify a novel role for IL-21 as an inducer of the costimulatory ligand CD86 on B lymphocytes. CD86 provides critical signals through T cell-expressed CD28 that promote T cell activation in response to Ag engagement. Expression levels of CD86 are tightly regulated in vivo, being actively decreased by regulatory T cells and increased in response to pathogen-derived signals. In this study, we demonstrate that IL-21 can trigger potent and sustained CD86 upregulation through a STAT3 and PI3K-dependent mechanism. We show that elevated CD86 expression has functional consequences for the magnitude of CD4 T cell responses both in vitro and in vivo. These data pinpoint CD86 upregulation as an additional mechanism by which IL-21 can elicit immunomodulatory effects. PMID:24470500

  3. The anti-apoptotic effect of IGF-1 on tissue resident stem cells is mediated via PI3-kinase dependent secreted frizzled related protein 2 (Sfrp2) release

    SciTech Connect

    Gehmert, Sebastian; Sadat, Sanga; Song Yaohua; Yan Yasheng [Department of Molecular Pathology, University of Texas M.D. Anderson Cancer Center, SCRB2, Box 951, 7435 Fannin Street, Houston, TX 77030 (United States); Alt, Eckhard [Department of Molecular Pathology, University of Texas M.D. Anderson Cancer Center, SCRB2, Box 951, 7435 Fannin Street, Houston, TX 77030 (United States)], E-mail: ealt@mdanderson.org

    2008-07-11

    Previous studies suggest that IGF-1 may be used as an adjuvant to stem cell transfer in order to improve cell engraftment in ischemic tissue. In the current study, we investigated the effect of IGF-1 on serum deprivation and hypoxia induced stem cell apoptosis and the possible mechanisms involved. Exposure of adipose tissue derived stem cells (ASCs) to serum deprivation and hypoxia resulted in significant apoptosis in ASC which is partially prevented by IGF-1. IGF-1's anti-apoptotic effect was abolished in ASCs transfected with Sfrp2 siRNA but not by the control siRNA. Using Western blot analysis, we demonstrated that serum deprivation and hypoxia reduced the expression of nuclear {beta}-catenin, which is reversed by IGF-1. IGF-1's effect on {beta}-catenin expression was abolished by the presence of PI3-kinase inhibitor LY294002 or in ASCs transfected with Sfrp2 siRNA. These results suggest that IGF-1, through the release of the Sfrp2, contributes to cell survival by stabilizing {beta}-catenin.

  4. Phosphoinositides in yeast: genetically tractable signalling

    Microsoft Academic Search

    Stefaan Wera; Jan C. T. Bergsma; Johan M. Thevelein

    2001-01-01

    Research on signalling through phosphoinositides has made tremendous advances over the last few years. Studies with budding yeast (Saccharomyces cerevisiae) combine the advantage of a eukaryotic system with those of a rapidly growing, genetically modifiable and tractable organism of which the genome is fully sequenced. Hence, despite some differences in phosphoinositide signalling between mammals and yeast (e.g. the absence of

  5. The Phosphoinositide 3-Kinase Pathway

    NSDL National Science Digital Library

    Lewis Cantley (Beth Israel Deaconess Medical Center; Department of Cell Biology, Harvard Medical School and Division of Signal Transduction)

    2002-05-31

    Phosphorylated lipids are produced at cellular membranes during signaling events and contribute to the recruitment and activation of various signaling components. The role of phosphoinositide 3-kinase (PI3K), which catalyzes the production of phosphatidylinositol-3,4,5-trisphosphate, in cell survival pathways; the regulation of gene expression and cell metabolism; and cytoskeletal rearrangements are highlighted. The PI3K pathway is implicated in human diseases including diabetes and cancer, and understanding the intricacies of this pathway may provide new avenues for therapuetic intervention.

  6. Stress-ING Out: Phosphoinositides Mediate the Cellular Stress Response

    NSDL National Science Digital Library

    Matthew W. Bunce (University of Wisconsin; Department of Pharmacology REV)

    2006-11-07

    Phosphoinositides regulate numerous cellular processes required for growth, proliferation, and motility. Whereas phosphoinositide signal transduction pathways within the cytosol have been well characterized, nuclear signaling pathways remain poorly understood. Accumulating experimental data have now started to uncover critical functions for nuclear phosphoinositides. In particular, phosphoinositides modulate the activity of the tumor suppressor protein ING2 in response to extracellular stress. These findings highlight a previously uncharacterized function for phosphoinositides and implicate their metabolism in signaling pathways critical for cell survival.

  7. Structure and function of phosphoinositide 3-kinases

    Microsoft Academic Search

    Matthias P Wymann; Luciano Pirola

    1998-01-01

    Phosphoinositide kinases (PI3Ks) play an important role in mitogenic signaling and cell survival, cytoskeletal remodeling, metabolic control and vesicular trafficking. Here we summarize the structure–function relationships delineating the activation process of class I PI3Ks involving various domains of adapter subunits, Ras, and interacting proteins. The resulting product, PtdIns(3,4,5)P3, targets Akt\\/protein kinase B (PKB), Bruton’s tyrosine kinase (Btk), phosphoinositide-dependent kinases (PDK),

  8. BIN1/M-Amphiphysin2 induces clustering of phosphoinositides to recruit its downstream partner dynamin

    NASA Astrophysics Data System (ADS)

    Picas, Laura; Viaud, Julien; Schauer, Kristine; Vanni, Stefano; Hnia, Karim; Fraisier, Vincent; Roux, Aurélien; Bassereau, Patricia; Gaits-Iacovoni, Frédérique; Payrastre, Bernard; Laporte, Jocelyn; Manneville, Jean-Baptiste; Goud, Bruno

    2014-12-01

    Phosphoinositides play a central role in many physiological processes by assisting the recruitment of proteins to membranes through specific phosphoinositide-binding motifs. How this recruitment is coordinated in space and time is not well understood. Here we show that BIN1/M-Amphiphysin2, a protein involved in T-tubule biogenesis in muscle cells and frequently mutated in centronuclear myopathies, clusters PtdIns(4,5)P2 to recruit its downstream partner dynamin. By using several mutants associated with centronuclear myopathies, we find that the N-BAR and the SH3 domains of BIN1 control the kinetics and the accumulation of dynamin on membranes, respectively. We show that phosphoinositide clustering is a mechanism shared by other proteins that interact with PtdIns(4,5)P2, but do not contain a BAR domain. Our numerical simulations point out that clustering is a diffusion-driven process in which phosphoinositide molecules are not sequestered. We propose that this mechanism plays a key role in the recruitment of downstream phosphoinositide-binding proteins.

  9. Phosphoinositide control of membrane protein function: a frontier led by studies on ion channels.

    PubMed

    Logothetis, Diomedes E; Petrou, Vasileios I; Zhang, Miao; Mahajan, Rahul; Meng, Xuan-Yu; Adney, Scott K; Cui, Meng; Baki, Lia

    2015-01-01

    Anionic phospholipids are critical constituents of the inner leaflet of the plasma membrane, ensuring appropriate membrane topology of transmembrane proteins. Additionally, in eukaryotes, the negatively charged phosphoinositides serve as key signals not only through their hydrolysis products but also through direct control of transmembrane protein function. Direct phosphoinositide control of the activity of ion channels and transporters has been the most convincing case of the critical importance of phospholipid-protein interactions in the functional control of membrane proteins. Furthermore, second messengers, such as [Ca(2+)]i, or posttranslational modifications, such as phosphorylation, can directly or allosterically fine-tune phospholipid-protein interactions and modulate activity. Recent advances in structure determination of membrane proteins have allowed investigators to obtain complexes of ion channels with phosphoinositides and to use computational and experimental approaches to probe the dynamic mechanisms by which lipid-protein interactions control active and inactive protein states. PMID:25293526

  10. Physical Foundations of PTEN/Phosphoinositide Interaction

    NASA Astrophysics Data System (ADS)

    Gericke, Arne; Jiang, Zhiping; Redfern, Roberta E.; Kooijman, Edgar E.; Ross, Alonzo H.

    2009-03-01

    Phosphoinositides act as signaling molecules by recruiting critical effectors to specific subcellular membranes to regulate cell proliferation, apoptosis and cytoskeletal reorganization, which requires a tight regulation of phosphoinositide generation and turnover as well as a high degree of compartmentalization. PTEN is a phosphatase specific for the 3 position of the phosophoinositide ring that is deleted or mutated in many different disease states. PTEN association with membranes requires the interaction of its C2 domain with phosphatidylserine and the interaction of its N-terminal end with phosphatidylinositol-4,5-bisphophate (PI(4,5)P2). We have investigated PTEN/PI(4,5)P2 interaction and found that Lys13 is crucial for the observed binding. We also found that the presence of cholesterol enhances PTEN binding to mixed PI(4,5)P2/POPC vesicles. Fluorescence microscopy experiments utilizing GUVs yielded results consistent with enhanced phosphoinositide domain formation in the presence of cholesterol. These experiments were accompanied by zeta potential measurements and solid state MAS ^31P-NMR experiments aimed at investigating the ionization behavior of phosphoinositides.

  11. Effectors of animal and plant pathogens use a common domain to bind host phosphoinositides.

    PubMed

    Salomon, Dor; Guo, Yirui; Kinch, Lisa N; Grishin, Nick V; Gardner, Kevin H; Orth, Kim

    2013-01-01

    Bacterial Type III Secretion Systems deliver effectors into host cells to manipulate cellular processes to the advantage of the pathogen. Many host targets of these effectors are found on membranes. Therefore, to identify their targets, effectors often use specialized membrane-localization domains to localize to appropriate host membranes. However, the molecular mechanisms used by many domains are unknown. Here we identify a conserved bacterial phosphoinositide-binding domain (BPD) that is found in functionally diverse Type III effectors of both plant and animal pathogens. We show that members of the BPD family functionally bind phosphoinositides and mediate localization to host membranes. Moreover, NMR studies reveal that the BPD of the newly identified Vibrio parahaemolyticus Type III effector VopR is unfolded in solution, but folds into a specific structure upon binding its ligand phosphatidylinositol-(4,5)-bisphosphate. Thus, our findings suggest a possible mechanism for promoting refolding of Type III effectors after delivery into host cells. PMID:24346350

  12. The chemical biology of phosphoinositide 3-kinases.

    PubMed

    Wymann, Matthias P; Schultz, Carsten

    2012-09-24

    Since its discovery in the late 1980s, phosphoinositide 3-kinase (PI3K), and its isoforms have arguably reached the forefront of signal transduction research. Regulation of this lipid kinase, its functions, its effectors, in short its entire signaling network, has been extensively studied. PI3K inhibitors are frequently used in biochemistry and cell biology. In addition, many pharmaceutical companies have launched drug-discovery programs to identify modulators of PI3Ks. Despite these efforts and a fairly good knowledge of the PI3K signaling network, we still have only a rudimentary picture of the signaling dynamics of PI3K and its lipid products in space and time. It is therefore essential to create and use novel biological and chemical tools to manipulate the phosphoinositide signaling network with spatial and temporal resolution. In this review, we discuss the current and potential future tools that are available and necessary to unravel the various functions of PI3K and its isoforms. PMID:22965647

  13. Caspase-activated phosphoinositide binding by CNT-1 promotes apoptosis by inhibiting the AKT pathway

    PubMed Central

    Nakagawa, Akihisa; Sullivan, Kelly D.; Xue, Ding

    2014-01-01

    Inactivation of cell survival factors is a crucial step in apoptosis. The phosphoinositide 3 kinase (PI3K) and AKT signaling pathway promotes cell growth, proliferation and survival and its deregulation causes cancer. How this pathway is suppressed to promote apoptosis is poorly understood. Here we report the identification of a CED-3 caspase substrate in C. elegans, CNT-1, that upon cleavage by CED-3 during apoptosis activates an N-terminal phosphoinositide-binding fragment (tCNT-1), which translocates from cytoplasm to plasma membrane to block AKT binding to phosphatidylinositol (3,4,5)-triphosphate (PIP3), thereby disabling AKT activation and its pro-survival activity. Our findings reveal a new mechanism that negatively regulates AKT cell signaling to promote apoptosis and that may restrict cell growth and proliferation in normal cells. PMID:25383666

  14. The Receptor Binding Domain of Botulinum Neurotoxin Stereotype C Binds Phosphoinositides

    SciTech Connect

    Zhang, Yanfeng; Varnum, Susan M.

    2012-03-01

    Botulinum neurotoxins (BoNTs) are the most toxic proteins known for humans and animals with an extremely low LD50 of {approx} 1 ng/kg. BoNTs generally require a protein and a ganglioside on the cell membrane surface for binding, which is known as a 'dual receptor' mechanism for host intoxication. Recent studies have suggested that in addition to gangliosides, other membrane lipids such as phosphoinositides may be involved in the interactions with the receptor binding domain (HCR) of BoNTs for better membrane penetration. Here, using two independent lipid-binding assays, we tested the interactions of BoNT/C-HCR with lipids in vitro. BoNT/C-HCR was found to bind negatively charged phospholipids, preferentially phosphoinositides. Additional interactions to phosphoinositides may help BoNT/C bind membrane more tightly and transduct signals for subsequent steps of intoxication. Our results provide new insights into the mechanisms of host cell membrane recognition by BoNTs.

  15. Urotensin-II receptor stimulation of cardiac L-type Ca2+ channels requires the ?? subunits of Gi/o-protein and phosphatidylinositol 3-kinase-dependent protein kinase C ?1 isoform.

    PubMed

    Zhang, Yuan; Ying, Jiaoqian; Jiang, Dongsheng; Chang, Zhigang; Li, Hua; Zhang, Guoqiang; Gong, Shan; Jiang, Xinghong; Tao, Jin

    2015-03-27

    Recent studies have demonstrated that urotensin-II (U-II) plays important roles in cardiovascular actions including cardiac positive inotropic effects and increasing cardiac output. However, the mechanisms underlying these effects of U-II in cardiomyocytes still remain unknown. We show by electrophysiological studies that U-II dose-dependently potentiates L-type Ca(2+) currents (ICa,L) in adult rat ventricular myocytes. This effect was U-II receptor (U-IIR)-dependent and was associated with a depolarizing shift in the voltage dependence of inactivation. Intracellular application of guanosine-5'-O-(2-thiodiphosphate) and pertussis toxin pretreatment both abolished the stimulatory effects of U-II. Dialysis of cells with the QEHA peptide, but not scrambled peptide SKEE, blocked the U-II-induced response. The phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin as well as the class I PI3K antagonist CH132799 blocked the U-II-induced ICa,L response. Protein kinase C antagonists calphostin C and chelerythrine chloride as well as dialysis of cells with 1,2bis(2aminophenoxy)ethaneN,N,N',N'-tetraacetic acid abolished the U-II-induced responses, whereas PKC? inhibition or PKA blockade had no effect. Exposure of ventricular myocytes to U-II markedly increased membrane PKC?1 expression, whereas inhibition of PKC?1 pharmacologically or by shRNA targeting abolished the U-II-induced ICa,L response. Functionally, we observed a significant increase in the amplitude of sarcomere shortening induced by U-II; blockade of U-IIR as well as PKC? inhibition abolished this effect, whereas Bay K8644 mimicked the U-II response. Taken together, our results indicate that U-II potentiates ICa,L through the ?? subunits of Gi/o-protein and downstream activation of the class I PI3K-dependent PKC?1 isoform. This occurred via the activation of U-IIR and contributes to the positive inotropic effect on cardiomyocytes. PMID:25678708

  16. Structure and function of phosphoinositide 3-kinases.

    PubMed

    Wymann, M P; Pirola, L

    1998-12-01

    Phosphoinositide kinases (PI3Ks) play an important role in mitogenic signaling and cell survival, cytoskeletal remodeling, metabolic control and vesicular trafficking. Here we summarize the structure-function relationships delineating the activation process of class I PI3Ks involving various domains of adapter subunits, Ras, and interacting proteins. The resulting product, PtdIns(3,4,5)P3, targets Akt/protein kinase B (PKB), Bruton's tyrosine kinase (Btk), phosphoinositide-dependent kinases (PDK), integrin-linked kinase (ILK), atypical protein kinases C (PKC), phospholipase Cgamma and more. Surface receptor-activated PI3Ks function in mammals, insects, nematodes and slime mold, but not yeast. While many members of the class II family have been identified and characterized biochemically, it is presently unknown how these C2-domain containing PI3Ks are activated, and which PI substrate they phosphorylate in vivo. PtdIns 3-P is produced by Vps34p/class III PI3Ks and operates via the PtdIns 3-P-binding proteins early endosomal antigen (EEA1), yeast Vac1p, Vps27p, Pip1p in lysosomal protein targeting. Besides the production of D3 phosphorylated lipids, PI3Ks have an intrinsic protein kinase activity. For trimeric GTP-binding protein-activated PI3Kgamma, protein kinase activity seems to be sufficient to trigger mitogen-activated protein kinase (MAPK). Recent disruption of PI3K genes in slime mold, Caenorhabditis elegans, Drosophila melanogaster and mice further underlines the importance of PI3K signaling systems and elucidates the role of PI3K signaling in multicellular organisms. PMID:9838078

  17. Autoradiographic imaging of phosphoinositide turnover in the brain

    SciTech Connect

    Hwang, P.M.; Bredt, D.S.; Snyder, S.H. (Johns Hopkins Univ. School of Medicine, Baltimore, MD (USA))

    1990-08-17

    With ({sup 3}H)cytidine as a precursor, phosphoinositide turnover can be localized in brain slices by selective autoradiography of the product ({sup 3}H)cytidine diphosphate diacylglycerol, which is membrane-bound. In the cerebellum, glutamatergic stimulation elicits an increase of phosphoinositide turnover only in Purkinje cells and the molecular layer. In the hippocampus, both glutamatergic and muscarinic cholinergic stimulation increase phosphoinositide turnover, but with distinct localizations. Cholinergic stimulation affects CA1, CA3, CA4, and subiculum, whereas glutamatergic effects are restricted to the subiculum and CA3. Imaging phosphoinositide turnover in brain slices, which are amenable to electrophysiologic studies, will permit a dynamic localized analysis of regulation of this second messenger in response to synaptic stimulation of specific neuronal pathways.

  18. Lipids on the move: phosphoinositide 3-kinases in leukocyte function

    Microsoft Academic Search

    Matthias P Wymann; Silvano Sozzani; Fiorella Altruda; Alberto Mantovani; Emilio Hirsch

    2000-01-01

    All four known cell surface receptor-activated phosphoinositide 3-kinases have been found in leukocytes. Matthias Wymann and colleagues discuss their nonredundant roles in leukocyte function, particularly in chemotaxis, and their promise as targets for therapeutic intervention.

  19. Phosphoinositide3-kinase regulates actin polymerization during delayed phagocytosis of Helicobacter pylori.

    PubMed

    Allen, Lee-Ann H; Allgood, J Aaron; Han, Xuemei; Wittine, Lara M

    2005-07-01

    We have shown previously that ulcerogenic (type I) strains of Helicobacter pylori (Hp) retard their entry into macrophages. However, the signaling pathways that regulate Hp phagocytosis are largely undefined. We show here that Hp strongly activated class IA phosphoinositide3-kinases (PI3Ks) in macrophages, coincident with phagocytosis, and endogenous p85 and active protein kinase Balpha accumulated on forming phagosomes. PI3K inhibitors, wortmannin and LY294002, inhibited phagocytosis of Hp in a dose-dependent manner, and blockade of engulfment correlated directly with loss of 3'-phosphoinositides in the membrane subjacent to attached bacteria. During uptake of large immunoglobulin G (IgG)-coated particles, PI3Ks regulate pseudopod extension and phagosome closure. In marked contrast, we show here that 3'-phosphoinositides regulated actin polymerization at sites of Hp uptake. Moreover, Hp and IgG beads activated distinct PI3K isoforms. Phagosomes containing IgG-coated particles accumulated 3'-phosphatase and tensin homologue deleted on chromosome 10 and Src homology 2 domain-containing inositol 5'-phosphatase, yet Hp phagosomes did not. Finally, rapid uptake of IgG-opsonized Hp or a less-virulent type II Hp was PI3K-independent. We conclude that Hp and IgG beads are ingested by distinct mechanisms and that PI3Ks regulate the actin cytoskeleton during slow phagocytosis of ulcerogenic Hp. PMID:15809290

  20. Phosphoinositide3-kinase regulates actin polymerization during delayed phagocytosis of Helicobacter pylori

    PubMed Central

    Allen, Lee-Ann H.; Allgood, J. Aaron; Han, Xuemei; Wittine, Lara M.

    2007-01-01

    We have shown previously that ulcerogenic (type I) strains of Helicobacter pylori (Hp) retard their entry into macrophages. However, the signaling pathways that regulate Hp phagocytosis are largely undefined. We show here that Hp strongly activated class IA phosphoinositide3-kinases (PI3Ks) in macrophages, coincident with phagocytosis, and endogenous p85 and active protein kinase B? accumulated on forming phagosomes. PI3K inhibitors, wortmannin and LY294002, inhibited phagocytosis of Hp in a dose-dependent manner, and blockade of engulfment correlated directly with loss of 3?-phosphoinositides in the membrane subjacent to attached bacteria. During uptake of large immunoglobulin G (IgG)-coated particles, PI3Ks regulate pseudopod extension and phagosome closure. In marked contrast, we show here that 3?-phosphoinositides regulated actin polymerization at sites of Hp uptake. Moreover, Hp and IgG beads activated distinct PI3K isoforms. Phagosomes containing IgG-coated particles accumulated 3?-phosphatase and tensin homologue deleted on chromosome 10 and Src homology 2 domain-containing inositol 5?-phosphatase, yet Hp phagosomes did not. Finally, rapid uptake of IgG-opsonized Hp or a less-virulent type II Hp was PI3K-independent. We conclude that Hp and IgG beads are ingested by distinct mechanisms and that PI3Ks regulate the actin cytoskeleton during slow phagocytosis of ulcerogenic Hp. PMID:15809290

  1. Phosphoinositide 3-kinase signalling – which way to target?

    Microsoft Academic Search

    Matthias P Wymann; Marketa Zvelebil; Muriel Laffargue

    2003-01-01

    Phosphoinositide 3-kinases (PI3Ks) are central to the control of cell growth, proliferation and survival, and drive the progression of tumours by activating phosphoinositide-dependent kinase, protein kinase B and the target of rapamycin. Other downstream effectors link PI3K to cell motility and the control of cardiovascular parameters. Current knowledge indicates that PI3Ks might qualify as drug targets for the treatment of

  2. CELLULAR AND MOLECULAR INTERACTIONS OF PHOSPHOINOSITIDES AND PERIPHERAL PROTEINS

    PubMed Central

    Stahelin, Robert V.; Scott, Jordan L.; Frick, Cary T.

    2015-01-01

    Anionic lipids act as signals for the recruitment of proteins containing cationic clusters to biological membranes. A family of anionic lipids known as the phosphoinositides (PIPs) are low in abundance, yet play a critical role in recruitment of peripheral proteins to the membrane interface. PIPs are mono-, bis-, or trisphosphorylated derivatives of phosphatidylinositol (PI) yielding seven species with different structure and anionic charge. The differential spatial distribution and temporal appearance of PIPs is key to their role in communicating information to target proteins. Selective recognition of PIPs came into play with the discovery that the substrate of protein kinase C termed pleckstrin possessed the first PIP binding region termed the pleckstrin homology (PH) domain. Since the discovery of the PH domain, more than ten PIP binding domains have been identified including PH, ENTH, FYVE, PX, and C2 domains. Representative examples of each of these domains have been thoroughly characterized to understand how they coordinate PIP headgroups in membranes, translocate to specific membrane docking sites in the cell, and function to regulate the activity of their full-length proteins. In addition, a number of novel mechanisms of PIP-mediated membrane association have emerged, such as coincidence detection – specificity for two distinct lipid headgroups. Other PIP-binding domains may also harbor selectivity for a membrane physical property such as charge or membrane curvature. This review summarizes the current understanding of the cellular distribution of PIPs and their molecular interaction with peripheral proteins. PMID:24556335

  3. Phosphoinositide metabolism and adrenergic receptors in astrocytes

    SciTech Connect

    Noble, E.P.; Ritchie, T.; de Vellis, J.

    1986-03-01

    Agonist-induced phosphoinositide (PI) breakdown functions as a signal generating system. Diacylglycerol, one breakdown product of phosphotidylinositol-4,5-diphosphate hydrolysis, can stimulate protein kinase C, whereas inositol triphosphate, the other product, has been proposed to be a second messenger for Ca/sup + +/ mobilization. Using purified astrocyte cultures from neonatal rat brain, the effects of adrenergic agonists and antagonists at 10/sup -5/ M were measured on PI breakdown. Astrocytes grown in culture were prelabeled with (/sup 3/H)inositol, and basal (/sup 3/H) inositol phosphate (IP/sub 1/) accumulation was measured in the presence of Li/sup +/. Epinephrine > norepinephrine (NE) were the most active stimulants of IP/sub 1/ production. The ..cap alpha../sub 1/ adrenoreceptor blockers, phentolamine and phenoxybenzamine, added alone had no effect on IP/sub 1/ production was reduced below basal levels. Propranolol partially blocked the effects of NE. Clonidine and isoproterenol, separately added, reduced IP/sub 1/ below basal levels and when added together diminished IP/sub 1/ accumulation even further. The role of adrenergic stimulation in the production of c-AMP.

  4. Schistosoma mansoni: characterization of phosphoinositide response.

    PubMed

    Wiest, P M; Li, Y N; Burnham, D C; Olds, G R; Bowen, W D

    1992-02-01

    Signal transduction pathways may have important regulatory roles in cellular events in the human parasite Schistosoma mansoni. The presence of the phosphoinositide response in S. mansoni was examined by radiolabeling intact worms with 20 muCi of [3H]myoinositol for 24 hr and stimulating parasites with 25 mM NaF and 10 microM AlCl3 in the presence of 10 mM LiCl. Total inositol phosphates were increased within 2 min and maximal accumulation was achieved after 30 min. Similar results were seen with the non-hydrolyzable GTP analogues GTP gamma S and GppNHp while only minimal changes were detected with GMP. Neomycin inhibited NaF-induced inositol phosphate production. NaF stimulated a significant 3.6-fold increase of inositol phosphates in females compared to males. These data suggest that stimulation of guanine nucleotide-binding regulatory proteins activates phospholipase C resulting in production of inositol phosphates in S. mansoni. PMID:1309701

  5. The emerging role of PtdIns5P: another signalling phosphoinositide takes its place.

    PubMed

    Grainger, Deborah L; Tavelis, Christodoulos; Ryan, Alexander J; Hinchliffe, Katherine A

    2012-02-01

    Of the seven phosphoinositides, PtdIns5P remains the most enigmatic. However, recent research has begun to elucidate its physiological functions. It is now clear that PtdIns5P is found in several distinct subcellular locations, and the identification of a number of PtdIns5P-binding proteins points to its involvement in a variety of key processes, including signal transduction, membrane trafficking and regulation of gene expression. Although the mechanisms that control its turnover are not yet fully understood, the existence of multiple pathways for PtdIns5P regulation is consistent with this emerging versatility. PMID:22260701

  6. Targeting phosphoinositide 3-kinase: moving towards therapy.

    PubMed

    Marone, Romina; Cmiljanovic, Vladimir; Giese, Bernd; Wymann, Matthias P

    2008-01-01

    Phosphoinositide 3-kinases (PI3K) orchestrate cell responses including mitogenic signaling, cell survival and growth, metabolic control, vesicular trafficking, degranulation, cytoskeletal rearrangement and migration. Deregulation of the PI3K pathway occurs by activating mutations in growth factor receptors or the PIK3CA locus coding for PI3Kalpha, by loss of function of the lipid phosphatase and tensin homolog deleted in chromosome ten (PTEN/MMAC/TEP1), by the up-regulation of protein kinase B (PKB/Akt), or the impairment of the tuberous sclerosis complex (TSC1/2). All these events are linked to growth and proliferation, and have thus prompted a significant interest in the pharmaceutical targeting of the PI3K pathway in cancer. Genetic targeting of PI3Kgamma (p110gamma) and PI3Kdelta (p110delta) in mice has underlined a central role of these PI3K isoforms in inflammation and allergy, as they modulate chemotaxis of leukocytes and degranulation in mast cells. Proof-of-concept molecules selective for PI3Kgamma have already successfully alleviated disease progress in murine models of rheumatoid arthritis and lupus erythematosus. As targeting PI3K moves forward to therapy of chronic, non-fatal disease, safety concerns for PI3K inhibitors increase. Many of the present inhibitor series interfere with target of rapamycin (TOR), DNA-dependent protein kinase (DNA-PK(cs)) and activity of the ataxia telangiectasia mutated gene product (ATM). Here we review the current disease-relevant knowledge for isoform-specific PI3K function in the above mentioned diseases, and review the progress of >400 recent patents covering pharmaceutical targeting of PI3K. Currently, several drugs targeting the PI3K pathway have entered clinical trials (phase I) for solid tumors and suppression of tissue damage after myocardial infarction (phases I,II). PMID:17997386

  7. Metabotropic glutamate receptor involvement in phosphoinositide hydrolysis stimulation by an endogenous Na +, K +ATPase inhibitor and ouabain in neonatal rat brain

    Microsoft Academic Search

    M. A Calvińo; C Peńa; G Rodr??guez de Lores Arnaiz

    2002-01-01

    The mechanism of action of an endogenous Na+, K+-ATPase inhibitor, termed endobain E, on phosphoinositide hydrolysis was studied in neonatal rat brain cortex and compared with that of ouabain. Lack of additivity for endobain E and glutamate paired stimulation on inositol phosphates accumulation suggested that they share at least a common step on inositol phosphate metabolism, as previously advanced for

  8. Novel Regulatory Mechanisms of mTOR Signaling

    Microsoft Academic Search

    J. Chen

    \\u000a As a master regulator of cellular processes ranging from cell growth and proliferation to differentiation, the mammalian target\\u000a of rapamycin (mTOR) is critically involved in a complex signaling network. mTOR appears to govern an amino acid sensing pathway\\u000a that integrates with a phosphatidylinositol 3-kinase-dependent mitogenic pathway to activate the downstream effectors. Recent\\u000a findings have revealed some unexpected regulatory mechanisms of

  9. Synthesis and Function of Membrane Phosphoinositides in Budding Yeast, Saccharomyces cerevisiae

    PubMed Central

    Strahl, Thomas; Thorner, Jeremy

    2007-01-01

    It is now well appreciated that derivatives of phosphatidylinositol (PtdIns) are key regulators of many cellular processes in eukaryotes. Of particular interest are phosphoinositides (mono- and polyphosphorylated adducts to the inositol ring in PtdIns), which are located at the cytoplasmic face of cellular membranes. Phosphoinositides serve both a structural and a signaling role via their recruitment of proteins that contain phosphoinositide-binding domains. Phosphoinositides also have a role as precursors of several types of second messengers for certain intracellular signaling pathways. Realization of the importance of phosphoinositides has brought increased attention to characterization of the enzymes that regulate their synthesis, interconversion, and turnover. Here we review the current state of our knowledge about the properties and regulation of the ATP-dependent lipid kinases responsible for synthesis of phosphoinositides and also the additional temporal and spatial controls exerted by the phosphatases and a phospholipase that act on phosphoinositides in yeast. PMID:17382260

  10. Nephrin mediates actin reorganization via phosphoinositide 3-kinase in podocytes

    Microsoft Academic Search

    J Zhu; N Sun; L Aoudjit; H Li; H Kawachi; S Lemay; T Takano

    2008-01-01

    Nephrin is a slit diaphragm protein critical for structural and functional integrity of visceral glomerular epithelial cells (podocytes) and is known to be tyrosine phosphorylated by Src family kinases. We studied the role of phosphoinositide 3-kinase (PI3K), activated via the phosphorylation of nephrin, in actin cytoskeletal reorganization of cultured rat podocytes. Phosphorylation of rat nephrin by the Fyn kinase markedly

  11. Sec14-nodulin proteins and the patterning of phosphoinositide landmarks for developmental control of membrane morphogenesis

    PubMed Central

    Ghosh, Ratna; de Campos, Marília K. F.; Huang, Jin; Huh, Seong K.; Orlowski, Adam; Yang, Yuan; Tripathi, Ashutosh; Nile, Aaron; Lee, Hsin-Chieh; Dynowski, Marek; Schäfer, Helen; Róg, Tomasz; Lete, Marta G.; Ahyayauch, Hasna; Alonso, Alicia; Vattulainen, Ilpo; Igumenova, Tatyana I.; Schaaf, Gabriel; Bankaitis, Vytas A.

    2015-01-01

    Polarized membrane morphogenesis is a fundamental activity of eukaryotic cells. This process is essential for the biology of cells and tissues, and its execution demands exquisite temporal coordination of functionally diverse membrane signaling reactions with high spatial resolution. Moreover, mechanisms must exist to establish and preserve such organization in the face of randomizing forces that would diffuse it. Here we identify the conserved AtSfh1 Sec14-nodulin protein as a novel effector of phosphoinositide signaling in the extreme polarized membrane growth program exhibited by growing Arabidopsis root hairs. The data are consistent with Sec14-nodulin proteins controlling the lateral organization of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) landmarks for polarized membrane morphogenesis in plants. This patterning activity requires both the PtdIns(4,5)P2 binding and homo-oligomerization activities of the AtSfh1 nodulin domain and is an essential aspect of the polarity signaling program in root hairs. Finally, the data suggest a general principle for how the phosphoinositide signaling landscape is physically bit mapped so that eukaryotic cells are able to convert a membrane surface into a high-definition lipid-signaling screen. PMID:25739452

  12. Activation of TRPV1 channels inhibits mechanosensitive Piezo channel activity by depleting membrane phosphoinositides.

    PubMed

    Borbiro, Istvan; Badheka, Doreen; Rohacs, Tibor

    2015-02-10

    Capsaicin is an activator of the heat-sensitive TRPV1 (transient receptor potential vanilloid 1) ion channels and has been used as a local analgesic. We found that activation of TRPV1 channels with capsaicin either in dorsal root ganglion neurons or in a heterologous expression system inhibited the mechanosensitive Piezo1 and Piezo2 channels by depleting phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and its precursor phosphatidylinositol 4-phosphate [PI(4)P] from the plasma membrane through Ca(2+)-induced phospholipase C? (PLC?) activation. Experiments with chemically inducible phosphoinositide phosphatases and receptor-induced activation of PLC? indicated that inhibition of Piezo channels required depletion of both PI(4)P and PI(4,5)P2. The mechanically activated current amplitudes decreased substantially in the excised inside-out configuration, where the membrane patch containing Piezo1 channels is removed from the cell. PI(4,5)P2 and PI(4)P applied to these excised patches inhibited this decrease. Thus, we concluded that Piezo channel activity requires the presence of phosphoinositides, and the combined depletion of PI(4,5)P2 and PI(4)P reduces channel activity. In addition to revealing a role for distinct membrane lipids in mechanosensitive ion channel regulation, these data suggest that inhibition of Piezo2 channels may contribute to the analgesic effect of capsaicin. PMID:25670203

  13. Sequential Activities of Phosphoinositide 3-Kinase, PKB/Akt, and Rab7 during Macropinosome Formation in Dictyostelium

    PubMed Central

    Rupper, Adam; Lee, Kyung; Knecht, David; Cardelli, James

    2001-01-01

    Macropinocytosis plays an important role in the internalization of antigens by dendritic cells and is the route of entry for many bacterial pathogens; however, little is known about the molecular mechanisms that regulate the formation or maturation of macropinosomes. Like dendritic cells, Dictyostelium amoebae are active in macropinocytosis, and various proteins have been identified that contribute to this process. As described here, microscopic analysis of null mutants have revealed that the class I phosphoinositide 3-kinases, PIK1 and PIK2, and the downstream effector protein kinase B (PKB/Akt) are important in regulating completion of macropinocytosis. Although actin-rich membrane protrusions form in these cell lines, they recede without forming macropinosomes. Imaging of cells expressing green fluorescent protein (GFP) fused to the pleckstrin homology domain (PH) of PKB (GFP-PHPKB) indicates that D3 phosphoinositides are enriched in the forming macropinocytic cup and remain associated with newly formed macropinosomes for <1 minute. A fusion protein, consisting of GFP fused to an F-actin binding domain, overlaps with GFP-PHPKB in the timing of association with forming macropinosomes. Although macropinocytosis is reduced in cells expressing dominant negative Rab7, microscopic imaging studies reveal that GFP-Rab7 associates only with formed macropinosomes at approximately the time that F-actin and D3 phosphoinositide levels decrease. These results support a model in which F-actin modulating proteins and vesicle trafficking proteins coordinately regulate the formation and maturation of macropinosomes. PMID:11553719

  14. Inhibition of phosphoinositide turnover by praziquantel in Schistosoma mansoni.

    PubMed

    Wiest, P M; Li, Y; Olds, G R; Bowen, W D

    1992-08-01

    The effect of praziquantel on phosphoinositide turnover was examined in Schistosoma mansoni to determine if this anthelminthic modulates signal transduction pathways in parasites. Adult worms were radiolabeled with [3H]myoinositol for 24 hr and total inositol phosphate levels determined in the presence of praziquantel. Praziquantel inhibited inositol phosphate turnover when activated with NaF plus AlCl3 or with the nonhydrolyzable guanine nucleotide-binding protein analogue GTP gamma S. Furthermore, praziquantel decreased basal turnover of inositol phosphates. Inhibition was seen in both male and female worms as well as in schistosomula. These data indicate that inhibition of phosphoinositide turnover may contribute to the effect of praziquantel on parasite survival within the definitive host. PMID:1321906

  15. Essential Role of Phosphoinositide Metabolism in Synaptic Vesicle Recycling

    Microsoft Academic Search

    Ottavio Cremona; Gilbert Di Paolo; Markus R Wenk; Anita Lüthi; Warren T Kim; Kohji Takei; Laurie Daniell; Yasuo Nemoto; Stephen B Shears; Richard A Flavell; David A McCormick; Pietro De Camilli

    1999-01-01

    Growing evidence suggests that phosphoinositides play an important role in membrane traffic. A polyphosphoinositide phosphatase, synaptojanin 1, was identified as a major presynaptic protein associated with endocytic coated intermediates. We report here that synaptojanin 1–deficient mice exhibit neurological defects and die shortly after birth. In neurons of mutant animals, PI(4,5)P2 levels are increased, and clathrin-coated vesicles accumulate in the cytomatrix-rich

  16. Analysis of the murine phosphoinositide 3-kinase ? gene

    Microsoft Academic Search

    Emilio Hirsch; Matthias P. Wymann; Enrico Patrucco; Emanuela Tolosano; Ginette Bulgarelli-Leva; Stefano Marengo; Mariano Rocchi; Fiorella Altruda

    2000-01-01

    Phosphoinositide 3-kinase ? is preferentially expressed in leukocytes. PI3K? is activated by ?? subunits of heterotrimeric G-proteins, which thus link seven transmembrane helix receptor activation to phosphatidylinositol (3,4,5)-trisphosphate production. Here we describe the molecular cloning of the murine PI3K? cDNA, the PI3K? gene structure, its chromosomal assignment and the analysis of promoter activity. The mouse cDNA shares 86% identity to

  17. Targeting phosphoinositide 3-kinase—Moving towards therapy

    Microsoft Academic Search

    Romina Marone; Vladimir Cmiljanovic; Bernd Giese; Matthias P. Wymann

    2008-01-01

    Phosphoinositide 3-kinases (PI3K) orchestrate cell responses including mitogenic signaling, cell survival and growth, metabolic control, vesicular trafficking, degranulation, cytoskeletal rearrangement and migration. Deregulation of the PI3K pathway occurs by activating mutations in growth factor receptors or the PIK3CA locus coding for PI3K?, by loss of function of the lipid phosphatase and tensin homolog deleted in chromosome ten (PTEN\\/MMAC\\/TEP1), by the

  18. Phosphoinositides: Tiny Lipids With Giant Impact on Cell Regulation

    PubMed Central

    2013-01-01

    Phosphoinositides (PIs) make up only a small fraction of cellular phospholipids, yet they control almost all aspects of a cell's life and death. These lipids gained tremendous research interest as plasma membrane signaling molecules when discovered in the 1970s and 1980s. Research in the last 15 years has added a wide range of biological processes regulated by PIs, turning these lipids into one of the most universal signaling entities in eukaryotic cells. PIs control organelle biology by regulating vesicular trafficking, but they also modulate lipid distribution and metabolism via their close relationship with lipid transfer proteins. PIs regulate ion channels, pumps, and transporters and control both endocytic and exocytic processes. The nuclear phosphoinositides have grown from being an epiphenomenon to a research area of its own. As expected from such pleiotropic regulators, derangements of phosphoinositide metabolism are responsible for a number of human diseases ranging from rare genetic disorders to the most common ones such as cancer, obesity, and diabetes. Moreover, it is increasingly evident that a number of infectious agents hijack the PI regulatory systems of host cells for their intracellular movements, replication, and assembly. As a result, PI converting enzymes began to be noticed by pharmaceutical companies as potential therapeutic targets. This review is an attempt to give an overview of this enormous research field focusing on major developments in diverse areas of basic science linked to cellular physiology and disease. PMID:23899561

  19. Phosphoinositide and Erk signaling pathways mediate activity-driven rodent olfactory sensory neuronal survival and stress mitigation.

    PubMed

    Kim, So Yeun; Mammen, Alex; Yoo, Seung-Jun; Cho, Bongki; Kim, Eun-Kyoung; Park, Jong-In; Moon, Cheil; Ronnett, Gabriele V

    2015-08-01

    Olfactory sensory neurons (OSNs) are the initial site for olfactory signal transduction. Therefore, their survival is essential to olfactory function. In the current study, we demonstrated that while odorant stimulation promoted rodent OSN survival, it induced generation of reactive oxygen species in a dose- and time-dependent manner as well as loss of membrane potential and fragmentation of mitochondria. The MEK-Erk pathway played a critical role in mediating these events, as its inhibition decreased odorant stimulation-dependent OSN survival and exacerbated intracellular stress measured by reactive oxygen species generation and heat-shock protein 70 expression. The phosphoinositide pathway, rather than the cyclic AMP pathway, mediated the odorant-induced activation of the MEK-Erk pathway. These findings provide important insights into the mechanisms of activity-driven OSN survival, the role of the phosphoinositide pathway in odorant signaling, and demonstrate that odorant detection and odorant stimulation-mediated survival proceed via independent signaling pathways. This mechanism, which permits independent regulation of odorant detection from survival signaling, may be advantageous if not diminished by repeated or prolonged odor exposure. We investigated the role of odorant stimulation in generating cellular stress and the molecular mechanisms mitigating such stress and promoting neuronal survival. Odorant stimulation promoted olfactory sensory neuron (OSN) survival and also induced intracellular oxidative stress, which was exacerbated when MEK/Erks pathway was inhibited. Sensory stimulation simultaneously activated at least two parallel pathways, the AC/cAMP cascade responsible for odorant detection, and phosphoinositide hydrolysis to promote odorant stimulation-dependent neuronal survival odorants may activate parallel signaling cascades to mediate sensory detection and sensory stimulation-dependent survival. AC, adenylyl cyclase; cAMP, cyclic adenosine monophosphate; Erk, extracellular signal-regulated kinase; MEK, MAPK/ERK kinase. PMID:25903517

  20. Phosphoinositide signalling links O-GlcNAc transferase to insulin resistance

    E-print Network

    Cai, Long

    ARTICLES Phosphoinositide signalling links O-GlcNAc transferase to insulin resistance Xiaoyong Yang unrecognized type of phosphoinositide-binding domain. After induction with insulin, phosphatidylinositol 3 modification of the insulin signalling pathway by O-GlcNAc. This results in the alteration in phosphorylation

  1. Antidepressants reduce phosphoinositide-specific phospholipase C (PI-PLC) activity and the mRNA and protein expression of selective PLC ? 1 isozyme in rat brain

    Microsoft Academic Search

    Y Dwivedi; A. K Agrawal; H. S Rizavi; G. N Pandey

    2002-01-01

    In order to examine whether antidepressants mediate their action by interacting with one of the key components of the phosphoinositide (PI) signaling pathway, i.e. PI-specific phospholipase C (PLC), and whether this represents a common mechanism of action of antidepressants, we determined the effects of antidepressants of various classes on PI-PLC activity and on the expression of PLC isozymes in rat

  2. Odorant-stimulated phosphoinositide signaling in mammalian olfactory receptor neurons

    PubMed Central

    Klasen, K.; Corey, E.A.; Kuck, F.; Wetzel, C.H.; Hatt, H.; Ache, B.W.

    2009-01-01

    Recent evidence has revived interest in the idea that phosphoinositides (PIs) may play a role in signal transduction in mammalian olfactory receptor neurons (ORNs). To provide direct evidence that odorants indeed activate PI signaling in ORNs, we used adenoviral vectors carrying two different fluorescently tagged probes, the pleckstrin homology (PH) domains of phospholipase C?1 (PLC?1) and the general receptor of phosphoinositides (GRP1), to monitor PI activity in the dendritic knobs of ORNs in vivo. Odorants mobilized PI(4,5)P2/IP3 and PI(3,4,5)P3, the substrates and products of PLC and PI3K. We then measured odorant activation of PLC and PI3K in olfactory ciliary-enriched membranes in vitro using a phospholipid overlay assay and ELISAs. Odorants activated both PLC and PI3K in the olfactory cilia within 2 sec of odorant stimulation. Odorant-dependent activation of PLC and PI3K in the olfactory epithelium could be blocked by enzyme-specific inhibitors. Odorants activated PLC and PI3K with partially overlapping specificity. These results provide direct evidence that odorants indeed activate PI signaling in mammalian ORNs in a manner that is consistent with the idea that PI signaling plays a role in olfactory transduction. PMID:19781634

  3. Unique cell wall abnormalities in the putative phosphoinositide phosphatase mutant AtSAC9

    Microsoft Academic Search

    Almut H. VollmerNabil; Nabil N. Youssef; Daryll B. DeWald

    SAC9 is a putative phosphoinositide phosphatase in Arabidopsis thaliana involved in phosphoinositide signaling. sac9-1 plants have a constitutively stressed phenotype with shorter roots which notably accumulate phosphatidylinositol 4,5-bisphosphate\\u000a and its hydrolysis product inositol trisphosphate. We investigated the primary roots of sac9-1 seedlings at the cytological and ultrastructural level to determine the structural basis for this altered growth. Despite\\u000a the normal

  4. Multiple roles for Plasmodium berghei phosphoinositide-specific phospholipase C in regulating gametocyte activation and differentiation

    PubMed Central

    Raabe, Andreas C; Wengelnik, Kai; Billker, Oliver; Vial, Henri J

    2011-01-01

    Critical events in the life cycle of malaria parasites are controlled by calcium-dependent signalling cascades, yet the molecular mechanisms of calcium release remain poorly understood. The synchronized development of Plasmodium berghei gametocytes relies on rapid calcium release from internal stores within 10 s of gametocytes being exposed to mosquito-derived xanthurenic acid (XA). Here we addressed the function of phosphoinositide-specific phospholipase C (PI-PLC) for regulating gametocyte activation. XA triggered the hydrolysis of PIP2 and the production of the secondary messenger IP3 in gametocytes. Both processes were selectively blocked by a PI-PLC inhibitor, which also reduced the early Ca2+ signal. However, microgametocyte differentiation into microgametes was blocked even when the inhibitor was added up to 5 min after activation, suggesting a requirement for PI-PLC beyond the early mobilization of calcium. In contrast, inhibitors of calcium release through ryanodine receptor channels were active only during the first minute of gametocyte activation. Biochemical determination of PI-PLC activity was confirmed using transgenic parasites expressing a fluorescent PIP2/IP3 probe that translocates from the parasite plasmalemma to the cytosol upon cell activation. Our study revealed a complex interdependency of Ca2+ and PI-PLC activity, with PI-PLC being essential throughout gamete formation, possibly explaining the irreversibility of this process. PMID:21518218

  5. Inhibition of phosphoinositide 3-kinase ? attenuates inflammation, obesity, and cardiovascular risk factors.

    PubMed

    Wymann, Matthias P; Solinas, Giovanni

    2013-03-01

    Phosphoinositide 3-kinase ? (PI3K?) plays a central role in inflammation, allergy, cardiovascular, and metabolic disease. Obesity is accompanied by chronic, low-grade inflammation. As PI3K? plays a major role in leukocyte recruitment, targeting of PI3K? has been considered to be a strategy for attenuating progression of obesity to insulin resistance and type 2 diabetes. Indeed, PI3K? null mice are protected from high fat diet-induced obesity, metabolic inflammation, fatty liver, and insulin resistance. The lean phenotype of the PI3K?-null mice has been linked to increased thermogenesis and energy expenditure. Surprisingly, the increase in fat mass and metabolic aberrations were not linked to PI3K? activity in the hematopoietic compartment. Thermogenesis and oxygen consumption are modulated by PI3K? lipid kinase-dependent and -independent signaling mechanisms. PI3K? signaling controls metabolic and inflammatory stress, and may provide an entry point for therapeutic strategies in metabolic disease, inflammation, and cardiovascular disease. PMID:23551103

  6. Airway inflammation: chemokine-induced neutrophilia and the class I phosphoinositide 3-kinases.

    PubMed

    Thomas, Matthew J; Smith, Alexandra; Head, Denise H; Milne, Laura; Nicholls, Andrew; Pearce, Wayne; Vanhaesebroeck, Bart; Wymann, Matthias P; Hirsch, Emilio; Trifilieff, Alexandre; Walker, Christoph; Finan, Peter; Westwick, John

    2005-04-01

    Class I phosphoinositide 3-kinases (PI3K) are known to play a significant role in neutrophil chemotaxis. However, the relative contributions of different PI3K isoforms, and how these impact on lung inflammation, have not been addressed. In vitro studies using wild-type and PI3Kgamma knockout neutrophils demonstrated the major role of the gamma isoform in chemotactic but not chemokinetic events. This was confirmed by a model of direct chemokine instillation into the airways in vivo. Within all studies, a low yet significant degree of neutrophil movement in the absence of PI3Kgamma could be observed. No role for the delta isoform was demonstrated both in vitro and in vivo using PI3Kdelta kinase-dead knock-in mice. Moreover, further studies using the broad-spectrum PI3K inhibitors wortmannin or LY294002 showed no other class I PI3K isoforms to be involved in these chemotactic processes. Here, we identify a contributory PI3K-independent mechanism of neutrophil movement, yet demonstrate PI3Kgamma as the pivotal mediator through which the majority of neutrophils migrate into the lung in response to chemokines. These data resolve the complexities of chemokine-induced neutrophilia and PI3K signaling and define the gamma isoform as a promising target for new therapeutics to treat airway inflammatory diseases. PMID:15739165

  7. Essential role of phosphoinositide 3-kinase gamma in eosinophil chemotaxis within acute pulmonary inflammation

    PubMed Central

    Thomas, Matthew; Edwards, Matthew J; Sawicka, Elzbieta; Duggan, Nicholas; Hirsch, Emilio; Wymann, Matthias P; Owen, Charles; Trifilieff, Alexandre; Walker, Christoph; Westwick, John; Finan, Peter

    2009-01-01

    We and others have established an important role for phosphoinositide-3 kinase gamma (PI3K?) in the chemotactic responses of macrophages and neutrophils. The involvement of this lipid kinase in allergic inflammatory responses is, however, yet to be fully determined. Here we compare wild-type (WT) and PI3K??/? (KO) mice within a model of ovalbumin (OVA) -specific pulmonary inflammation. Upon OVA aerosol challenge, cell influx into the bronchoalveolar lavage (BAL) fluid consisted of neutrophils, macrophages and, more significantly, eosinophils – which are key effector cells in allergic inflammation. Each population was reduced by up to 80% in KO mice, demonstrating a role for PI3K? in cell infiltration into the airways. The mechanism of reduced eosinophilia was analysed within both development and effector stages of the immune response. Comparable levels of OVA-specific T-cell proliferation and immunoglobulin production were established in both strains. Furthermore, no significant differences between WT and KO chemokine production were observed. Having identified the critical point of PI3K? involvement, KO eosinophil chemotactic dysfunction was confirmed in vitro. These data are the first to demonstrate the vital role of PI3K? in acute allergic inflammation. The profound dependency of eosinophils on PI3K? for pulmonary influx identifies this lipid kinase as an attractive target for the pharmacological intervention of asthma. PMID:18754810

  8. Essential role of phosphoinositide 3-kinase gamma in eosinophil chemotaxis within acute pulmonary inflammation.

    PubMed

    Thomas, Matthew; Edwards, Matthew J; Sawicka, Elzbieta; Duggan, Nicholas; Hirsch, Emilio; Wymann, Matthias P; Owen, Charles; Trifilieff, Alexandre; Walker, Christoph; Westwick, John; Finan, Peter

    2009-03-01

    We and others have established an important role for phosphoinositide-3 kinase gamma (PI3Kgamma) in the chemotactic responses of macrophages and neutrophils. The involvement of this lipid kinase in allergic inflammatory responses is, however, yet to be fully determined. Here we compare wild-type (WT) and PI3Kgamma(-/-) (KO) mice within a model of ovalbumin (OVA) -specific pulmonary inflammation. Upon OVA aerosol challenge, cell influx into the bronchoalveolar lavage (BAL) fluid consisted of neutrophils, macrophages and, more significantly, eosinophils - which are key effector cells in allergic inflammation. Each population was reduced by up to 80% in KO mice, demonstrating a role for PI3Kgamma in cell infiltration into the airways. The mechanism of reduced eosinophilia was analysed within both development and effector stages of the immune response. Comparable levels of OVA-specific T-cell proliferation and immunoglobulin production were established in both strains. Furthermore, no significant differences between WT and KO chemokine production were observed. Having identified the critical point of PI3Kgamma involvement, KO eosinophil chemotactic dysfunction was confirmed in vitro. These data are the first to demonstrate the vital role of PI3Kgamma in acute allergic inflammation. The profound dependency of eosinophils on PI3Kgamma for pulmonary influx identifies this lipid kinase as an attractive target for the pharmacological intervention of asthma. PMID:18754810

  9. Neural stem cell targeting of glioma is dependent on phosphoinositide 3-kinase signaling.

    PubMed

    Kendall, Stephen E; Najbauer, Joseph; Johnston, Heather F; Metz, Marianne Z; Li, Shan; Bowers, Marisa; Garcia, Elizabeth; Kim, Seung U; Barish, Michael E; Aboody, Karen S; Glackin, Carlotta A

    2008-06-01

    The utility of neural stem cells (NSCs) has extended beyond regenerative medicine to targeted gene delivery, as NSCs possess an inherent tropism to solid tumors, including invasive gliomas. However, for optimal clinical implementation, an understanding of the molecular events that regulate NSC tumor tropism is needed to ensure their safety and to maximize therapeutic efficacy. We show that human NSC lines responded to multiple tumor-derived growth factors and that hepatocyte growth factor (HGF) induced the strongest chemotactic response. Gliomatropism was critically dependent on c-Met signaling, as short hairpin RNA-mediated ablation of c-Met significantly attenuated the response. Furthermore, inhibition of Ras-phosphoinositide 3-kinase (PI3K) signaling impaired the migration of human neural stem cells (hNSCs) toward HGF and other growth factors. Migration toward tumor cells is a highly regulated process, in which multiple growth factor signals converge on Ras-PI3K, causing direct modification of the cytoskeleton. The signaling pathways that regulate hNSC migration are similar to those that promote unregulated glioma invasion, suggesting shared cellular mechanisms and responses. Disclosure of potential conflicts of interest is found at the end of this article. PMID:18339768

  10. Role of phosphoinositide 3-OH kinase p110? in skeletal myogenesis.

    PubMed

    Matheny, Ronald W; Riddle-Kottke, Melissa A; Leandry, Luis A; Lynch, Christine M; Abdalla, Mary N; Geddis, Alyssa V; Piper, David R; Zhao, Jean J

    2015-04-01

    Phosphoinositide 3-OH kinase (PI3K) regulates a number of developmental and physiologic processes in skeletal muscle; however, the contributions of individual PI3K p110 catalytic subunits to these processes are not well-defined. To address this question, we investigated the role of the 110-kDa PI3K catalytic subunit ? (p110?) in myogenesis and metabolism. In C2C12 cells, pharmacological inhibition of p110? delayed differentiation. We next generated mice with conditional deletion of p110? in skeletal muscle (p110? muscle knockout [p110?-mKO] mice). While young p110?-mKO mice possessed a lower quadriceps mass and exhibited less strength than control littermates, no differences in muscle mass or strength were observed between genotypes in old mice. However, old p110?-mKO mice were less glucose tolerant than old control mice. Overexpression of p110? accelerated differentiation in C2C12 cells and primary human myoblasts through an Akt-dependent mechanism, while expression of kinase-inactive p110? had the opposite effect. p110? overexpression was unable to promote myoblast differentiation under conditions of p110? inhibition, but expression of p110? was able to promote differentiation under conditions of p110? inhibition. These findings reveal a role for p110? during myogenesis and demonstrate that long-term reduction of skeletal muscle p110? impairs whole-body glucose tolerance without affecting skeletal muscle size or strength in old mice. PMID:25605332

  11. The phosphoinositide kinase PIKfyve mediates epidermal growth factor receptor trafficking to the nucleus.

    PubMed

    Kim, Jayoung; Jahng, Wan Jin; Di Vizio, Dolores; Lee, Julie S; Jhaveri, Raj; Rubin, Mark A; Shisheva, Assia; Freeman, Michael R

    2007-10-01

    ErbB receptor tyrosine kinases can transit to nuclei in tumor cells, where they have been shown to regulate gene expression as components of transcriptional complexes. Quantitative analysis of a human bladder cancer tissue microarray identified nuclear epidermal growth factor receptor (EGFR) in tumor cells and also showed an increased frequency of this histologic feature in cancer relative to normal tissues. This observation suggests a potential role for nuclear EGFR in bladder cancer. We confirmed that EGFR could be induced to transit to nuclei in cultured human bladder cancer cells in response to the urothelial cell growth factor and EGFR ligand heparin-binding EGF-like growth factor (HB-EGF). Mass spectrometric analysis of EGFR immune complexes from a transitional carcinoma cell line (TCCSUP) identified the phosphoinositide kinase, PIKfyve, as a potential component of the EGFR trafficking mechanism. RNA silencing indicated that PIKfyve is a mediator of HB-EGF-stimulated EGFR nuclear trafficking, EGFR binding to the cyclin D1 promoter, and cell cycle progression. These results identify a novel mediator of the EGFR transcription function and further suggest that nuclear EGFR and the lipid kinase PIKfyve may play a role in bladder oncogenesis. PMID:17909029

  12. Kappa opioid receptors stimulate phosphoinositide turnover in rat brain

    SciTech Connect

    Periyasamy, S.; Hoss, W. (Univ. of Toledo, OH (USA))

    1990-01-01

    The effects of various subtype-selective opioid agonists and antagonists on the phosphoinositide (PI) turnover response were investigated in the rat brain. The {kappa}-agonists U-50,488H and ketocyclazocine produced a concentration-dependent increase in the accumulation of IP's in hippocampal slices. The other {kappa}-agonists Dynorphin-A (1-13) amide, and its protected analog D(Ala){sup 2}-dynorphin-A (1-13) amide also produced a significant increase in the formation of ({sup 3}H)-IP's, whereas the {mu}-selective agonists (D-Ala{sup 2}-N-Me-Phe{sup 4}-Gly{sup 5}-ol)-enkephalin and morphine and the {delta}-selective agonist (D-Pen{sup 2,5})-enkephalin were ineffective. The increase in IP's formation elicited by U-50,488H was partially antagonized by naloxone and more completely antagonized by the {kappa}-selective antagonists nor-binaltorphimine and MR 2266. The formation of IP's induced by U-50,488H varies with the regions of the brain used, being highest in hippocampus and amygdala, and lowest in striatum and pons-medullar. The results indicate that brain {kappa}- but neither {mu}- nor {delta}- receptors are coupled to the PI turnover response.

  13. Analysis of the murine phosphoinositide 3-kinase gamma gene.

    PubMed

    Hirsch, E; Wymann, M P; Patrucco, E; Tolosano, E; Bulgarelli-Leva, G; Marengo, S; Rocchi, M; Altruda, F

    2000-10-01

    Phosphoinositide 3-kinase gamma is preferentially expressed in leukocytes. PI3Kgamma is activated by betagamma subunits of heterotrimeric G-proteins, which thus link seven transmembrane helix receptor activation to phosphatidylinositol (3,4,5)-trisphosphate production. Here we describe the molecular cloning of the murine PI3Kgamma cDNA, the PI3Kgamma gene structure, its chromosomal assignment and the analysis of promoter activity. The mouse cDNA shares 86% identity to its pig and human orthologues at the nucleotide level. The MmPI3Kgamma gene spans approximately 30kb and comprises 11 exons. RACE-PCR indicated the presence of multiple start sites generating 5' UTRs with different lengths, the longest being 874bp. The putative promoter region contains no TATA box but several putative binding sites for hematopoietic specific transcription factors. A 1200bp long sequence upstream the first transcription start site was found to possess tissue specific promoter activity. Deletion constructs revealed two contiguous regions, with activator function, ranging from positions -139 to -557, and with inhibitory function, ranging from positions -557 to -892. FISH analysis revealed that the MmPI3Kgamma is located on chromosome 12 band B and that the human orthologue is positioned on chromosome 7q22.2-22.3. In spite of some differences in the ATP-binding site, recombinant murine PI3Kgamma protein is equally sensitive to wortmannin as its human counterpart. This suggests that mouse models will provide reliable results in the assessments of novel PI3Kgamma inhibitors. PMID:11054537

  14. Transient receptor potential melastatin 3 is a phosphoinositide-dependent ion channel.

    PubMed

    Badheka, Doreen; Borbiro, Istvan; Rohacs, Tibor

    2015-07-01

    Phosphoinositides are emerging as general regulators of the functionally diverse transient receptor potential (TRP) ion channel family. Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) has been reported to positively regulate many TRP channels, but in several cases phosphoinositide regulation is controversial. TRP melastatin 3 (TRPM3) is a heat-activated ion channel that is also stimulated by chemical agonists, such as pregnenolone sulfate. Here, we used a wide array of approaches to determine the effects of phosphoinositides on TRPM3. We found that channel activity in excised inside-out patches decreased over time (rundown), an attribute of PI(4,5)P2-dependent ion channels. Channel activity could be restored by application of either synthetic dioctanoyl (diC8) or natural arachidonyl stearyl (AASt) PI(4,5)P2. The PI(4,5)P2 precursor phosphatidylinositol 4-phosphate (PI(4)P) was less effective at restoring channel activity. TRPM3 currents were also restored by MgATP, an effect which was inhibited by two different phosphatidylinositol 4-kinase inhibitors, or by pretreatment with a phosphatidylinositol-specific phospholipase C (PI-PLC) enzyme, indicating that MgATP acted by generating phosphoinositides. In intact cells, reduction of PI(4,5)P2 levels by chemically inducible phosphoinositide phosphatases or a voltage-sensitive 5'-phosphatase inhibited channel activity. Activation of PLC via muscarinic receptors also inhibited TRPM3 channel activity. Overall, our data indicate that TRPM3 is a phosphoinositide-dependent ion channel and that decreasing PI(4,5)P2 abundance limits its activity. As all other members of the TRPM family have also been shown to require PI(4,5)P2 for activity, our data establish PI(4,5)P2 as a general positive cofactor of this ion channel subfamily. PMID:26123195

  15. Genome-Wide Analysis of the Phosphoinositide Kinome from Two Ciliates Reveals Novel Evolutionary Links for Phosphoinositide Kinases in Eukaryotic Cells

    PubMed Central

    Leondaritis, George; Siokos, John; Skaripa, Irini; Galanopoulou, Dia

    2013-01-01

    Background The complexity of phosphoinositide signaling in higher eukaryotes is partly due to expansion of specific families and types of phosphoinositide kinases (PIKs) that can generate all phosphoinositides via multiple routes. This is particularly evident in the PI3Ks and PIPKs, and it is considered an evolutionary trait associated with metazoan diversification. Yet, there are limited comprehensive studies on the PIK repertoire of free living unicellular organisms. Methodology/Principal Findings We undertook a genome-wide analysis of putative PIK genes in two free living ciliated cells, Tetrahymena and Paramecium. The Tetrahymena thermophila and Paramecium tetraurelia genomes were probed with representative kinases from all families and types. Putative homologs were verified by EST, microarray and deep RNA sequencing database searches and further characterized for domain structure, catalytic efficiency, expression patterns and phylogenetic relationships. In total, we identified and characterized 22 genes in the Tetrahymena thermophila genome and 62 highly homologues genes in Paramecium tetraurelia suggesting a tight evolutionary conservation in the ciliate lineage. Comparison to the kinome of fungi reveals a significant expansion of PIK genes in ciliates. Conclusions/Significance Our study highlights four important aspects concerning ciliate and other unicellular PIKs. First, ciliate-specific expansion of PI4KIII-like genes. Second, presence of class I PI3Ks which, at least in Tetrahymena, are associated with a metazoan-type machinery for PIP3 signaling. Third, expansion of divergent PIPK enzymes such as the recently described type IV transmembrane PIPKs. Fourth, presence of possible type II PIPKs and presumably inactive PIKs (hence, pseudo-PIKs) not previously described. Taken together, our results provide a solid framework for future investigation of the roles of PIKs in ciliates and indicate that novel functions and novel regulatory pathways of phosphoinositides may be more widespread than previously thought in unicellular organisms. PMID:24244373

  16. Ras is an indispensable coregulator of the class IB phosphoinositide 3-kinase p87/p110?

    PubMed Central

    Kurig, Barbara; Shymanets, Aliaksei; Bohnacker, Thomas; Prajwal; Brock, Carsten; Ahmadian, Mohammad Reza; Schaefer, Michael; Gohla, Antje; Harteneck, Christian; Wymann, Matthias P.; Jeanclos, Elisabeth; Nürnberg, Bernd

    2009-01-01

    Class IB phosphoinositide 3-kinase ? (PI3K?) elicits various immunologic and cardiovascular responses; however, the molecular basis for this signal heterogeneity is unclear. PI3K? consists of a catalytic p110? and a regulatory p87PIKAP (p87, also p84) or p101 subunit. Hitherto p87 and p101 are generally assumed to exhibit redundant functions in receptor-induced and G protein ?? (G??)-mediated PI3K? regulation. Here we investigated the molecular mechanism for receptor-dependent p87/p110? activation. By analyzing GFP-tagged proteins expressed in HEK293 cells, PI3K?-complemented bone marrow–derived mast cells (BMMCs) from p110?-/- mice, and purified recombinant proteins reconstituted to lipid vesicles, we elucidated a novel pathway of p87-dependent, G protein–coupled receptor (GPCR)-induced PI3K? activation. Although p101 strongly interacted with G??, thereby mediating PI3K? membrane recruitment and stimulation, p87 exhibited only a weak interaction, resulting in modest kinase activation and lack of membrane recruitment. Surprisingly, Ras-GTP substituted the missing G??-dependent membrane recruitment of p87/p110? by direct interaction with p110?, suggesting the indispensability of Ras for activation of p87/p110?. Consequently, interference with Ras signaling indeed selectively blocked p87/p110?, but not p101/p110?, kinase activity in HEK293 and BMMC cells, revealing an important crosstalk between monomeric and trimeric G proteins for p87/p110? activation. Our data display distinct signaling requirements of p87 and p101, conferring signaling specificity to PI3K? that could open up new possibilities for therapeutic intervention. PMID:19906996

  17. Reduced Phosphoinositide 3-Kinase (p110?) Activation Increases the Susceptibility to Atrial Fibrillation

    PubMed Central

    Pretorius, Lynette; Du, Xiao-Jun; Woodcock, Elizabeth A.; Kiriazis, Helen; Lin, Ruby C.Y.; Marasco, Silvana; Medcalf, Robert L.; Ming, Ziqiu; Head, Geoffrey A.; Tan, Joon Win; Cemerlang, Nelly; Sadoshima, Junichi; Shioi, Tetsuo; Izumo, Seigo; Lukoshkova, Elena V.; Dart, Anthony M.; Jennings, Garry L.; McMullen, Julie R.

    2009-01-01

    Atrial fibrillation (AF) is the most common sustained arrhythmia presenting at cardiology departments. A limited understanding of the molecular mechanisms responsible for the development of AF has hindered treatment strategies. The purpose of this study was to assess whether reduced activation of phosphoinositide 3-kinase (PI3K, p110?) makes the compromised heart susceptible to AF. Risk factors for AF, including aging, obesity, and diabetes, have been associated with insulin resistance that leads to depressed/defective PI3K signaling. However, to date, there has been no link between PI3K(p110?) and AF. To address this question, we crossed a cardiac-specific transgenic mouse model of dilated cardiomyopathy (DCM) with a cardiac-specific transgenic mouse expressing a dominant negative mutant of PI3K (dnPI3K; reduces PI3K activity). Adult (?4.5 months) double-transgenic (dnPI3K-DCM), single-transgenic (DCM-Tg, dnPI3K-Tg), and nontransgenic mice were subjected to morphological, functional/ECG, microarray, and biochemical analyses. dnPI3K-DCM mice developed AF and had depressed cardiac function as well as greater atrial enlargement and fibrosis than DCM-Tg mice. AF was not detected in other groups. Aged DCM-Tg mice (?15 months) with a similar phenotype to dnPI3K-DCM mice (4.5 months) did not develop AF, suggesting loss of PI3K activity directly contributed to the AF phenotype. Furthermore, increasing PI3K activity reduced atrial fibrosis and improved cardiac conduction in DCM-Tg mice. Finally, in atrial appendages from patients with AF, PI3K activation was lower compared with tissue from patients in sinus rhythm. These results suggest a link between PI3K(p110?) and AF. PMID:19679877

  18. IN VITRO ALUMINUM INHIBITION OF BRAIN PHOSPHOINOSITIDE METABOLISM:COMPARISON OF NEONATAL AND ADULTS RATS

    EPA Science Inventory

    Recent evidence indicates that the neurotoxic metal aluminum interferes with the phosphoinositide second messenger system in adult rats both in vitro and in vivo. e have examined the age-related effects of aluminum chloride (AlCl3) on receptor-stimulated inositol phosphate (IP) a...

  19. Oxidized low density lipoproteins stimulate phosphoinositide turnover in cultured vascular smooth muscle cells.

    PubMed

    Resink, T J; Tkachuk, V A; Bernhardt, J; Bühler, F R

    1992-03-01

    Atherogenesis is associated with alterations in the properties of different cell types, including monocytes/macrophages (foam cell formation), platelets (increased aggregation), endothelial cells (injury), and smooth muscle cells (SMCs) (lipid accumulation or foam cell formation). Oxidized low density lipoproteins (ox-LDL) play a key role in this vascular pathology. This study investigated the ability of ox-LDL to elicit chemical signaling events in cultured human vascular smooth muscle cells (VSMCs). Ox-LDL was found to stimulate phospholipase C-mediated phosphoinositide turnover in human VSMCs. This response occurred rapidly (within 1 minute) and at low concentrations of ox-LDL (half-maximal effective concentration, approximately 5 micrograms/ml). Ox-LDL-stimulated inositol phosphate accumulation in human VSMCs was inhibited by pretreatment of cells with phorbol 12-myristate 13-acetate and with compounds that elevate cyclic AMP or cyclic GMP. Ca2+ antagonists also blocked the effects of ox-LDL on phosphoinositide turnover. Inhibitors of receptor-endocytotic processes (including receptor clustering, cross-linking, and cytoskeleton-dependent internalization) effectively prevented ox-LDL-induced inositol phosphate generation. The data suggest that ox-LDL promotes phospholipase C-mediated phosphoinositide turnover in a manner analogous to that for other Ca(2+)-mobilizing hormones. The results also support an association between phosphoinositide turnover and receptor-mediated endocytosis. Prevention of the direct effects of ox-LDL on SMCs could prove an interesting therapeutic avenue for the prevention of atherosclerosis. PMID:1312338

  20. Coordinated Expression of Phosphoinositide Metabolic Genes during Development and Aging of Human Dorsolateral Prefrontal Cortex

    PubMed Central

    Rapoport, Stanley I.; Primiani, Christopher T.; Chen, Chuck T.; Ahn, Kwangmi; Ryan, Veronica H.

    2015-01-01

    Background Phosphoinositides, lipid-signaling molecules, participate in diverse brain processes within a wide metabolic cascade. Hypothesis Gene transcriptional networks coordinately regulate the phosphoinositide cascade during human brain Development and Aging. Methods We used the public BrainCloud database for human dorsolateral prefrontal cortex to examine age-related expression levels of 49 phosphoinositide metabolic genes during Development (0 to 20+ years) and Aging (21+ years). Results We identified three groups of partially overlapping genes in each of the two intervals, with similar intergroup correlations despite marked phenotypic differences between Aging and Development. In each interval, ITPKB, PLCD1, PIK3R3, ISYNA1, IMPA2, INPPL1, PI4KB, and AKT1 are in Group 1, PIK3CB, PTEN, PIK3CA, and IMPA1 in Group 2, and SACM1L, PI3KR4, INPP5A, SYNJ1, and PLCB1 in Group 3. Ten of the genes change expression nonlinearly during Development, suggesting involvement in rapidly changing neuronal, glial and myelination events. Correlated transcription for some gene pairs likely is facilitated by colocalization on the same chromosome band. Conclusions Stable coordinated gene transcriptional networks regulate brain phosphoinositide metabolic pathways during human Development and Aging. PMID:26168237

  1. Oncogenic mutations mimic and enhance dynamic events in the natural activation of phosphoinositide

    E-print Network

    Williams, Roger L.

    Oncogenic mutations mimic and enhance dynamic events in the natural activation of phosphoinositide110/p85 and a spectrum of oncogenic mutants using hydrogen/deuterium exchange mass spectrometry (HDX to an activated form on membranes entails four distinct events. Analysis of oncogenic mutations shows that all up

  2. Phosphatidylinositol 3-kinase-dependent, MEK-independent proliferation in response to CaR activation

    SciTech Connect

    Bilderback, Tim R.; Lee, Fred; Auersperg, Nelly; Rodland, Karin D.

    2002-07-02

    Although ovarian surface epithelial (OSE) cells are responsible for the majority of ovarian tumors, we know relatively little about the pathway(s) that are responsible for regulating their proliferation. We found that phosphatidylinositol 3-kinase (PI3K) is activated in OSE cells in response to elevated extracellular calcium, and the PI3K inhibitors wortmannin and LY29004 inhibited ERK activation by approximately 75%, similar to effects of the MEK2 inhibitor PD98059. However, in assays of proliferation we found that PD98059 inhibited proliferation by approximately 50%, while wortmannin inhibited greater than 90% of the proliferative response to elevated calcium. Expression of a dominant negative PI3K totally inhibited ERK activation in response to calcium. These results demonstrate that ERK activation cannot account for the full proliferative effect of elevated calcium in OSE cells, and suggest the presence of an ERK independent, PI3K dependant component in the proliferative response.

  3. IGF-1 Modulates N and L Calcium Channels in a PI 3-Kinase-Dependent Manner

    Microsoft Academic Search

    Lesley A. C Blair; John Marshall

    1997-01-01

    Receptor tyrosine kinases (RTKs) have long been associated with proliferation in non-neural cells, although they are also expressed in postmitotic neurons. We demonstrate that insulin-like growth factor-1 (IGF-1) induces within seconds a large, tyrosine- kinase-dependent increase in calcium channel currents in cerebellar granule neurons. Separation of channel subtypes reveals that, while P, Q, and R channels are unaffected, N and

  4. Maintenance of hormone-sensitive phosphoinositide pools in the plasma membrane requires phosphatidylinositol 4-kinase IIIalpha.

    PubMed

    Balla, Andras; Kim, Yeun Ju; Varnai, Peter; Szentpetery, Zsofia; Knight, Zachary; Shokat, Kevan M; Balla, Tamas

    2008-02-01

    Type III phosphatidylinositol (PtdIns) 4-kinases (PI4Ks) have been previously shown to support plasma membrane phosphoinositide synthesis during phospholipase C activation and Ca(2+) signaling. Here, we use biochemical and imaging tools to monitor phosphoinositide changes in the plasma membrane in combination with pharmacological and genetic approaches to determine which of the type III PI4Ks (alpha or beta) is responsible for supplying phosphoinositides during agonist-induced Ca(2+) signaling. Using inhibitors that discriminate between the alpha- and beta-isoforms of type III PI4Ks, PI4KIIIalpha was found indispensable for the production of phosphatidylinositol 4-phosphate (PtdIns4P), phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P(2)], and Ca(2+) signaling in angiotensin II (AngII)-stimulated cells. Down-regulation of either the type II or type III PI4K enzymes by small interfering RNA (siRNA) had small but significant effects on basal PtdIns4P and PtdIns(4,5)P(2) levels in (32)P-labeled cells, but only PI4KIIIalpha down-regulation caused a slight impairment of PtdIns4P and PtdIns(4,5)P(2) resynthesis in AngII-stimulated cells. None of the PI4K siRNA treatments had a measurable effect on AngII-induced Ca(2+) signaling. These results indicate that a small fraction of the cellular PI4K activity is sufficient to maintain plasma membrane phosphoinositide pools, and they demonstrate the value of the pharmacological approach in revealing the pivotal role of PI4KIIIalpha enzyme in maintaining plasma membrane phosphoinositides. PMID:18077555

  5. Effects of thyroxine and 1-methyl, 2-mercaptoimidazol on phosphoinositides synthesis in rat liver

    PubMed Central

    Babenko, Nataliya A; Krasilnikova, Oksana A

    2004-01-01

    Background Phosphoinositides mediate one of the intracellular signal transduction pathways and produce a class of second messengers that are involved in the action of hormones and neurotransmitters on target cells. Thyroid hormones are well known regulators of lipid metabolism and modulators of signal transduction in cells. However, little is known about phosphoinositides cycle regulation by thyroid hormones. The present paper deals with phosphoinositides synthesis de novo and acylation in liver at different thyroid status of rats. Results The experiments were performed in either the rat liver or hepatocytes of 90- and 720-day-old rats. Myo-[3H]inositol, [14C]CH3COONa, [14C]oleic and [3H]arachidonic acids were used to investigate the phosphatidylinositol (PtdIns), phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate (PtdInsP2) synthesis. 1-methyl, 2-mercaptoimidazol-induced hypothyroidism was associated with the decrease of myo-[3H]inositol and [3H]arachidonic acids incorporation into liver phosphoinositides and total phospholipids, respectively. The thyroxine (L-T4) injection to hypothyroid animals increased the hormones contents in blood serum and PtdInsP2 synthesis de novo as well as [3H]arachidonic acids incorporation into the PtdIns and PtdInsP2. Under the hormone action, the [14C]oleic acid incorporation into PtdIns reduced in the liver of hypothyroid animals. A single injection of L-T4 to the euthyroid [14C]CH3COONa-pre-treated animals or addition of the hormone to a culture medium of hepatocytes was accompanied by the rapid prominent increase in the levels of the newly synthesized PtdIns and PtdInsP2 and in the mass of phosphatidic acid in the liver or the cells. Conclusions The data obtained have demonstrated that thyroid hormones are of vital importance in the regulation of arachidonate-containing phosphoinositides metabolism in the liver. The drug-induced malfunction of thyroid gland noticeably changed the phosphoinositides synthesis de novo. The L-T4 injection to the animals was followed by the time-dependent increase of polyphosphoinositide synthesis in the liver. The both long-term and short-term hormone effects on the newly synthesized PtdInsP2 have been determined. PMID:15588322

  6. Effect of ACTH on rate of /sup 32/P-orthophosphate uptake into synaptosomal phosphoinositides of the ischemic rat brain

    SciTech Connect

    Pavlinova, L.I.; Tyul'kova, E.I.; Gasteva, S.V.

    1987-06-01

    The aim of this investigation was to study phosphoinositide metabolism in rat brain synaptosomes under normal conditions, under the influence of ACTH, and during depolarization of the synaptosomes, and also to compare it with the metabolic response of synaptosomes obtained from the ischemic rat brain. The rate of incorporation of orthophosphate, labelled with phosphorus 32, was expressed as the ratio of specific radioactivity of phosphorus in the phosphoinositide to the protein content.

  7. Profiling and relative quantitation of phosphoinositides by multiple precursor ion scanning based on phosphate methylation and isotopic labeling.

    PubMed

    Cai, Tanxi; Shu, Qingbo; Hou, JunJie; Liu, Peibin; Niu, Lili; Guo, Xiaojing; Liu, Charles C; Yang, Fuquan

    2015-01-01

    Phosphoinositides, the phosphorylated derivatives of phosphatidylinositol (PtdIns), are key regulators of many fundamental biological processes, including cell growth, proliferation, and motility. Here, we present a novel method for rapid, sensitive, and simultaneous profiling of phosphatidylinositol trisphosphate (PtdInsP3), phosphatidylinositol bisphosphate (PtdInsP2), and phosphatidylinositol phosphate (PtdInsP) of different fatty acid compositions. This method is based on a technique called "charged diacylglycerol fragment ion-specific multiple precursor ion scanning" (DAG(+)-specific MPIS), coupled with prior phosphate methylation. Using DAG(+)-specific MPIS, we were able to identify 32 PtdIns, 28 PtdInsP, 30 PtdInsP2, and 3 PtdInsP3 molecular species from bovine brain extracts or prostatic cancer cell lines in an efficient and time-saving manner. Our analysis revealed a large range of fatty acyl compositions in phosphoinositides not obtained previously from mammalian samples. We also developed a method that involves isotopic labeling of endogenous phosphoinositides with deuterated diazomethane (CD2N2) for quantitation of phosphoinositides. CD2N2 was generated in situ through acid-catalyzed H/D exchange and methanolysis of trimethylsilyl diazomethane (TMS-diazomethane). Phosphoinositides, extracted from a PC3 prostatic cancer cell line, were labeled either with CH2N2 or CD2N2 and mixed in known proportions for DAG(+)-specific MPIS-based mass spectrometry (MS) analysis. The results indicate that isotopic labeling is capable of providing accurate quantitation of PtdInsP3, PtdInsP2, and PtdInsP with adequate linearity as well as high reproducibility with an average coefficient variation of 18.9%. More importantly, this new methods excluded the need for multiple phosphoinositide internal standards. DAG(+)-specific MPIS and isotopic labeling based MS analysis of phosphoinositides offers unique advantages over existing approaches and presents a powerful tool for research of phosphoinositide metabolism. PMID:25495789

  8. GABAA receptor-associated phosphoinositide 3-kinase is required for insulin-induced recruitment of postsynaptic GABAA receptors.

    PubMed

    Vetiska, S M; Ahmadian, G; Ju, W; Liu, L; Wymann, M P; Wang, Y T

    2007-01-01

    Type A gamma-aminobutyric acid (GABAA) receptors mediate most of the fast inhibitory synaptic transmission within the vertebrate brain. The regulation of this inhibition is vital in modulating neural activity. One regulator of GABAA receptor function is insulin, which can serve to enhance GABAA receptor-mediated miniature inhibitory postsynaptic currents, via an increase in the number of receptors at the plasma membrane. We set out to investigate the molecular mechanisms involved in the insulin-induced potentiation of GABAA receptor-mediated responses, by examining the role of phosphoinositide 3-kinase (PI3-K), a key mediator of the insulin response within the brain. We found that PI3-K associates with the GABAA receptor, and this interaction is increased following insulin treatment. Additionally, the beta2 subunit of the GABAA receptor appears to mediate the insulin-stimulated association with the N-terminal SH2 domain of the p85 subunit of PI3-K. Our results imply a mechanism whereby insulin can regulate changes in synaptic transmission through its downstream actions on the GABAA receptor. PMID:16890252

  9. [Effects of Salvia miltiorrhizae compositae on phosphoinositides metabolism in acute myocardial ischemia].

    PubMed

    Tao, Y Y

    1993-06-01

    The effects of Salvia miltiorrhizae compositae (SMC) on phosphoinositides metabolism of acute myocardial ischemia in rats were studied. The results showed that the function of the phosphoinositide signaling system was enhanced 10 min. after acute myocardial ischemia. Both of the accumulations of phosphatidylinositol-4,5-biphosphate (PIP2) and inositol-1,4,5-triphosphate (IP3) were higher than those of the non-ischemic control group (P < 0.01, n = 7). The rise of the levels of PIP2 and IP3 caused by acute myocardial ischemia was remarkably inhibited by the SMC (2g/kg body wt.), and the levels of PIP2 and IP3 were lower than those of the ischemic group (P < 0.01, n = 7). PMID:8257842

  10. LIVE CELL IMAGING OF PHOSPHOINOSITIDES WITH EXPRESSED INOSITIDE-BINDING PROTEIN DOMAINS

    PubMed Central

    Várnai, Péter; Balla, Tamas

    2008-01-01

    Summary Inositol lipids and calcium signaling has been inseparable twins during the 1980s when the molecular details of phospholipase C-mediated generation of inositol 1,4,5-trisphosphate (InsP3) and its Ca2+ mobilizing action were discovered. Since then, both the Ca2+- and inositol lipid signaling fields have hugely expanded and the tools allowing dissection of the finest details of their molecular organization also followed closely. Although phosphoinositides regulate many cell functions unrelated to Ca2+ signaling there are still many open questions even in the Ca2+ field that would benefit from single cell monitoring of PtdIns(4,5)P2 or InsP3 changes during agonist stimulation. This chapter is designed to provide practical guidance as well as some theoretical background on measurements of phosphoinositides in live cells using protein domain-GFP chimeras that could be also useful for people working on calcium signaling. PMID:18930153

  11. Abscisic Acid-Induced Phosphoinositide Turnover in Guard Cell Protoplasts of Vicia faba.

    PubMed Central

    Lee, Y.; Choi, Y. B.; Suh, S.; Lee, J.; Assmann, S. M.; Joe, C. O.; Kelleher, J. F.; Crain, R. C.

    1996-01-01

    Guard cell protoplasts of Vicia faba treated with 10 [mu]M (+)abscisic acid (ABA) in the light exhibited a 20% decrease in diameter within 1.5 h, from 24.1 to 19.6 [mu]m. Within 10 s of administration of ABA, a 90% increase in levels of inositol 1,4,5-trisphosphate was observed, provided that cells were treated with Li+, an inhibitor of inositol phosphatase activity, prior to incubation. Concomitantly, levels of 32P-labeled phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol 4-phosphate decreased 20% compared to levels in control cells; levels of label in the membrane lipids phosphatidylcholine, phosphatidylethanolamine, and phosphatidylglycerol did not change significantly in response to ABA treatment. These results show that phosphoinositide turnover is activated in response to ABA in guard cells. We conclude that phosphoinositide signaling is likely to be a step in the biochemical cascade that couples ABA to guard cell shrinking and stomatal closure. PMID:12226236

  12. Atg18 phosphoregulation controls organellar dynamics by modulating its phosphoinositide-binding activity

    PubMed Central

    Tamura, Naoki; Oku, Masahide; Ito, Moemi; Noda, Nobuo N.; Inagaki, Fuyuhiko

    2013-01-01

    The PROPPIN family member Atg18 is a phosphoinositide-binding protein that is composed of a seven ?-propeller motif and is part of the conserved autophagy machinery. Here, we report that the Atg18 phosphorylation in the loops in the propellar structure of blade 6 and blade 7 decreases its binding affinity to phosphatidylinositol 3,5-bisphosphate in the yeast Pichia pastoris. Dephosphorylation of Atg18 was necessary for its association with the vacuolar membrane and caused septation of the vacuole. Upon or after dissociation from the vacuolar membrane, Atg18 was rephosphorylated, and the vacuoles fused and formed a single rounded structure. Vacuolar dynamics were regulated according to osmotic changes, oxidative stresses, and nutrient conditions inducing micropexophagy via modulation of Atg18 phosphorylation. This study reveals how the phosphoinositide-binding activity of the PROPPIN family protein Atg18 is regulated at the membrane association domain and highlights the importance of such phosphoregulation in coordinated intracellular reorganization. PMID:23940117

  13. Phosphoinositide signalling links O-GlcNAc transferase to insulin resistance

    Microsoft Academic Search

    Xiaoyong Yang; Pat P. Ongusaha; Philip D. Miles; Joyce C. Havstad; Fengxue Zhang; W. Venus So; Jeffrey E. Kudlow; Robert H. Michell; Jerrold M. Olefsky; Seth J. Field; Ronald M. Evans

    2008-01-01

    Glucose flux through the hexosamine biosynthetic pathway leads to the post-translational modification of cytoplasmic and nuclear proteins by O-linked beta-N-acetylglucosamine (O-GlcNAc). This tandem system serves as a nutrient sensor to couple systemic metabolic status to cellular regulation of signal transduction, transcription, and protein degradation. Here we show that O-GlcNAc transferase (OGT) harbours a previously unrecognized type of phosphoinositide-binding domain. After

  14. Protein kinase activity of phosphoinositide 3-kinase regulates ?-adrenergic receptor endocytosis

    Microsoft Academic Search

    Sathyamangla V. Naga Prasad; Arundathi Jayatilleke; Aasakiran Madamanchi; Howard A. Rockman

    2005-01-01

    Phosphoinositide 3-kinase (PI(3)K) is a unique enzyme characterized by both lipid and protein kinase activities. Here, we demonstrate a requirement for the protein kinase activity of PI(3)K in agonist-dependent ?-adrenergic receptor (?AR) internalization. Using PI(3)K mutants with either protein or lipid phosphorylation activity, we identify the cytoskeletal protein non-muscle tropomyosin as a substrate of PI(3)K, which is phosphorylated in a

  15. Thrombin-induced phosphoinositide hydrolysis in platelets. Receptor occupancy and desensitization.

    PubMed Central

    Huang, E M; Detwiler, T C

    1987-01-01

    The relationship between occupancy of thrombin receptors on platelets and enhanced phosphoinositide hydrolysis was analysed by examination of the dose-response relationship, the effects of thrombin inhibitors and the contribution of secondary effects. Washed human platelets were labelled with [3H]inositol, and agonist-induced accumulation of labelled inositol phosphates was measured. The dose-response curves and the time courses for alpha-thrombin- or gamma-thrombin-induced accumulation of inositol phosphates were similar to those for dense-granule secretion. Addition of the thrombin inhibitor hirudin to thrombin-activated platelets revealed that the continuous presence of active thrombin was required to maintain the accumulation of labelled inositol phosphates; the total production of inositol phosphates increased with longer periods of exposure to thrombin, reaching a maximum between 5 and 10 min. After activation with thrombin, the ability of a second, greater, addition of thrombin to induce additional phosphoinositide hydrolysis decreased with time; it was absent within 10 min after the first addition. The failure to sustain accumulation of labelled inositol phosphates or to respond to a second addition of thrombin beyond 10 min was not due to depletion of the pool of labelled precursors, because the platelets retained their ability to respond to collagen. Addition of ADP-consuming enzymes decreased sensitivity to thrombin, but inhibition of cyclo-oxygenase with indomethacin did not impair the thrombin-induced hydrolysis of phosphoinositides. It was concluded that thrombin-induced hydrolysis of phosphoinositides has characteristics consistent with mediation by a receptor that is similar to that that triggers dense-granule secretion, requires continuous presence of active thrombin to be maintained, is mediated by a receptor that displays thrombin-induced desensitization, and is only partially enhanced by secondary agents. PMID:3036080

  16. Rapid Turnover Rate of Phosphoinositides at the Front of Migrating MDCK Cells

    Microsoft Academic Search

    Teruko Nishioka; Kazuhiro Aoki; Kazuhiro Hikake; Hisayoshi Yoshizaki; Etsuko Kiyokawa; Michiyuki Matsuda

    2008-01-01

    Phosphoinositides (PtdInss) play key roles in cell polarization and motility. With a series of biosensors based on Forster resonance energy transfer, we examined the distribution and metabolism of PtdInss and diacylglycerol (DAG) in stochastically migrating Madin-Darby canine kidney (MDCK) cells. The concentrations of phosphatidylinositol (4,5)- bisphosphate, phosphatidylinositol (3,4,5)-trisphosphate (PIP3), phosphatidylinositol (3,4)-bisphosphate, and DAG were higher at the plasma membrane in

  17. Central Role for G Protein-Coupled Phosphoinositide 3Kinase gamma in Inflammation

    Microsoft Academic Search

    Emilio Hirsch; Vladimir L. Katanaev; Cecilia Garlanda; Ornella Azzolino; Luciano Pirola; Lorenzo Silengo; Silvano Sozzani; Alberto Mantovani; Fiorella Altruda; Matthias P. Wymann

    2000-01-01

    Phosphoinositide 3-kinase (PI3K) activity is crucial for leukocyte function, but the roles of the four receptor-activated isoforms are unclear. Mice lacking heterotrimeric guanine nucleotide-binding protein (G protein)-coupled PI3Kgamma were viable and had fully differentiated neutrophils and macrophages. Chemoattractant-stimulated PI3Kgamma-\\/- neutrophils did not produce phosphatidylinositol 3,4,5-trisphosphate, did not activate protein kinase B, and displayed impaired respiratory burst and motility. Peritoneal PI3Kgamma-null

  18. Structural Determinants of Phosphoinositide 3Kinase Inhibition by Wortmannin, LY294002, Quercetin, Myricetin, and Staurosporine

    Microsoft Academic Search

    Edward H. Walker; Michael E. Pacold; Olga Perisic; Len Stephens; Philip T. Hawkins; Matthias P. Wymann; Roger L. Williams

    2000-01-01

    The specific phosphoinositide 3-kinase (PI3K) inhibitors wortmannin and LY294002 have been invaluable tools for elucidating the roles of these enzymes in signal transduction pathways. The X-ray crystallographic structures of PI3K? bound to these lipid kinase inhibitors and to the broad-spectrum protein kinase inhibitors quercetin, myricetin, and staurosporine reveal how these compounds fit into the ATP binding pocket. With a nanomolar

  19. Phosphoinositide 3Kinase ? Is an Essential Amplifier of Mast Cell Function

    Microsoft Academic Search

    Muriel Laffargue; Ronan Calvez; Peter Finan; Alexandre Trifilieff; Maryse Barbier; Fiorella Altruda; Emilio Hirsch; Matthias P Wymann

    2002-01-01

    Mast cells are key regulators in allergy and inflammation, and release histamine upon clustering of their IgE receptors. Here we demonstrate that murine mast cell responses are exacerbated in vitro and in vivo by autocrine signals through G protein-coupled receptors (GPCRs) and require functional phosphoinositide 3-kinase ? (PI3K?). Adenosine, acting through the A3 adenosine receptor (A3AR) as well as other

  20. Ankyrin-G palmitoylation and ?II-spectrin binding to phosphoinositide lipids drive lateral membrane assembly

    PubMed Central

    He, Meng; Abdi, Khadar M.

    2014-01-01

    Ankyrin-G and ?II-spectrin colocalize at sites of cell–cell contact in columnar epithelial cells and promote lateral membrane assembly. This study identifies two critical inputs from lipids that together provide a rationale for how ankyrin-G and ?II-spectrin selectively localize to Madin-Darby canine kidney (MDCK) cell lateral membranes. We identify aspartate-histidine-histidine-cysteine 5/8 (DHHC5/8) as ankyrin-G palmitoyltransferases required for ankyrin-G lateral membrane localization and for assembly of lateral membranes. We also find that ?II-spectrin functions as a coincidence detector that requires recognition of both ankyrin-G and phosphoinositide lipids for its lateral membrane localization. DHHC5/8 and ?II-spectrin colocalize with ankyrin-G in micrometer-scale subdomains within the lateral membrane that are likely sites for palmitoylation of ankyrin-G. Loss of either DHHC5/8 or ankyrin-G–?II-spectrin interaction or ?II-spectrin–phosphoinositide recognition through its pleckstrin homology domain all result in failure to build the lateral membrane. In summary, we identify a functional network connecting palmitoyltransferases DHHC5/8 with ankyrin-G, ankyrin-G with ?II-spectrin, and ?II-spectrin with phosphoinositides that is required for the columnar morphology of MDCK epithelial cells. PMID:25049274

  1. Phosphoinositide Kinase-3 Status Associated With Presence or Absence of Human Papillomavirus in Head and Neck Squamous Cell Carcinomas

    SciTech Connect

    Yarbrough, Wendell G. [Department of Otolaryngology, Vanderbilt University, Nashville, TN (United States); Department of Cancer Biology, Vanderbilt University, Nashville, TN (United States); Ingram Cancer Center, Vanderbilt University, Nashville, TN (United States)], E-mail: Wendell.yarbrough@vanderbilt.edu; Whigham, Amy; Brown, Brandee [Department of Otolaryngology, Vanderbilt University, Nashville, TN (United States); Roach, Michael [Department of Cancer Biology, Vanderbilt University, Nashville, TN (United States); Slebos, Robbert [Department of Otolaryngology, Vanderbilt University, Nashville, TN (United States); Department of Cancer Biology, Vanderbilt University, Nashville, TN (United States)

    2007-10-01

    Purpose: To investigate phosphoinositide kinase-3 (PI3K) activation in relation to human papillomavirus (HPV) status in head and neck squamous cell carcinoma (HNSCC). Methods and Materials: Gene expression microarray data were analyzed to determine differentially expressed genes between HPV(+) and HPV(-) HNSCC. PIK3CA gene expression was confirmed by quantitative reverse transcriptase-polymerase chain reaction in seven HPV(+) and seven HPV(-) primary HNSCCs. PIK3CA mutation status in three HPV(+) and nine HPV(-) cell lines was determined by polymerase chain reaction amplification of hot spot exons (1, 9, 20) followed by direct sequencing. Results: PIK3CA was overexpressed in HPV(+)-associated HNSCC compared with the expression in HPV(-) HNSCC. Activation of PIK3CA by mutation was found in 1 of the 12 tested HNSCC cell lines. Conclusion: Activation of PI3K by mutation of PIK3CA is rare in HNSCC cell lines and was not found in three HPV(+) cell lines. One mechanism by which HPV-associated HNSCC might activate PI3K is increased expression of PIK3CA.

  2. Mammalian phosphoinositide-specic phospholipase C Roger L. Williams *

    E-print Network

    Williams, Roger L.

    of regulatory mechanisms control- ling the mammalian PI-PLCs. Ten mammalian PLC isozymes have been reported: PLC messengers, inositol-1,4,5-trisphosphate (Ins(1,4,5)P3) and diacylglycerol. These two products of PLC, the mammalian phospholipases are dis- tinct from bacterial PI-PLC that cannot hydrolyse phosphorylated

  3. Assessing the subcellular distribution of oncogenic phosphoinositide 3-kinase using microinjection into live cells

    PubMed Central

    Layton, Meredith J.; Rynkiewicz, Natalie K.; Ivetac, Ivan; Horan, Kristy A.; Mitchell, Christina A.; Phillips, Wayne A.

    2014-01-01

    Oncogenic mutations in PIK3CA lead to an increase in intrinsic phosphoinositide kinase activity, but it is thought that increased access of PI3K? (phosphoinositide 3-kinase ?) to its PM (plasma membrane) localized substrate is also required for increased levels of downstream PIP3/Akt [phosphoinositide-3,4,5-trisphosphate/also called PKB (protein kinase B)] signalling. We have studied the subcellular localization of wild-type and the two most common oncogenic mutants of PI3K? in cells maintained in growth media, and starved or stimulated cells using a novel method in which PI3K? is pre-formed as a 1:1 p110?:p85? complex in vitro then introduced into live cells by microinjection. Oncogenic E545K and H1047R mutants did not constitutively interact with membrane lipids in vitro or in cells maintained in 10% (v/v) FBS. Following stimulation of RTKs (receptor tyrosine kinases), microinjected PI3K? was recruited to the PM, but oncogenic forms of PI3K? were not recruited to the PM to a greater extent and did not reside at the PM longer than the wild-type PI3K?. Instead, the E545K mutant specifically bound activated Cdc42 in vitro and microinjection of E545K was associated with the formation of cellular protrusions, providing some preliminary evidence that changes in protein–protein interactions may play a role in the oncogenicity of the E545K mutant in addition to the well-known changes in lipid kinase activity. PMID:24597785

  4. Membrane targeting by APPL1 and APPL2: dynamic scaffolds that oligomerize and bind phosphoinositides.

    PubMed

    Chial, Heidi J; Wu, Ruping; Ustach, Carolyn V; McPhail, Linda C; Mobley, William C; Chen, Yong Q

    2008-02-01

    Human adaptor protein, phosphotyrosine interaction, PH domain and leucine zipper containing 1 (APPL1) and adaptor protein, phosphotyrosine interaction, PH domain and leucine zipper containing 2 (APPL2) are homologous effectors of the small guanosine triphosphatase RAB5 that interact with a diverse set of receptors and signaling proteins and are proposed to function in endosome-mediated signaling. Herein, we investigated the membrane-targeting properties of the APPL1 and APPL2 Bin/Amphiphysin/Rvs (BAR), pleckstrin homology (PH) and phosphotyrosine binding (PTB) domains. Coimmunoprecipitation and yeast two-hybrid studies demonstrated that full-length APPL proteins formed homooligomers and heterooligomers and that the APPL minimal BAR domains were necessary and sufficient for mediating APPL-APPL interactions. When fused to a fluorescent protein and overexpressed, all three domains (minimal BAR, PH and PTB) were targeted to cell membranes. Furthermore, full-length APPL proteins bound to phosphoinositides, and the APPL isolated PH or PTB domains were sufficient for in vitro phosphoinositide binding. Live cell imaging showed that full-length APPL-yellow fluorescent protein (YFP) fusion proteins associated with cytosolic membrane structures that underwent movement, fusion and fission events. Overexpression of full-length APPL-YFP fusion proteins was sufficient to recruit endogenous RAB5 to enlarged APPL-associated membrane structures, although APPL1 was not necessary for RAB5 membrane targeting. Taken together, our findings suggest a role for APPL proteins as dynamic scaffolds that modulate RAB5-associated signaling endosomal membranes by their ability to undergo domain-mediated oligomerization, membrane targeting and phosphoinositide binding. PMID:18034774

  5. Epigallocatechin gallate (EGCG), a major component of green tea, is a dual phosphoinositide-3-kinase/mTOR inhibitor

    SciTech Connect

    Van Aller, Glenn S., E-mail: glenn.s.van.aller@gsk.com [Department of Cancer Research, GlaxoSmithKline, Collegeville, PA 19426 (United States); Carson, Jeff D. [Department of Cancer Research, GlaxoSmithKline, Collegeville, PA 19426 (United States)] [Department of Cancer Research, GlaxoSmithKline, Collegeville, PA 19426 (United States); Tang, Wei; Peng, Hao; Zhao, Lin [Discovery Biology, BioDuro, No. 29 Life Science Park Road, Changping, Beijing (China)] [Discovery Biology, BioDuro, No. 29 Life Science Park Road, Changping, Beijing (China); Copeland, Robert A.; Tummino, Peter J. [Department of Cancer Research, GlaxoSmithKline, Collegeville, PA 19426 (United States)] [Department of Cancer Research, GlaxoSmithKline, Collegeville, PA 19426 (United States); Luo, Lusong [Discovery Biology, BioDuro, No. 29 Life Science Park Road, Changping, Beijing (China)] [Discovery Biology, BioDuro, No. 29 Life Science Park Road, Changping, Beijing (China)

    2011-03-11

    Research highlights: {yields} Epigallocatechin-3-gallate (EGCG) is an ATP-competitive inhibitor of PI3K and mTOR with Ki values around 300 nM. {yields} EGCG inhibits cell proliferation and AKT phosphorylation at Ser473 in MDA-MB-231and A549 cells. {yields} Molecular docking studies show that EGCG binds well to the PI3K kinase domain active site. {yields} These results suggest another important molecular mechanism for the anticancer activities of EGCG. -- Abstract: The PI3K signaling pathway is activated in a broad spectrum of human cancers, either directly by genetic mutation or indirectly via activation of receptor tyrosine kinases or inactivation of the PTEN tumor suppressor. The key nodes of this pathway have emerged as important therapeutic targets for the treatment of cancer. In this study, we show that (-)-epigallocatechin-3-gallate (EGCG), a major component of green tea, is an ATP-competitive inhibitor of both phosphoinositide-3-kinase (PI3K) and mammalian target of rapamycin (mTOR) with K{sub i} values of 380 and 320 nM respectively. The potency of EGCG against PI3K and mTOR is within physiologically relevant concentrations. In addition, EGCG inhibits cell proliferation and AKT phosphorylation at Ser473 in MDA-MB-231 and A549 cells. Molecular docking studies show that EGCG binds well to the PI3K kinase domain active site, agreeing with the finding that EGCG competes for ATP binding. Our results suggest another important molecular mechanism for the anticancer activities of EGCG.

  6. Threonine 34 phosphorylation by phosphoinositide-dependent protein kinase 1 facilitates dissociation of Akt from the plasma membrane.

    PubMed

    Huang, Bill X; Lee, Rachel; Akbar, Mohammed; Kim, Hee-Yong

    2015-07-01

    Akt is a key mediator of cell proliferation, survival and metabolism. After translocation to the membrane and phosphorylation at T308 and S473, the activated Akt dissociates from the plasma membrane to cytoplasm, which is an important step to phosphorylate its downstream targets. In addition to its central role in regulating the kinase activity, phosphorylation of T308 in the kinase loop has been reported to be necessary for this dissociation process. However, it is not clear whether the membrane detachment requires further mechanisms. In the present report, we demonstrate that membrane dissociation of Akt requires phosphoinositide-dependent protein kinase 1 (PDK1) which directly phosphorylates not only T308 but also T34 in the pleckstrin homology (PH) domain. Like T308, T34 was phosphorylated in a phosphatidylinositol 3,4,5-trisphosphate- and phosphatidylserine-dependent manner. Phosphorylation of T34 also occurred in cells following growth factor stimulation, concurrently with T308 phosphorylation. Moreover, when T34 was mutated to aspartic acid (T34D) to mimic its phosphorylation, Akt-membrane association assessed by surface plasmon resonance spectroscopy was significantly reduced. In cells, this mutation impaired the IGF-induced Akt membrane translocation and subsequent phosphorylation at T308 and S473. Taken together, our results demonstrate that T34 phosphorylation by PDK1 promotes the membrane dissociation of activated Akt for its downstream action through attenuating membrane binding affinity. This membrane dissociation mechanism offers a new insight for Akt activation process and provides a potential new target for controlling the Akt-dependent cellular processes. PMID:25912234

  7. The phosphoinositide kinase PIKfyve is vital in early embryonic development: preimplantation lethality of PIKfyve-/- embryos but normality of PIKfyve+/- mice.

    PubMed

    Ikonomov, Ognian C; Sbrissa, Diego; Delvecchio, Khortnal; Xie, Yufen; Jin, Jian-Ping; Rappolee, Daniel; Shisheva, Assia

    2011-04-15

    Gene mutations in the phosphoinositide-metabolizing enzymes are linked to various human diseases. In mammals, PIKfyve synthesizes PtdIns(3,5)P(2) and PtdIns5P lipids that regulate endosomal trafficking and responses to extracellular stimuli. The consequence of pikfyve gene ablation in mammals is unknown. To clarify the importance of PIKfyve and PIKfyve lipid products, in this study, we have characterized the first mouse model with global deletion of the pikfyve gene using the Cre-loxP approach. We report that nearly all PIKfyve(KO/KO) mutant embryos died before the 32-64-cell stage. Cultured fibroblasts derived from PIKfyve(flox/flox) embryos and rendered pikfyve-null by Cre recombinase expression displayed severely reduced DNA synthesis, consistent with impaired cell division causing early embryo lethality. The heterozygous PIKfyve(WT/KO) mice were born at the expected Mendelian ratio and developed into adulthood. PIKfyve(WT/KO) mice were ostensibly normal by several other in vivo, ex vivo, and in vitro criteria despite the fact that their levels of the PIKfyve protein and in vitro enzymatic activity in cells and tissues were 50-55% lower than those of wild-type mice. Consistently, steady-state levels of the PIKfyve products PtdIns(3,5)P(2) and PtdIns5P selectively decreased, but this reduction (35-40%) was 10-15% less than that expected based on PIKfyve protein reduction. The nonlinear decrease of the PIKfyve protein versus PIKfyve lipid products, the potential mechanism(s) discussed herein, may explain how one functional allele in PIKfyve(WT/KO) mice is able to support the demands for PtdIns(3,5)P(2)/PtdIns5P synthesis during life. Our data also shed light on the known human disorder linked to PIKFYVE mutations. PMID:21349843

  8. Organophosphorous and organochlorine pesticides affect human placental phosphoinositides metabolism and PI-4 kinase activity.

    PubMed

    Souza, María S; Magnarelli de Potas, Gladis; Pechén de D'Angelo, Ana M

    2004-01-01

    The objective of this work was to describe the effect of organophosphorous and organochlorine pesticides on phosphoinositides metabolism in human placenta. Pesticides concentration (10 microM) was used for in vitro incubations of cell-free homogenates labelled with (32)P orthophosphate. Heptachlor (HC) and dichloro-diphenyl-trichloroethane (o-p' DDT) increased phosphatidyl-inositol, phosphatidylinositolphosphate, and phosphatidyl-inositolbiphosphate phosphorylation while azinphosmethyl (AM) increased phosphatidylinositolbiphosphate labeling. Decreased (32)P incorporation in phosphatidylinositol was found with phosmet (PM), AM, and chlorpyriphos (CHL). The effects of these xenobiotics on PI4-kinase activity using different subcellular fractions were also examined. Both type of pesticides affected the postmembrane supernatant enzyme activity. A biphasic effect on membrane and nuclear PI4-kinase activity was seen with HC. The strongest effect found was seen with o-p' DDT in nuclear kinase activity while substantial changes were also observed in membrane. These data demonstrate the sensitivity of human placental PI4-kinase to pesticides currently found in human tissues and suggest deleterious consequences in different processes regulated by 4-phosphoinositides. PMID:14994277

  9. Yeast-based methods to assess PTEN phosphoinositide phosphatase activity in vivo.

    PubMed

    Rodríguez-Escudero, Isabel; Fernández-Acero, Teresa; Bravo, Ignacio; Leslie, Nicholas R; Pulido, Rafael; Molina, María; Cid, Víctor J

    2015-05-01

    The PTEN phosphoinositide 3-phosphatase is a tumor suppressor commonly targeted by pathologic missense mutations. Subject to multiple complex layers of regulation, its capital role in cancer relies on its counteracting function of class I phosphoinositide 3-kinase (PI3K), a key feature in oncogenic signaling pathways. Precise assessment of the involvement of PTEN mutations described in the clinics in loss of catalytic activity requires either tedious in vitro phosphatase assays or in vivo experiments involving transfection into mammalian cell lines. Taking advantage of the versatility of the model organism Saccharomyces cerevisiae, we have developed different functional assays by reconstitution of the mammalian PI3K-PTEN switch in this lower eukaryote. This methodology is based on the fact that regulated PI3K expression in yeast cells causes conversion of PtdIns(4,5)P2 in PtdIns(3,4,5)P3 and co-expression of PTEN counteracts this effect. This can be traced by monitoring growth, given that PtdIns(4,5)P2 pools are essential for the yeast cell, or by using fluorescent reporters amenable for microscopy or flow cytometry. Here we describe the methodology and review its application to evaluate the functionality of PTEN mutations. We show that the technique is amenable to both directed and systematic structure-function relationship studies, and present an example of its use for the study of the recently discovered PTEN-L variant. PMID:25448481

  10. Phosphoinositide Regulation of Integrin Trafficking Required for Muscle Attachment and Maintenance

    PubMed Central

    Ribeiro, Inęs; Yuan, Lin; Tanentzapf, Guy; Dowling, James J.; Kiger, Amy

    2011-01-01

    Muscles must maintain cell compartmentalization when remodeled during development and use. How spatially restricted adhesions are regulated with muscle remodeling is largely unexplored. We show that the myotubularin (mtm) phosphoinositide phosphatase is required for integrin-mediated myofiber attachments in Drosophila melanogaster, and that mtm-depleted myofibers exhibit hallmarks of human XLMTM myopathy. Depletion of mtm leads to increased integrin turnover at the sarcolemma and an accumulation of integrin with PI(3)P on endosomal-related membrane inclusions, indicating a role for Mtm phosphatase activity in endocytic trafficking. The depletion of Class II, but not Class III, PI3-kinase rescued mtm-dependent defects, identifying an important pathway that regulates integrin recycling. Importantly, similar integrin localization defects found in human XLMTM myofibers signify conserved MTM1 function in muscle membrane trafficking. Our results indicate that regulation of distinct phosphoinositide pools plays a central role in maintaining cell compartmentalization and attachments during muscle remodeling, and they suggest involvement of Class II PI3-kinase in MTM-related disease. PMID:21347281

  11. Yunis-Varón Syndrome Is Caused by Mutations in FIG4, Encoding a Phosphoinositide Phosphatase

    PubMed Central

    Campeau, Philippe M.; Lenk, Guy M.; Lu, James T.; Bae, Yangjin; Burrage, Lindsay; Turnpenny, Peter; Román Corona-Rivera, Jorge; Morandi, Lucia; Mora, Marina; Reutter, Heiko; Vulto-van Silfhout, Anneke T.; Faivre, Laurence; Haan, Eric; Gibbs, Richard A.; Meisler, Miriam H.; Lee, Brendan H.

    2013-01-01

    Yunis-Varón syndrome (YVS) is an autosomal-recessive disorder with cleidocranial dysplasia, digital anomalies, and severe neurological involvement. Enlarged vacuoles are found in neurons, muscle, and cartilage. By whole-exome sequencing, we identified frameshift and missense mutations of FIG4 in affected individuals from three unrelated families. FIG4 encodes a phosphoinositide phosphatase required for regulation of PI(3,5)P2 levels, and thus endosomal trafficking and autophagy. In a functional assay, both missense substitutions failed to correct the vacuolar phenotype of Fig4-null mouse fibroblasts. Homozygous Fig4-null mice exhibit features of YVS, including neurodegeneration and enlarged vacuoles in neurons. We demonstrate that Fig4-null mice also have small skeletons with reduced trabecular bone volume and cortical thickness and that cultured osteoblasts accumulate large vacuoles. Our findings demonstrate that homozygosity or compound heterozygosity for null mutations of FIG4 is responsible for YVS, the most severe known human phenotype caused by defective phosphoinositide metabolism. In contrast, in Charcot-Marie-Tooth disease type 4J (also caused by FIG4 mutations), one of the FIG4 alleles is hypomorphic and disease is limited to the peripheral nervous system. This genotype-phenotype correlation demonstrates that absence of FIG4 activity leads to central nervous system dysfunction and extensive skeletal anomalies. Our results describe a role for PI(3,5)P2 signaling in skeletal development and maintenance. PMID:23623387

  12. Galectin-1 augments Ras activation and diverts Ras signals to Raf-1 at the expense of phosphoinositide 3-kinase.

    PubMed

    Elad-Sfadia, Galit; Haklai, Roni; Ballan, Eyal; Gabius, Hans-Joachim; Kloog, Yoel

    2002-10-01

    Ras proteins activate diverse effector molecules. Depending on the cellular context, Ras activation may have different biological consequences: induction of cell proliferation, senescence, survival, or death. Augmentation and selective activation of particular effector molecules may underlie various Ras actions. In fact, Ras effector-loop mutants interacting with distinctive effectors provide evidence for such selectivity. Interactions of active Ras with escort proteins, such as galectin-1, could also direct Ras selectivity. Here we show that in comparison with Ras transfectants, H-Ras/galectin-1 or K-Ras4B/galectin-1 co-transfectants exhibit enhanced and prolonged epidermal growth factor (EGF)-stimulated increases in Ras-GTP, Raf-1 activity, and active extracellular signal-regulated kinase. Galectin-1 antisense RNA inhibited these EGF responses. Conversely, Ras and galectin-1 co-transfection inhibited the EGF-stimulated increase in phosphoinositide 3-kinase (PI3K) activity. Galectin-1 transfection also inhibited Ras(G12V)-induced PI3K but not Raf-1 activity. Galectin-1 co-immunoprecipitated with Ras(G12V) or with Ras(G12V/T35S) that activate Raf-1 but not with Ras(G12V/Y40C) that activates PI3K. Thus, galectin-1 binds active Ras and diverts its signal to Raf-1 at the expense of PI3K. This demonstrates a novel mechanism controlling the duration and selectivity of the Ras signal. Ras gains selectivity when it is associated with galectin-1, mimicking the selectivity of Ras(T35S), which activates Raf-1 but not PI3K. PMID:12149263

  13. Cell autonomous phosphoinositide 3-kinase activation in oocytes disrupts normal ovarian function through promoting survival and overgrowth of ovarian follicles.

    PubMed

    Kim, So-Youn; Ebbert, Katherine; Cordeiro, Marilia H; Romero, Megan; Zhu, Jie; Serna, Vanida Ann; Whelan, Kelly A; Woodruff, Teresa K; Kurita, Takeshi

    2015-04-01

    In this study, we explored the effects of oocytic phosphoinositide 3-kinase (PI3K) activation on folliculogensis by generating transgenic mice, in which the oocyte-specific Cre-recombinase induces the expression of constitutively active mutant PI3K during the formation of primordial follicles. The ovaries of neonatal transgenic (Cre+) mice showed significantly reduced apoptosis in follicles, which resulted in an excess number of follicles per ovary. Thus, the elevation of phosphatidylinositol (3,4,5)-trisphosphate levels within oocytes promotes the survival of follicles during neonatal development. Despite the increase in AKT phosphorylation, primordial follicles in neonatal Cre+ mice remained dormant demonstrating a nuclear accumulation of phosphatase and tensin homolog deleted on chromosome 10 (PTEN). These primordial follicles containing a high level of nuclear PTEN persisted in postpubertal females, suggesting that PTEN is the dominant factor in the maintenance of female reproductive lifespan through the regulation of primordial follicle recruitment. Although the oocytic PI3K activity and PTEN levels were elevated, the activation of primordial follicles and the subsequent accumulation of antral follicles with developmentally competent oocytes progressed normally in prepubertal Cre+ mice. However, mature Cre+ female mice were anovulatory. Because postnatal day 50 Cre+ mice released cumulus-oocyte complexes with developmentally competent oocytes in response to super-ovulation treatment, the anovulatory phenotype was not due to follicular defects but rather endocrine abnormalities, which were likely caused by the excess number of overgrown follicles. Our current study has elucidated the critical role of oocytic PI3K activity in follicular function, as well as the presence of a PTEN-mediated mechanism in the prevention of immature follicle activation. PMID:25594701

  14. Ras is an indispensable coregulator of the class IB phosphoinositide 3-kinase p87/p110gamma.

    PubMed

    Kurig, Barbara; Shymanets, Aliaksei; Bohnacker, Thomas; Prajwal; Brock, Carsten; Ahmadian, Mohammad Reza; Schaefer, Michael; Gohla, Antje; Harteneck, Christian; Wymann, Matthias P; Jeanclos, Elisabeth; Nürnberg, Bernd

    2009-12-01

    Class I(B) phosphoinositide 3-kinase gamma (PI3Kgamma) elicits various immunologic and cardiovascular responses; however, the molecular basis for this signal heterogeneity is unclear. PI3Kgamma consists of a catalytic p110gamma and a regulatory p87(PIKAP) (p87, also p84) or p101 subunit. Hitherto p87 and p101 are generally assumed to exhibit redundant functions in receptor-induced and G protein betagamma (Gbetagamma)-mediated PI3Kgamma regulation. Here we investigated the molecular mechanism for receptor-dependent p87/p110gamma activation. By analyzing GFP-tagged proteins expressed in HEK293 cells, PI3Kgamma-complemented bone marrow-derived mast cells (BMMCs) from p110gamma(-/-) mice, and purified recombinant proteins reconstituted to lipid vesicles, we elucidated a novel pathway of p87-dependent, G protein-coupled receptor (GPCR)-induced PI3Kgamma activation. Although p101 strongly interacted with Gbetagamma, thereby mediating PI3Kgamma membrane recruitment and stimulation, p87 exhibited only a weak interaction, resulting in modest kinase activation and lack of membrane recruitment. Surprisingly, Ras-GTP substituted the missing Gbetagamma-dependent membrane recruitment of p87/p110gamma by direct interaction with p110gamma, suggesting the indispensability of Ras for activation of p87/p110gamma. Consequently, interference with Ras signaling indeed selectively blocked p87/p110gamma, but not p101/p110gamma, kinase activity in HEK293 and BMMC cells, revealing an important crosstalk between monomeric and trimeric G proteins for p87/p110gamma activation. Our data display distinct signaling requirements of p87 and p101, conferring signaling specificity to PI3Kgamma that could open up new possibilities for therapeutic intervention. PMID:19906996

  15. Disulfiram treatment facilitates phosphoinositide 3-kinase inhibition in human breast cancer cells in vitro and in vivo.

    PubMed

    Zhang, Haijun; Chen, Di; Ringler, Jonathan; Chen, Wei; Cui, Qiuzhi Cindy; Ethier, Stephen P; Dou, Q Ping; Wu, Guojun

    2010-05-15

    Frequent genetic alterations of the components in the phosphoinositide 3-kinase (PI3K)/PTEN/AKT signaling pathway contribute greatly to breast cancer initiation and progression, which makes targeting this signaling pathway a promising therapeutic strategy for breast cancer treatment. In this study, we showed that in the presence of copper (Cu), disulfiram (DSF), a clinically used antialcoholism drug, could potently inhibit breast cancer cell growth regardless of the PIK3CA status. Surprisingly, the treatment with a mixture of DSF and copper (DSF-Cu) led to the decreased expression of PTEN protein and the activation of AKT in a dose- and time-dependent manner in different cell lines with or without PIK3CA mutations. Treatment of breast cancer cell lines with a combination of DSF-Cu and LY294002, a pan-PI3K inhibitor, resulted in the significant inhibition of cell growth when compared with either drug alone. In addition, the combined treatment of DSF and LY294002 significantly inhibited the growth of the breast tumor xenograft in nude mice induced by MDA-MB-231 cells expressing mutant PIK3CA-H1047R and PIK3CA-E545K, whereas neither DSF nor LY294002 alone could significantly retard tumor growth. Finally, the observed in vivo inhibitory effects are found associated with aberrant signaling alterations and apoptosis-inducing activities in tumor samples. Thus, our finding shows for the first time that treatment of breast cancer with DSF results in a novel feedback mechanism that activates AKT signaling. Our study also suggests that the combination of DSF and a PI3K inhibitor may offer a new combinational treatment model for breast cancer, particularly for those with PIK3CA mutations. PMID:20424113

  16. Depletion of plasma membrane PtdIns(4,5)P2 reveals essential roles for phosphoinositides in flagellar biogenesis

    PubMed Central

    Wei, Ho-Chun; Rollins, Janet; Fabian, Lacramioara; Hayes, Madeline; Polevoy, Gordon; Bazinet, Christopher; Brill, Julie A.

    2011-01-01

    Summary Axonemes are microtubule-based organelles of crucial importance in the structure and function of eukaryotic cilia and flagella. Despite great progress in understanding how axonemes are assembled, the signals that initiate axoneme outgrowth remain unknown. Here, we identified phosphatidylinositol phosphates (phosphoinositides) as key regulators of early stages of axoneme outgrowth in Drosophila melanogaster spermatogenesis. In a study of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] function in developing Drosophila male germ cells, we depleted PtdIns(4,5)P2 by expression of a potent phosphoinositide phosphatase. Phosphatase expression dramatically inhibited sperm tail formation and perturbed microtubule organization in a manner reversible by co-expression of a PtdIns 4-phosphate 5-kinase. Depletion of PtdIns(4,5)P2 caused increased levels of basal body ?-tubulin and altered the distribution of proteins known to be required for axoneme assembly. Examination of PtdIns(4,5)P2-depleted spermatids by transmission electron microscopy revealed defects in basal body docking to the nuclear envelope, and in axoneme architecture and integrity of the developing flagellar axoneme and axial sheath. Our results provide the first evidence that phosphoinositides act at several steps during flagellar biogenesis, coordinately regulating microtubule and membrane organization. They further suggest that phosphoinositides play evolutionarily conserved roles in flagella and cilia, across phyla and in structurally diverse cell types. PMID:18334551

  17. Interleukin-18 Is a Pro-hypertrophic Cytokine That Acts through a Phosphatidylinositol 3-Kinase-Phosphoinositide-dependent Kinase-

    E-print Network

    Wojcik, Edward J.

    -Phosphoinositide-dependent Kinase- 1-Akt-GATA4 Signaling Pathway in Cardiomyocytes* Received for publication, October 18, 2004, effects that were abolished by Akt inhibitor or knockdown. IL-18 stimulated GATA4 DNA binding activity and increased transcription of a re- porter gene driven by multimerized GATA4-binding DNA elements

  18. Actin and phosphoinositide recruitment to fully-formed Candida albicans phagosomes in mouse macrophages

    PubMed Central

    Heinsbroek, Sigrid E.M.; Kamen, Lynn A.; Taylor, Philip R.; Brown, Gordon D.; Swanson, Joel; Gordon, Siamon

    2010-01-01

    Candida albicans is a dimorphic yeast that enters macrophages via the ?-glucan receptor Dectin-1. Phagocytosis of C. albicans is characterized by actin polymerization, Syk kinase activation and rapid acquisition of phagolysosomal markers. In mice, C. albicans are able to resist the harsh environment of the phagosome and form pseudohyphae inside the phagolysosomal compartment, eventually extending from the macrophage. In this study, we investigated these unique C. albicans phagosomes and found that actin dynamically localized around the phagosomes, before disintegrating. Membrane phosphoinositides, PI(4,5)P2, PI(3,4,5)P3, PI(3,4)P2, and PI(3)P also localized to the phagosomes. This localization was not related to actin polymerization and inhibitor studies showed that polymerization of actin on the C. albicans phagosome was independent of PI3K. The ability of mature C. albicans phagosomes to stimulate actin polymerization could facilitate the escape of the growing yeast from the macrophage. PMID:20375582

  19. The phosphoinositide 3-kinase pathway is crucial for the growth of canine mast cell tumors.

    PubMed

    Amagai, Yosuke; Tanaka, Akane; Matsuda, Akira; Oida, Kumiko; Jung, Kyungsook; Matsuda, Hiroshi

    2013-01-01

    Mast cell tumors (MCTs) are the most common tumors in dogs, accounting for 16-21% of cutaneous tumors. Although several small molecule inhibitors, including imatinib mesylate, have been used for the treatment of MCTs, the response rate remains less than 50%. In this study, the effects of different selective signal inhibitors on MCT cell growth were evaluated using 4 different cell lines derived from dogs. We found that the phosphoinositide 3-kinase (PI3K) signaling pathway is crucial for the proliferation of MCT cells in the presence or absence of c-kit gene mutations. Here, we propose a novel therapeutic strategy to target the PI3K pathway for the treatment of canine MCTs. PMID:23328607

  20. Phosphoinositide 3-kinase ? mediates microglial phagocytosis via lipid kinase-independent control of cAMP.

    PubMed

    Schmidt, C; Schneble, N; Müller, J P; Bauer, R; Perino, A; Marone, R; Rybalkin, S D; Wymann, M P; Hirsch, E; Wetzker, R

    2013-03-13

    Microglial phagocytosis plays a key role in neuroprotective and neurodegenerative responses of the innate immune system in the brain. Here we investigated the regulatory function of phosphoinositide 3-kinase ? (PI3K?) in phagocytosis of bacteria and Zymosan particles by mouse brain microglia in vitro and in vivo. Using genetic and pharmacological approaches our data revealed PI3K? as an essential mediator of microglial phagocytosis. Unexpectedly, microglia expressing lipid kinase deficient mutant PI3K? exhibited similar phagocytosis as wild-type cells. These data suggest kinase-independent stimulation of cAMP phosphodiesterase activity by PI3K? as a crucial mediator of phagocytosis. In sum our findings indicate PI3K?-dependent suppression of cAMP signaling as a critical regulatory element of microglial phagocytosis. PMID:23276671

  1. Central role for G protein-coupled phosphoinositide 3-kinase gamma in inflammation.

    PubMed

    Hirsch, E; Katanaev, V L; Garlanda, C; Azzolino, O; Pirola, L; Silengo, L; Sozzani, S; Mantovani, A; Altruda, F; Wymann, M P

    2000-02-11

    Phosphoinositide 3-kinase (PI3K) activity is crucial for leukocyte function, but the roles of the four receptor-activated isoforms are unclear. Mice lacking heterotrimeric guanine nucleotide-binding protein (G protein)-coupled PI3Kgamma were viable and had fully differentiated neutrophils and macrophages. Chemoattractant-stimulated PI3Kgamma-/- neutrophils did not produce phosphatidylinositol 3,4,5-trisphosphate, did not activate protein kinase B, and displayed impaired respiratory burst and motility. Peritoneal PI3Kgamma-null macrophages showed a reduced migration toward a wide range of chemotactic stimuli and a severely defective accumulation in a septic peritonitis model. These results demonstrate that PI3Kgamma is a crucial signaling molecule required for macrophage accumulation in inflammation. PMID:10669418

  2. Phosphoinositide-signaling is one component of a robust plant defense response

    PubMed Central

    Hung, Chiu-Yueh; Aspesi Jr, Peter; Hunter, Melissa R.; Lomax, Aaron W.; Perera, Imara Y.

    2014-01-01

    The phosphoinositide pathway and inositol-1,4,5-triphosphate (InsP3) have been implicated in plant responses to many abiotic stresses; however, their role in response to biotic stress is not well characterized. In the current study, we show that both basal defense and systemic acquired resistance responses are affected in transgenic plants constitutively expressing the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase) which have greatly reduced InsP3 levels. Flagellin induced Ca2+-release as well as the expressions of some flg22 responsive genes were attenuated in the InsP 5-ptase plants. Furthermore, the InsP 5-ptase plants were more susceptible to virulent and avirulent strains of Pseudomonas syringae pv. tomato (Pst) DC3000. The InsP 5-ptase plants had lower basal salicylic acid (SA) levels and the induction of SAR in systemic leaves was reduced and delayed. Reciprocal exudate experiments showed that although the InsP 5-ptase plants produced equally effective molecules that could trigger PR-1 gene expression in wild type plants, exudates collected from either wild type or InsP 5-ptase plants triggered less PR-1 gene expression in InsP 5-ptase plants. Additionally, expression profiles indicated that several defense genes including PR-1, PR-2, PR-5, and AIG1 were basally down regulated in the InsP 5-ptase plants compared with wild type. Upon pathogen attack, expression of these genes was either not induced or showed delayed induction in systemic leaves. Our study shows that phosphoinositide signaling is one component of the plant defense network and is involved in both basal and systemic responses. The dampening of InsP3-mediated signaling affects Ca2+ release, modulates defense gene expression and compromises plant defense responses. PMID:24966862

  3. Pharmacological characterization of the phosphoinositide second messenger system in the rabbit kidney

    SciTech Connect

    McArdle, S.

    1988-01-01

    The cellular response to hormones and neurotransmitters is a result of receptor activation of a second messenger system to initiate the intracellular cascade. In several tissues, such as brain and liver, one of the second messenger systems involves the hydrolysis of phosphoinositides (PIs) for the formation of inositol phosphate and diacylglycerol as the intracellular messengers. In the present study, they examined the effect of various agents on the hydrolysis of PIs in the rabbit kidney. In the kidney, the effect of the various hormones and neurotransmitters was region specific. Hydrolysis of PIs was stimulated in the inner medulla by (arg{sup 8})-vasopressin, angiotensin II, and atriopeptin I, and in the outer medulla by histamine, adenosine, and secretin. Only carbachol was able to stimulate the hydrolysis of PIs in both the inner and outer medulla. None of the substances tested were able to stimulate this response in the cortex. The following agents did not have an effect in any of the three zones of the kidney: norepinephrine, dopamine, atriopeptins II, and III. They have directly demonstrated the presence of a high affinity saturable binding site on inner medullary collecting duct (IMCD) cells with studies of binding characteristics of the radiolabelled muscarinic antagonist, 1-quinuclidinyl (phenyl-4-{sup 3}H) benzilate (({sup 3}H)QNB). The K{sub d} of 0.27 nM and the B{sub max} of 27.5 fmol/mg protein were determined from Scatchard analysis of the saturation data. In summary, they have demonstrated that cholinergic muscarinic receptors are present in the rabbit kidney, specifically in the IMCD cells. These receptors, which are coupled to the hydrolysis of phosphoinositides, may be involved in the vasodilatory and/or diuretic effects of cholinergic agents.

  4. Inhibition by taurine of the phosphorylation of specific synaptosomal proteins in the rat cortex: effects of taurine on the stimulation of calcium uptake in mitochondria and inhibition of phosphoinositide turnover.

    PubMed

    Li, Y P; Lombardini, J B

    1991-07-01

    It has been previously observed that taurine inhibits PKC-activated phosphorylation of specific proteins including a approximately 20k Mr protein in rat cortical synaptosomes. In the present study, the mechanism of the above effects of taurine were investigated. In an intrasynaptosomal cytosol fraction obtained by subcellular fractionation, taurine did not have inhibitory effects on protein phosphorylation. However, taurine did inhibit the phosphorylation of the approximately 20k Mr protein in a reconstituted preparation containing intrasynaptosomal cytosol and mitochondria. Experiments measuring calcium uptake demonstrated that taurine increased the accumulation of 45Ca2+ in the mitochondrial fraction in incubation systems both in the absence and presence of added ATP. In addition, taurine inhibited the accumulation of 32P-labeled phosphatidic acid in synaptosomes indicative of a reduction in the levels of diacylglycerol. These results suggest that taurine may inhibit specific protein phosphorylation both by reducing cytosolic calcium levels and by inhibiting the turnover of phosphoinositides. These effects of taurine on the signal transduction cascade involving PKC and phosphoinositide metabolism indicate a potential biological role for taurine in the nervous system. PMID:1657280

  5. Antitumor effect of resveratrol on chondrosarcoma cells via phosphoinositide 3-kinase/AKT and p38 mitogen-activated protein kinase pathways.

    PubMed

    Dai, Zixun; Lei, Pengfei; Xie, Jie; Hu, Yihe

    2015-08-01

    Chondrosarcoma is one of the most common types of primary bone cancer that develops in cartilage cells. Resveratrol (Res), a natural polyphenol compound isolated from various fruits, has a suppressive effect on various human malignancies. It has been shown that Res inhibits matrix metalloproteinase (MMP)?induced differentiation in chondrosarcoma cells. However, the effects of Res on cell proliferation, apoptosis and invasion of chondrosarcoma cells, as well as the underlying mechanisms, remain largely unknown. To the best of our knowledge, the present study showed for the first time that Res inhibited proliferation and induced apoptosis in chondrosarcoma cells in a dose-dependent manner. Furthermore, it was shown that Res also suppressed chondrosarcoma cell invasion in a dose-dependent manner, probably via the inhibition of MMP2 and MMP9 protein expression. Molecular mechanism investigations revealed that Res could inhibit the activity of phosphoinositide 3-kinase/AKT and p38 mitogen?activated protein kinase signaling pathways, which has been demonstrated to be important in the regulation of proliferation, apoptosis and invasion in various cancer cell types. In conclusion, this study suggests that Res may serve as a promising agent for the treatment of chondrosarcoma. PMID:25936406

  6. Fucoidan inhibits the migration and proliferation of HT-29 human colon cancer cells via the phosphoinositide-3 kinase/Akt/mechanistic target of rapamycin pathways.

    PubMed

    Han, Yong-Seok; Lee, Jun Hee; Lee, Sang Hun

    2015-09-01

    Fucoidan, a sulfated polysaccharide, has a variety of biological activities, including anti-cancer, anti-angiogenic and anti-inflammatory effects. However, the underlying mechanisms of fucoidan as an anti?cancer agent remain to be elucidated. The present study examined the anti?cancer effect of fucoidan on HT?29 human colon cancer cells. The cell growth of HT29 cells was significantly decreased following treatment with fucoidan (200 µg/ml). In addition, fucoidan inhibited the migration of HT?29 cells by decreasing the expression levels of matrix metalloproteinase?2 in a dose?dependent manner (0?200 µg/ml). The underlying mechanism of these inhibitory effects included the downregulation of phosphoinositide 3?kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) by treatment with fucoidan. Furthermore, fucoidan increased the expression of cleaved caspase?3 and decreased cancer sphere formation. The present study suggested that fucoidan exerts an anti?cancer effect on HT?29 human colon cancer cells by downregulating the PI3K?Akt?mTOR signaling pathway. Therefore, fucoidan may be a potential therapeutic reagent against the growth of human colon cancer cells. PMID:25998232

  7. Requirement for phosphoinositide 3-OH kinase in growth hormone signalling to the mitogen-activated protein kinase and p70s6k pathways.

    PubMed Central

    Kilgour, E; Gout, I; Anderson, N G

    1996-01-01

    Pituitary growth hormone (GH) co-ordinately stimulates three distinct signalling pathways in 3T3-F442A preadipocytes, the STAT (signal transducer and activator of transcription) pathway, the mitogen-activated protein (MAP) kinase cascade and p70s6k. The mechanisms linking the GH receptor to these signals have not been fully identified. In this study we have examined the role of phosphoinositide 3-OH kinase (PI 3-kinase). Pretreatment of cells with wortmannin, a specific inhibitor of PI 3-kinase, prevented the activation of p70s6k and partially inhibited the activation of p42 and p44 MAP kinases by GH. In contrast, wortmannin failed to appreciably affect the GH-stimulated tyrosyl phosphorylation of JAK-2 or STAT-1. GH transiently increased the activity of PI 3-kinase recovered in antiphosphotyrosine immunoprecipitates. In addition, several tyrosyl-phosphorylated proteins were specifically adsorbed from lysates of cells exposed to GH by a glutathione S-transferase fusion protein containing the 85 kDa regulatory subunit of PI 3-kinase. GH also induced an increase in the PI 3-kinase activity associated with both JAK-2 and insulin receptor substrate-1 (IRS-1) immunoprecipitates. These results establish PI 3-kinase as an important mediator of GH signalling to the MAP kinase and p70s6k pathways and suggest that PI 3-kinase is activated by a mechanism involving JAK-2 and IRS-1. PMID:8615823

  8. In utero hypoxic ischemia decreases the cholinergic agonist-stimulated poly-phosphoinositide turnover in the developing rat brain

    Microsoft Academic Search

    K. Hersey; Z. Y. Hu; J. P. Zhang; P. G. Rhodes; G. Y. Sun

    1995-01-01

    Perinatal hypoxic-ischemic (HI) insult is known to cause cellular and molecular disturbances leading to functional and behavioral abnormalities during brain development. In this study, we examined the effects of an in utero HI insult on poly-phosphoinositide turnover in vivo in the cerebrum and cerebellum as well as cholinergic-stimulated turnover in cortical slices from developing rat brain. In utero HI treatment

  9. Wortmannin Inactivates Phosphoinositide 3Kinase by Covalent Modification of Lys802, a Residue Involved in the Phosphate Transfer Reaction

    Microsoft Academic Search

    MATTHIAS P. WYMANN; GINETTE BULGARELLI-LEVA; MARKETA J. ZVELEBIL; LUCIANO PIROLA; BART VANHAESEBROECK; MICHAEL D. WATERFIELD

    Wortmannin at nanomolar concentrations is a potent and specific inhibitor of phosphoinositide (PI) 3-kinase and has been used extensively to demonstrate the role of this enzyme in diverse signal transduction processes. At higher concentrations, wortmannin inhibits the ataxia telangiectasia gene (ATM)-related DNA- dependent protein kinase (DNA-PKcs). We report here the identification of the site of interaction of wortman- nin on

  10. Pharmacological Characterization of Endothelin-stimulated Phosphoinositide Breakdown and Cytosolic Free Ca2+ Rise in Rat C, Glioma Cells

    Microsoft Academic Search

    Wan-Wan Lin; Juliann G. Kiang; De-Maw Chuangl

    1992-01-01

    Because increasing evidence indicates that glial cells are a target of endothelin, we have characterized endothelin-in- duced phosphoinositide (PI) turnover and Ca2+ homeostasis in C,glioma cells. Endothelin-1 (ET) increased formation of 3H-inositol phosphate (IP) from PI and elicited an increase in cytosolic free Ca*+ ((Caz+)J in rat C,glioma. In the presence of Li+, the increase in 3H-inositol trisphosphateformation was rapid,

  11. Structural basis for different phosphoinositide specificities of the PX domains of sorting nexins regulating G-protein signaling.

    PubMed

    Mas, Caroline; Norwood, Suzanne J; Bugarcic, Andrea; Kinna, Genevieve; Leneva, Natalya; Kovtun, Oleksiy; Ghai, Rajesh; Ona Yanez, Lorena E; Davis, Jasmine L; Teasdale, Rohan D; Collins, Brett M

    2014-10-10

    Sorting nexins (SNXs) or phox homology (PX) domain containing proteins are central regulators of cell trafficking and signaling. A subfamily of PX domain proteins possesses two unique PX-associated domains, as well as a regulator of G protein-coupled receptor signaling (RGS) domain that attenuates G?s-coupled G protein-coupled receptor signaling. Here we delineate the structural organization of these RGS-PX proteins, revealing a protein family with a modular architecture that is conserved in all eukaryotes. The one exception to this is mammalian SNX19, which lacks the typical RGS structure but preserves all other domains. The PX domain is a sensor of membrane phosphoinositide lipids and we find that specific sequence alterations in the PX domains of the mammalian RGS-PX proteins, SNX13, SNX14, SNX19, and SNX25, confer differential phosphoinositide binding preferences. Although SNX13 and SNX19 PX domains bind the early endosomal lipid phosphatidylinositol 3-phosphate, SNX14 shows no membrane binding at all. Crystal structures of the SNX19 and SNX14 PX domains reveal key differences, with alterations in SNX14 leading to closure of the binding pocket to prevent phosphoinositide association. Our findings suggest a role for alternative membrane interactions in spatial control of RGS-PX proteins in cell signaling and trafficking. PMID:25148684

  12. Ion channel regulation by phosphoinositides analyzed with VSPs—PI(4,5)P2 affinity, phosphoinositide selectivity, and PI(4,5)P2 pool accessibility

    PubMed Central

    Rjasanow, Alexandra; Leitner, Michael G.; Thallmair, Veronika; Halaszovich, Christian R.; Oliver, Dominik

    2015-01-01

    The activity of many proteins depends on the phosphoinositide (PI) content of the membrane. E.g., dynamic changes of the concentration of PI(4,5)P2 are cellular signals that regulate ion channels. The susceptibility of a channel to such dynamics depends on its affinity for PI(4,5)P2. Yet, measuring affinities for endogenous PIs has not been possible directly, but has relied largely on the response to soluble analogs, which may not quantitatively reflect binding to native lipids. Voltage-sensitive phosphatases (VSPs) turn over PI(4,5)P2 to PI(4)P when activated by depolarization. In combination with voltage-clamp electrophysiology VSPs are useful tools for rapid and reversible depletion of PI(4,5)P2. Because cellular PI(4,5)P2 is resynthesized rapidly, steady state PI(4,5)P2 changes with the degree of VSP activation and thus depends on membrane potential. Here we show that titration of endogenous PI(4,5)P2 with Ci-VSP allows for the quantification of relative PI(4,5)P2 affinities of ion channels. The sensitivity of inward rectifier and voltage-gated K+ channels to Ci-VSP allowed for comparison of PI(4,5)P2 affinities within and across channel subfamilies and detected changes of affinity in mutant channels. The results also reveal that VSPs are useful only for PI effectors with high binding specificity among PI isoforms, because PI(4,5)P2 depletion occurs at constant overall PI level. Thus, Kir6.2, a channel activated by PI(4,5)P2 and PI(4)P was insensitive to VSP. Surprisingly, despite comparable PI(4,5)P2 affinity as determined by Ci-VSP, the Kv7 and Kir channel families strongly differed in their sensitivity to receptor-mediated depletion of PI(4,5)P2. While Kv7 members were highly sensitive to activation of PLC by Gq-coupled receptors, Kir channels were insensitive even when PI(4,5)P2 affinity was lowered by mutation. We hypothesize that different channels may be associated with distinct pools of PI(4,5)P2 that differ in their accessibility to PLC and VSPs.

  13. Osteopontin is a myosphere-derived secretory molecule that promotes angiogenic progenitor cell proliferation through the phosphoinositide 3-kinase/Akt pathway

    SciTech Connect

    Ogata, Takehiro [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan); Ueyama, Tomomi [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan)]. E-mail: tueyama@kuhp.kyoto-u.ac.jp; Nomura, Tetsuya [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan); Department of Cardiovascular Medicine, Kyoto Prefectural University School of Medicine, Kyoto 602-8566 (Japan); Asada, Satoshi [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan); Department of Cardiovascular Medicine, Kyoto Prefectural University School of Medicine, Kyoto 602-8566 (Japan); Tagawa, Masashi [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan); Department of Cardiovascular Medicine, Kyoto Prefectural University School of Medicine, Kyoto 602-8566 (Japan); Nakamura, Tomoyuki [Department of Pharmacology, Kansai Medical University, Moriguchi, Osaka 570-8507 (Japan); Takahashi, Tomosaburo [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan); Department of Cardiovascular Medicine, Kyoto Prefectural University School of Medicine, Kyoto 602-8566 (Japan); Matsubara, Hiroaki [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan); Department of Cardiovascular Medicine, Kyoto Prefectural University School of Medicine, Kyoto 602-8566 (Japan); Oh, Hidemasa [Department of Experimental Therapeutics, Translational Research Center, Kyoto University Hospital, Kyoto 606-8507 (Japan)]. E-mail: hidemasa@kuhp.kyoto-u.ac.jp

    2007-07-27

    We have reported that skeletal myosphere-derived progenitor cells (MDPCs) can differentiate into vascular cells, and that MDPC transplantation into cardiomyopathic hearts improves cardiac function. However, the autocrine/paracrine molecules and underlying mechanisms responsible for MDPC growth have not yet been determined. To explore the molecules enhancing the proliferation of MDPCs, we performed serial analysis of gene expression and signal sequence trap methods using RNA isolated from MDPCs. We identified osteopontin (OPN), a secretory molecule, as one of most abundant molecules expressed in MDPCs. OPN provided a proliferative effect for MDPCs. MDPCs treated with OPN showed Akt activation, and inhibition of the phosphoinositide 3-kinase (PI3K)/Akt pathway repressed the proliferative effect of OPN. Furthermore, OPN-pretreated MDPCs maintained their differentiation potential into endothelial and vascular smooth muscle cells. These findings indicate an important role of OPN as an autocrine/paracrine molecule in regulating the proliferative growth of muscle-derived angiogenic progenitor cells via the PI3K/Akt pathway.

  14. Phosphoinositide 3-kinase inhibition restores neutrophil accuracy in the elderly: toward targeted treatments for immunosenescence.

    PubMed

    Sapey, Elizabeth; Greenwood, Hannah; Walton, Georgia; Mann, Elizabeth; Love, Alexander; Aaronson, Natasha; Insall, Robert H; Stockley, Robert A; Lord, Janet M

    2014-01-01

    Immunosenescence is the functional deterioration of the immune system during natural aging. Despite increased susceptibility to bacterial infections in older adults, age-associated changes to neutrophil responses are only partially understood, and neutrophil migration has not been characterized in detail. Here we describe reduced chemotaxis but preserved chemokinesis toward a range of inflammatory stimuli in migrating neutrophils isolated from healthy older subjects. Cross-sectional data indicate that migratory behavior changes in the sixth decade of life. Crucially, aberrant migration may increase "bystander" tissue damage and heighten inflammation as a result of excess proteinase release during inaccurate chemotaxis, as well as reducing pathogen clearance. We show evidence of increased neutrophil proteinase activity in older adults, namely, raised levels of neutrophil proteinase substrate-derived peptides and evidence of primary granule release, associated with increased systemic inflammation. Inaccurate migration was causally associated with increased constitutive phosphoinositide 3-kinase (PI3K) signaling; untreated neutrophils from old donors demonstrated significant PI3K activation compared with cells from young donors. PI3K-blocking strategies, specifically inhibition of PI3K? or PI3K?, restored neutrophil migratory accuracy, whereas SHIP1 inhibition worsened migratory flaws. Targeting PI3K signaling may therefore offer a new strategy in improving neutrophil functions during infections and reduce inappropriate inflammation in older patients. PMID:24191150

  15. TRAF4 Is a Novel Phosphoinositide-Binding Protein Modulating Tight Junctions and Favoring Cell Migration

    PubMed Central

    Rousseau, Adrien; McEwen, Alastair G.; Poussin-Courmontagne, Pierre; Rognan, Didier; Nominé, Yves; Rio, Marie-Christine; Tomasetto, Catherine; Alpy, Fabien

    2013-01-01

    Tumor necrosis factor (TNF) receptor-associated factor 4 (TRAF4) is frequently overexpressed in carcinomas, suggesting a specific role in cancer. Although TRAF4 protein is predominantly found at tight junctions (TJs) in normal mammary epithelial cells (MECs), it accumulates in the cytoplasm of malignant MECs. How TRAF4 is recruited and functions at TJs is unclear. Here we show that TRAF4 possesses a novel phosphoinositide (PIP)-binding domain crucial for its recruitment to TJs. Of interest, this property is shared by the other members of the TRAF protein family. Indeed, the TRAF domain of all TRAF proteins (TRAF1 to TRAF6) is a bona fide PIP-binding domain. Molecular and structural analyses revealed that the TRAF domain of TRAF4 exists as a trimer that binds up to three lipids using basic residues exposed at its surface. Cellular studies indicated that TRAF4 acts as a negative regulator of TJ and increases cell migration. These functions are dependent from its ability to interact with PIPs. Our results suggest that TRAF4 overexpression might contribute to breast cancer progression by destabilizing TJs and favoring cell migration. PMID:24311986

  16. Nonenzymatic domains of Kalirin7 contribute to spine morphogenesis through interactions with phosphoinositides and Abl

    PubMed Central

    Ma, Xin-Ming; Miller, Megan B.; Vishwanatha, K. S.; Gross, Maegan J.; Wang, Yanping; Abbott, Thomas; Lam, TuKiet T.; Mains, Richard E.; Eipper, Betty A.

    2014-01-01

    Like several Rho GDP/GTP exchange factors (GEFs), Kalirin7 (Kal7) contains an N-terminal Sec14 domain and multiple spectrin repeats. A natural splice variant of Kalrn lacking the Sec14 domain and four spectrin repeats is unable to increase spine formation; our goal was to understand the function of the Sec14 and spectrin repeat domains. Kal7 lacking its Sec14 domain still increased spine formation, but the spines were short. Strikingly, Kal7 truncation mutants containing only the Sec14 domain and several spectrin repeats increased spine formation. The Sec14 domain bound phosphoinositides, a minor but crucial component of cellular membranes, and binding was increased by a phosphomimetic mutation. Expression of KalSec14-GFP in nonneuronal cells impaired receptor-mediated endocytosis, linking Kal7 to membrane trafficking. Consistent with genetic studies placing Abl, a non–receptor tyrosine kinase, and the Drosophila orthologue of Kalrn into the same signaling pathway, Abl1 phosphorylated two sites in the fourth spectrin repeat of Kalirin, increasing its sensitivity to calpain-mediated degradation. Treating cortical neurons of the wild-type mouse, but not the Kal7KO mouse, with an Abl inhibitor caused an increase in linear spine density. Phosphorylation of multiple sites in the N-terminal Sec14/spectrin region of Kal7 may allow coordination of the many signaling pathways contributing to spine morphogenesis. PMID:24600045

  17. Assessing the subcellular distribution of oncogenic phosphoinositide 3-kinase using microinjection into live cells.

    PubMed

    Layton, Meredith J; Rynkiewicz, Natalie; Ivetac, Ivan; Horan, Kristy A; Mitchell, Christina A; Phillips, Wayne A

    2014-03-01

    Oncogenic mutations in PIK3CA lead to an increase in instrinsic phosphoinositide kinase activity, but it is thought that increased access of PI3K? to its plasma membrane localised substrate is also required for increased levels of downstream PIP3/Akt signalling. We have studied the subcellular localisation of wild type and the two most common oncogenic mutants of PI3K? in cells maintained in growth media, and starved or stimulated cells using a novel method in which PI3K? is pre-formed as a 1:1 p110?:p85? complex in vitro then introduced into live cells by microinjection. Oncogenic E545K and H1047R mutants did not constitutively interact with membrane lipid in vitro or in cells maintained in 10% FCS. Following stimulation of receptor tyrosine kinases, microinjected PI3K? was recruited to the plasma membrane, but oncogenic forms of PI3K? were not recruited to the plasma membrane to a greater extent and did not reside at the plasma membrane longer than wild type PI3K?. Instead, the E545K mutant specifically bound activated Cdc42 in vitro and microinjection of E545K was associated with the formation of cellular protrusions, providing some preliminary evidence that changes in protein-protein interactions may play a role in the oncogenicity of the E545K mutant in addition to the well-known changes in lipid kinase activity. PMID:24597785

  18. Effect of albumin-bound DHA on phosphoinositide phosphorylation in collagen stimulated human platelets

    SciTech Connect

    Gaudette, D.C.; Holub, B.J. (Univ. of Guelph, Ontario (Canada))

    1990-05-15

    The effect of exogenous albumin-bound docosahexaenoic acid (22:6n-3) (DHA), arachidonic acid (20:4n-6) (AA), and eicosapendaenoic acid (20:5n-3) (EPA) on phosphoinositide metabolism following collagen stimulation was studied using (3H)inositol prelabelled platelets. Collagen stimulation (3 min, 1.8 micrograms/ml) increased the labelling of both phosphatidylinositol 4-monophosphate (PIP), and phosphatidylinositol 4,5-biphosphate (PIP2). Of the fatty acids tested, only pre-incubation (2 min) with DHA (20 microM) significantly attenuated the collagen-induced increased PIP and PIP2 labelling; EPA was without effect, while AA enhanced PIP labelling. Forty microM DHA was less effective at attenuating the increased PIP and PIP2 labelling even though this concentration of DHA resulted in greater inhibition of platelet aggregation. Neither concentration of DHA attenuated the increased polyphosphoinositide labelling resulting from stimulation by the endoperoxide analogue U46619, or the phorbol ester, PMA. These data suggest that the effect of DHA on attenuating the increased PIP and PIP2 labelling following collagen stimulation likely occurs before thromboxane receptor occupancy, may not occur at the level of protein kinase C activation, and could be mediated in part via a lessened synthesis of thromboxane A2.

  19. Ablation of phosphoinositide-3-kinase class II alpha suppresses hepatoma cell proliferation

    SciTech Connect

    Ng, Stanley K.L. [Singapore Immunology Network A-STAR (Singapore)] [Singapore Immunology Network A-STAR (Singapore); Neo, Soek-Ying, E-mail: neo_soek_ying@sics.a-star.edu.sg [Singapore Immunology Network A-STAR (Singapore)] [Singapore Immunology Network A-STAR (Singapore); Yap, Yann-Wan [Singapore Immunology Network A-STAR (Singapore)] [Singapore Immunology Network A-STAR (Singapore); Karuturi, R. Krishna Murthy; Loh, Evelyn S.L. [Genome Institute of Singapore A-STAR (Singapore)] [Genome Institute of Singapore A-STAR (Singapore); Liau, Kui-Hin [Department of General Surgery, Tan Tock Seng Hospital (Singapore)] [Department of General Surgery, Tan Tock Seng Hospital (Singapore); Ren, Ee-Chee, E-mail: ren_ee_chee@immunol.a-star.edu.sg [Singapore Immunology Network A-STAR (Singapore) [Singapore Immunology Network A-STAR (Singapore); Department of Microbiology, Yong Loo Lin School of Medicine, National University of Singapore (Singapore)

    2009-09-18

    Cancer such as hepatocellular carcinoma (HCC) is characterized by complex perturbations in multiple signaling pathways, including the phosphoinositide-3-kinase (PI3K/AKT) pathways. Herein we investigated the role of PI3K catalytic isoforms, particularly class II isoforms in HCC proliferation. Among the siRNAs tested against the eight known catalytic PI3K isoforms, specific ablation of class II PI3K alpha (PIK3C2{alpha}) was the most effective in impairing cell growth and this was accompanied by concomitant decrease in PIK3C2{alpha} mRNA and protein levels. Colony formation ability of cells deficient for PIK3C2{alpha} was markedly reduced and growth arrest was associated with increased caspase 3 levels. A small but significant difference in gene dosage and expression levels was detected between tumor and non-tumor tissues in a cohort of 19 HCC patients. Taken together, these data suggest for the first time that in addition to class I PI3Ks in cancer, class II PIK3C2{alpha} can modulate HCC cell growth.

  20. Phosphoinositide-3 kinase ? required for LPS-induced transepithelial neutrophil trafficking in the lung

    PubMed Central

    Reutershan, Jörg; Saprito, Mary S.; Wu, Dan; Rückle, Thomas; Ley, Klaus

    2009-01-01

    Phosphoinositide 3-kinase ? (PI3K?) is a critical mediator of directional cell movement. Here, we sought to characterize the role of PI3K? in mediating the different steps of PMN trafficking in the lung. In a murine model of LPS-induced lung injury, PMN migration into the different lung compartments was determined in PI3K? gene-deficient (PI3K??/?) and wildtype mice. Bone marrow chimeras were created to characterize the role of PI3K? on hematopoietic vs. non-hematopoietic cells. A small molecule PI3K? inhibitor was tested in vitro and in vivo. PMN adhesion to the pulmonary endothelium and transendothelial migration into the lung interstitium was enhanced in PI3K??/? mice. However, transepithelial migration into the alveolar space was reduced in these mice. When irradiated PI3K??/? mice were reconstituted with bone marrow from wildtype mice, migratory activity into the alveolar space was restored partially. A small molecule PI3K? inhibitor reduced chemokine-induced PMN migration in vitro when PMNs or epithelial cells but not when endothelial cells were treated. The inhibitor also reduced LPS-induced PMN migration in vivo. We conclude that PI3K? is required for transepithelial but not for transendothelial migration in LPS-induced lung injury. Inhibition of PI3K? activity may be effective at curbing excessive PMN infiltration in lung injury. PMID:19797129

  1. Essential Role of the p110? Subunit of Phosphoinositide 3-OH Kinase in Male Fertility

    PubMed Central

    Ciraolo, Elisa; Morello, Fulvio; Hobbs, Robin M.; Wolf, Frieder; Marone, Romina; Iezzi, Manuela; Lu, Xiaoyun; Mengozzi, Giulio; Altruda, Fiorella; Sorba, Giovanni; Guan, Kaomei; Pandolfi, Pier Paolo; Wymann, Matthias P.

    2010-01-01

    Phosphoinositide 3-kinases (PI3K) are key molecular players in male fertility. However, the specific roles of different p110 PI3K catalytic subunits within the spermatogenic lineage have not been characterized so far. Herein, we report that male mice expressing a catalytically inactive p110? develop testicular hypotrophy and impaired spermatogenesis, leading to a phenotype of oligo-azoospermia and defective fertility. The examination of testes from p110?-defective tubules demonstrates a widespread loss in spermatogenic cells, due to defective proliferation and survival of pre- and postmeiotic cells. In particular, p110? is crucially needed in c-Kit–mediated spermatogonial expansion, as c-Kit–positive cells are lost in the adult testis and activation of Akt by SCF is blocked by a p110? inhibitor. These data establish that activation of the p110? PI3K isoform by c-Kit is required during spermatogenesis, thus opening the way to new treatments for c-Kit positive testicular cancers. PMID:20053680

  2. Ablation of phosphoinositide 3-kinase-gamma reduces the severity of acute pancreatitis.

    PubMed

    Lupia, Enrico; Goffi, Alberto; De Giuli, Paolo; Azzolino, Ornella; Bosco, Ornella; Patrucco, Enrico; Vivaldo, Maria Cristina; Ricca, Marco; Wymann, Matthias P; Hirsch, Emilio; Montrucchio, Giuseppe; Emanuelli, Giorgio

    2004-12-01

    In pancreatic acini, the G-protein-activated phosphoinositide 3-kinase-gamma (PI3K gamma) regulates several key pathological responses to cholecystokinin hyperstimulation in vitro. Thus, using mice lacking PI3K gamma, we studied the function of this enzyme in vivo in two different models of acute pancreatitis. The disease was induced by supramaximal concentrations of cerulein and by feeding mice a choline-deficient/ethionine-supplemented diet. Although the secretive function of isolated pancreatic acini was identical in mutant and control samples, in both models, genetic ablation of PI3K gamma significantly reduced the extent of acinar cell injury/necrosis. In agreement with a protective role of apoptosis in pancreatitis, PI3K gamma-deficient pancreata showed an increased number of apoptotic acinar cells, as determined by terminal dUTP nick-end labeling and caspase-3 activity. In addition, neutrophil infiltration within the pancreatic tissue was also reduced, suggesting a dual action of PI3K gamma, both in the triggering events within acinar cells and in the subsequent neutrophil recruitment and activation. Finally, the lethality of the choline-deficient/ethionine-supplemented diet-induced pancreatitis was significantly reduced in mice lacking PI3K gamma. Our results thus suggest that inhibition of PI3K gamma may be of therapeutic value in acute pancreatitis. PMID:15579443

  3. Ablation of Phosphoinositide 3-Kinase-? Reduces the Severity of Acute Pancreatitis

    PubMed Central

    Lupia, Enrico; Goffi, Alberto; De Giuli, Paolo; Azzolino, Ornella; Bosco, Ornella; Patrucco, Enrico; Vivaldo, Maria Cristina; Ricca, Marco; Wymann, Matthias P.; Hirsch, Emilio; Montrucchio, Giuseppe; Emanuelli, Giorgio

    2004-01-01

    In pancreatic acini, the G-protein-activated phosphoinositide 3-kinase-? (PI3K?) regulates several key pathological responses to cholecystokinin hyperstimulation in vitro. Thus, using mice lacking PI3K?, we studied the function of this enzyme in vivo in two different models of acute pancreatitis. The disease was induced by supramaximal concentrations of cerulein and by feeding mice a choline-deficient/ethionine-supplemented diet. Although the secretive function of isolated pancreatic acini was identical in mutant and control samples, in both models, genetic ablation of PI3K? significantly reduced the extent of acinar cell injury/necrosis. In agreement with a protective role of apoptosis in pancreatitis, PI3K?-deficient pancreata showed an increased number of apoptotic acinar cells, as determined by terminal dUTP nick-end labeling and caspase-3 activity. In addition, neutrophil infiltration within the pancreatic tissue was also reduced, suggesting a dual action of PI3K?, both in the triggering events within acinar cells and in the subsequent neutrophil recruitment and activation. Finally, the lethality of the choline-deficient/ethionine-supplemented diet-induced pancreatitis was significantly reduced in mice lacking PI3K?. Our results thus suggest that inhibition of PI3K? may be of therapeutic value in acute pancreatitis. PMID:15579443

  4. Phosphoinositide 3-kinase gamma participates in T cell receptor-induced T cell activation.

    PubMed

    Alcázar, Isabela; Marqués, Miriam; Kumar, Amit; Hirsch, Emilio; Wymann, Matthias; Carrera, Ana C; Barber, Domingo F

    2007-11-26

    Class I phosphoinositide 3-kinases (PI3Ks) constitute a family of enzymes that generates 3-phosphorylated polyphosphoinositides at the cell membrane after stimulation of protein tyrosine (Tyr) kinase-associated receptors or G protein-coupled receptors (GPCRs). The class I PI3Ks are divided into two types: class I(A) p85/p110 heterodimers, which are activated by Tyr kinases, and the class I(B) p110gamma isoform, which is activated by GPCR. Although the T cell receptor (TCR) is a protein Tyr kinase-associated receptor, p110gamma deletion affects TCR-induced T cell stimulation. We examined whether the TCR activates p110gamma, as well as the consequences of interfering with p110gamma expression or function for T cell activation. We found that after TCR ligation, p110gamma interacts with G alpha(q/11), lymphocyte-specific Tyr kinase, and zeta-associated protein. TCR stimulation activates p110gamma, which affects 3-phosphorylated polyphosphoinositide levels at the immunological synapse. We show that TCR-stimulated p110gamma controls RAS-related C3 botulinum substrate 1 activity, F-actin polarization, and the interaction between T cells and antigen-presenting cells, illustrating a crucial role for p110gamma in TCR-induced T cell activation. PMID:17998387

  5. Structural determinants of phosphoinositide 3-kinase inhibition by wortmannin, LY294002, quercetin, myricetin, and staurosporine.

    PubMed

    Walker, E H; Pacold, M E; Perisic, O; Stephens, L; Hawkins, P T; Wymann, M P; Williams, R L

    2000-10-01

    The specific phosphoinositide 3-kinase (PI3K) inhibitors wortmannin and LY294002 have been invaluable tools for elucidating the roles of these enzymes in signal transduction pathways. The X-ray crystallographic structures of PI3Kgamma bound to these lipid kinase inhibitors and to the broad-spectrum protein kinase inhibitors quercetin, myricetin, and staurosporine reveal how these compounds fit into the ATP binding pocket. With a nanomolar IC50, wortmannin most closely fits and fills the active site and induces a conformational change in the catalytic domain. Surprisingly, LY294002 and the lead compound on which it was designed, quercetin, as well as the closely related flavonoid myricetin bind PI3K in remarkably different orientations that are related to each other by 180 degrees rotations. Staurosporine/PI3K interactions are reminiscent of low-affinity protein kinase/staurosporine complexes. These results provide a rich basis for development of isoform-specific PI3K inhibitors with therapeutic potential. PMID:11090628

  6. Essential role of the p110beta subunit of phosphoinositide 3-OH kinase in male fertility.

    PubMed

    Ciraolo, Elisa; Morello, Fulvio; Hobbs, Robin M; Wolf, Frieder; Marone, Romina; Iezzi, Manuela; Lu, Xiaoyun; Mengozzi, Giulio; Altruda, Fiorella; Sorba, Giovanni; Guan, Kaomei; Pandolfi, Pier Paolo; Wymann, Matthias P; Hirsch, Emilio

    2010-03-01

    Phosphoinositide 3-kinases (PI3K) are key molecular players in male fertility. However, the specific roles of different p110 PI3K catalytic subunits within the spermatogenic lineage have not been characterized so far. Herein, we report that male mice expressing a catalytically inactive p110beta develop testicular hypotrophy and impaired spermatogenesis, leading to a phenotype of oligo-azoospermia and defective fertility. The examination of testes from p110beta-defective tubules demonstrates a widespread loss in spermatogenic cells, due to defective proliferation and survival of pre- and postmeiotic cells. In particular, p110beta is crucially needed in c-Kit-mediated spermatogonial expansion, as c-Kit-positive cells are lost in the adult testis and activation of Akt by SCF is blocked by a p110beta inhibitor. These data establish that activation of the p110beta PI3K isoform by c-Kit is required during spermatogenesis, thus opening the way to new treatments for c-Kit positive testicular cancers. PMID:20053680

  7. Phosphoinositide 3–kinase ? participates in T cell receptor–induced T cell activation

    PubMed Central

    Alcázar, Isabela; Marqués, Miriam; Kumar, Amit; Hirsch, Emilio; Wymann, Matthias; Carrera, Ana C.; Barber, Domingo F.

    2007-01-01

    Class I phosphoinositide 3–kinases (PI3Ks) constitute a family of enzymes that generates 3-phosphorylated polyphosphoinositides at the cell membrane after stimulation of protein tyrosine (Tyr) kinase–associated receptors or G protein–coupled receptors (GPCRs). The class I PI3Ks are divided into two types: class IA p85/p110 heterodimers, which are activated by Tyr kinases, and the class IB p110? isoform, which is activated by GPCR. Although the T cell receptor (TCR) is a protein Tyr kinase–associated receptor, p110? deletion affects TCR-induced T cell stimulation. We examined whether the TCR activates p110?, as well as the consequences of interfering with p110? expression or function for T cell activation. We found that after TCR ligation, p110? interacts with G?q/11, lymphocyte-specific Tyr kinase, and ?-associated protein. TCR stimulation activates p110?, which affects 3-phosphorylated polyphosphoinositide levels at the immunological synapse. We show that TCR-stimulated p110? controls RAS-related C3 botulinum substrate 1 activity, F-actin polarization, and the interaction between T cells and antigen-presenting cells, illustrating a crucial role for p110? in TCR-induced T cell activation. PMID:17998387

  8. Phosphoinositide 3-kinase gamma is an essential amplifier of mast cell function.

    PubMed

    Laffargue, Muriel; Calvez, Ronan; Finan, Peter; Trifilieff, Alexandre; Barbier, Maryse; Altruda, Fiorella; Hirsch, Emilio; Wymann, Matthias P

    2002-03-01

    Mast cells are key regulators in allergy and inflammation, and release histamine upon clustering of their IgE receptors. Here we demonstrate that murine mast cell responses are exacerbated in vitro and in vivo by autocrine signals through G protein-coupled receptors (GPCRs) and require functional phosphoinositide 3-kinase gamma (PI3Kgamma). Adenosine, acting through the A(3) adenosine receptor (A(3)AR) as well as other agonists of G(alphai)-coupled GPCRs, transiently increased PtdIns(3,4,5)P(3) exclusively via PI3Kgamma. PI3Kgamma-derived PtdIns(3,4,5)P(3) was instrumental for initiating a sustained influx of external Ca(2+) and degranulation. Mice lacking PI3Kgamma did not form edema after intradermal injection of adenosine and when challenged by passive systemic anaphylaxis. PI3Kgamma thus relays inflammatory signals through various G(i)-coupled receptors and is central to mast cell function. PMID:11911828

  9. Short-Form Ron Promotes Spontaneous Breast Cancer Metastasis through Interaction with Phosphoinositide 3-Kinase

    PubMed Central

    Liu, Xuemei; Zhao, Ling; DeRose, Yoko S.; Lin, Yi-Chun; Bieniasz, Magdalena; Eyob, Henok; Buys, Saundra S.; Neumayer, Leigh

    2011-01-01

    Receptor tyrosine kinases (RTKs) have been the subject of intense investigation due to their widespread deregulation in cancer and the prospect of developing targeted therapeutics against these proteins. The Ron RTK has been implicated in tumor aggressiveness and is a developing target for therapy, but its function in tumor progression and metastasis is not fully understood. We examined Ron activity in human breast cancers and found striking predominance of an activated Ron isoform known as short-form Ron (sfRon), whose function in breast tumors has not been explored. We found that sfRon plays a significant role in aggressiveness of breast cancer in vitro and in vivo. sfRon expression was sufficient to convert slow-growing, nonmetastatic tumors into rapidly growing tumors that spontaneously metastasized to liver and bones. Mechanistic studies revealed that sfRon promotes epithelial-mesenchymal transition, invasion, tumor growth, and metastasis through interaction with p85, the regulatory subunit of phosphoinositide 3-kinase (PI3K). Inhibition of PI3K activity, or introduction of a single mutation in the p85 docking site on sfRon, completely eliminated the ability of sfRon to promote tumor growth, invasion, and metastasis. These findings reveal sfRon as an important new player in breast cancer and validate Ron and PI3K as therapeutic targets in this disease. PMID:22207901

  10. Tyrosol Suppresses Allergic Inflammation by Inhibiting the Activation of Phosphoinositide 3-Kinase in Mast Cells

    PubMed Central

    Kim, Sung-Wan; Lee, Soyoung; Lee, Hyun-Shik; Park, Eui Kyun; Khang, Dongwoo; Kim, Sang-Hyun

    2015-01-01

    Allergic diseases such as atopic dermatitis, rhinitis, asthma, and anaphylaxis are attractive research areas. Tyrosol (2-(4-hydroxyphenyl)ethanol) is a polyphenolic compound with diverse biological activities. In this study, we investigated whether tyrosol has anti-allergic inflammatory effects. Ovalbumin-induced active systemic anaphylaxis and immunoglobulin E-mediated passive cutaneous anaphylaxis models were used for the immediate-type allergic responses. Oral administration of tyrosol reduced the allergic symptoms of hypothermia and pigmentation in both animal models. Mast cells that secrete allergic mediators are key regulators on allergic inflammation. Tyrosol dose-dependently decreased mast cell degranulation and expression of inflammatory cytokines. Intracellular calcium levels and activation of inhibitor of ?B kinase (IKK) regulate cytokine expression and degranulation. Tyrosol blocked calcium influx and phosphorylation of the IKK complex. To define the molecular target for tyrosol, various signaling proteins involved in mast cell activation such as Lyn, Syk, phosphoinositide 3-kinase (PI3K), and Akt were examined. Our results showed that PI3K could be a molecular target for tyrosol in mast cells. Taken together, these findings indicated that tyrosol has anti-allergic inflammatory effects by inhibiting the degranulation of mast cells and expression of inflammatory cytokines; these effects are mediated via PI3K. Therefore, we expect tyrosol become a potential therapeutic candidate for allergic inflammatory disorders. PMID:26068872

  11. Changing phosphoinositides "on the fly": how trafficking vesicles avoid an identity crisis.

    PubMed

    Botelho, Roberto J

    2009-10-01

    Joining an antagonistic phosphoinositide (PtdInsP) kinase and phosphatase into a single protein complex may regulate rapid and local PtdInsP changes. This may be important for processes such as membrane fission that require a specific PtdInsP and that are innately local and rapid. Such a complex could couple vesicle formation, with erasing of the identity of the donor organelle from the vesicle prior to its fusion with target organelles, thus preventing organelle identity intermixing. Coordinating signals are postulated to switch the relative activities of the kinase and phosphatase in a spatio-temporal manner that matches membrane fission events. The discovery of two such complexes supports this hypothesis. One regulates the interconversion of phosphatidylinositol and PtdIns(3)P by joining the Vps34 PtdIns 3-kinase and the myotubularin 3-phosphatases. The other regulates the interconversion between PtdIns(3)P and PtdIns(3,5)P(2) through the Fab1/PIKfyve kinase and the Fig4/mFig4 phosphatase. These lipids are essential components of the endosomal identity code. PMID:19708025

  12. RAS and RHO Families of GTPases Directly Regulate Distinct Phosphoinositide 3-Kinase Isoforms

    PubMed Central

    Fritsch, Ralph; de Krijger, Inge; Fritsch, Kornelia; George, Roger; Reason, Beth; Kumar, Madhu S.; Diefenbacher, Markus; Stamp, Gordon; Downward, Julian

    2013-01-01

    Summary RAS proteins are important direct activators of p110?, p110?, and p110? type I phosphoinositide 3-kinases (PI3Ks), interacting via an amino-terminal RAS-binding domain (RBD). Here, we investigate the regulation of the ubiquitous p110? isoform of PI3K, implicated in G-protein-coupled receptor (GPCR) signaling, PTEN-loss-driven cancers, and thrombocyte function. Unexpectedly, RAS is unable to interact with p110?, but instead RAC1 and CDC42 from the RHO subfamily of small GTPases bind and activate p110? via its RBD. In fibroblasts, GPCRs couple to PI3K through Dock180/Elmo1-mediated RAC activation and subsequent interaction with p110?. Cells from mice carrying mutations in the p110? RBD show reduced PI3K activity and defective chemotaxis, and these mice are resistant to experimental lung fibrosis. These findings revise our understanding of the regulation of type I PI3K by showing that both RAS and RHO family GTPases directly regulate distinct ubiquitous PI3K isoforms and that RAC activates p110? downstream of GPCRs. PMID:23706742

  13. Phosphoinositide hydrolysis by guanosine 5'-[gamma-thio]triphosphate-activated phospholipase C of turkey erythrocyte membranes.

    PubMed Central

    Harden, T K; Hawkins, P T; Stephens, L; Boyer, J L; Downes, C P

    1988-01-01

    Phosphatidylinositol (PtdIns), phosphatidylinositol 4-phosphate (PtdIns4P) and phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] of turkey erythrocytes were labelled by using either [32P]Pi or [3H]inositol. Although there was little basal release of inositol phosphates from membranes purified from labelled cells, in the presence of guanosine 5'-[gamma-thio]triphosphate (GTP[S]) the rate of accumulation of inositol bis-, tris- and tetrakis-phosphate (InsP2, InsP3 and InsP4) was increased 20-50-fold. The enhanced rate of accumulation of 3H-labelled inositol phosphates was linear for up to 20 min; owing to decreases in 32P specific radioactivity of phosphoinositides during incubation of membranes with unlabelled ATP, the accumulation of 32P-labelled inositol phosphates was linear for only 5 min. In the absence of ATP and a nucleotide-regenerating system, no InsP4 was formed, and the overall inositol phosphate response to GTP[S] was decreased. Analyses of phosphoinositides during incubation with ATP indicated that interconversions of PtdIns to PtdIns4P and PtdIns4P to PtdIns(4,5)P2 occurred to maintain PtdIns(4,5)P2 concentrations; GTP[S]-induced inositol phosphate formation was accompanied by a corresponding decrease in 32P- and 3H-labelled PtdIns, PtdIns4P and PtdIns(4,5)P2. In the absence of ATP, only GTP[S]-induced decreases in PtdIns(4,5)P2 occurred. Since inositol monophosphate was not formed under any condition, PtdIns is not a substrate for the phospholipase C. The production of InsP2 was decreased markedly, but not blocked, under conditions where Ins(1,4,5)P3 5-phosphomonoesterase activity in the preparation was inhibited. Thus the predominant substrate of the GTP[S]-activated phospholipase C of turkey erythrocyte membranes is PtdIns(4,5)P2. Ins(1,4,5)P3 was the major product of this reaction; only a small amount of Ins(1:2-cyclic, 4,5)P3 was released. The effects of ATP on inositol phosphate formation apparently involve the contributions of two phenomena. First, the P2-receptor agonist 2-methylthioadenosine triphosphate (2MeSATP) greatly increased inositol phosphate formation and decreased [3H]PtdIns4P and [3H]PtdIns(4,5)P2 in the presence of a low (0.1 microM) concentration of GTP[S]. ATP over the concentration range 0-100 microM produced effects in the presence of 0.1 microM-GTP[S] essentially identical with those observed with 2MeSATP, suggesting that the effects of low concentrations of ATP are also explained by a stimulation of P2-receptors. Higher concentrations of ATP also increase inositol phosphate formation, apparently by supporting the synthesis of substrate phospholipids.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:2843174

  14. Molecular mechanism of membrane targeting by the GRP1 PH domain,boxs

    Microsoft Academic Search

    Ju He; Rachel M. Haney; Mohsin Vora; Vladislav V. Verkhusha; Robert V. Stahelin; Tatiana G. Kutateladze

    2008-01-01

    Abstract,The general,receptor,for phosphoinositides iso- form,1 (GRP1) is recruited,to the plasma,membrane,in re- sponse,to activation,of phosphoinositide,3-kinases and accumulation of phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P3]. GRP1?s pleckstrin homology (PH) do- main recognizes PtdIns(3,4,5)P3 with high specificity and af- finity, however, the precise mechanism of its association with membranes remains unclear. Here, we detail the mo- lecular basis of membrane,anchoring,by the GRP1 PH do- main. Our

  15. Triggering actin polymerization in Xenopus egg extracts from phosphoinositide-containing lipid bilayers.

    PubMed

    Walrant, Astrid; Saxton, Daniel S; Correia, Guilherme Pereira; Gallop, Jennifer L

    2015-01-01

    Xenopus egg extracts are a powerful tool to reconstitute complex cell biological processes using a cell-free strategy. When used in conjunction with liposomes and supported lipid bilayers, they can recapitulate the biochemical activities occurring at the cytosol/plasma membrane interface of the cell that underlie remodeling of the actin cytoskeleton. We use these in vitro systems to elucidate how membranes and proteins collaborate to make the appropriate actin structure at a given time and place. We have recently broadened the types of membrane substrate used, and also optimized protocols for preparation of Xenopus egg extracts for actin assembly assays from membranes. Tuning the lipid composition and curvature appropriately demands an appreciation of the native phospholipid and curvature environments that can form transiently in cells. Supported lipid bilayers on glass coverslips that contain phosphatidylserine and phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) make actin bundles termed filopodia-like structures that contain fascin and have vasodilator-stimulated phosphoprotein (VASP) at their growing tips, indicating that these resemble filopodia growing from the plasma membrane. The combination of PI(4,5)P2 and phosphatidylinositol 3-phosphate in curved liposomes or supported bilayers on glass nanospheres uses Snx9, Cdc42, N-WASP (neuronal-Wiskott-Aldrich syndrome protein), and Arp2/3 complex for actin polymerization suggesting that this membrane may mimic the progression from plasma membrane to endosomes. Here we describe how to prepare high-speed supernatant frog egg extracts and phosphoinositide-containing liposomes and supported lipid bilayers that can assemble actin structures. We also describe the methods we use to assay actin polymerization using microscopy and spectrofluorometry and our protocol for immunodepleting specific proteins from extracts. PMID:25997346

  16. Endosomal Maturation, Rab7 GTPase and Phosphoinositides in African Swine Fever Virus Entry

    PubMed Central

    Cuesta-Geijo, Miguel A.; Galindo, Inmaculada; Hernáez, Bruno; Quetglas, Jose Ignacio; Dalmau-Mena, Inmaculada; Alonso, Covadonga

    2012-01-01

    Here we analyzed the dependence of African swine fever virus (ASFV) infection on the integrity of the endosomal pathway. Using confocal immunofluorescence with antibodies against viral capsid proteins, we found colocalization of incoming viral particles with early endosomes (EE) during the first minutes of infection. Conversely, viral capsid protein was not detected in acidic late endosomal compartments, multivesicular bodies (MVBs), late endosomes (LEs) or lysosomes (LY). Using an antibody against a viral inner core protein, we found colocalization of viral cores with late compartments from 30 to 60 minutes postinfection. The absence of capsid protein staining in LEs and LYs suggested that virus desencapsidation would take place at the acid pH of these organelles. In fact, inhibitors of intraluminal acidification of endosomes caused retention of viral capsid staining virions in Rab7 expressing endosomes and more importantly, severely impaired subsequent viral protein production. Endosomal acidification in the first hour after virus entry was essential for successful infection but not thereafter. In addition, altering the balance of phosphoinositides (PIs) which are responsible of the maintenance of the endocytic pathway impaired ASFV infection. Early infection steps were dependent on the production of phosphatidylinositol 3-phosphate (PtdIns3P) which is involved in EE maturation and multivesicular body (MVB) biogenesis and on the interconversion of PtdIns3P to phosphatidylinositol 3, 5-biphosphate (PtdIns(3,5)P2). Likewise, GTPase Rab7 activity should remain intact, as well as processes related to LE compartment physiology, which are crucial during early infection. Our data demonstrate that the EE and LE compartments and the integrity of the endosomal maturation pathway orchestrated by Rab proteins and PIs play a central role during early stages of ASFV infection. PMID:23133661

  17. Characterization of human recombinant somatostatin sst5 receptors mediating activation of phosphoinositide metabolism

    PubMed Central

    Wilkinson, Graeme F; Feniuk, Wasyl; Humphrey, Patrick P A

    1997-01-01

    We have functionally characterized the human recombinant somatostatin (SRIF) sst5 receptor in Chinese hamster ovary-K1 (CHOsst5) cells by measuring total [3H]-inositol phosphate ([3H]-InsPx) accumulation, in the presence of 10?mM LiCl, in cells labelled with [3H]-myo-inositol. In CHOsst5 cells, SRIF, SRIF-28 and the cyclic hexapeptide, L-362,855, produced time- and concentration-related increases in [3H]-InsPx accumulation, with similar potency (pEC50 values of 6.5, 6.8 and 7.2, respectively). L-362,855 behaved as a partial agonist, producing approximately 30% of the SRIF maximum response. The other peptide analogues of SRIF, BIM-23027 and BIM-23056, were inactive as agonists. Increasing concentrations of L-362,855 increased [3H]-InsPx accumulation and simultaneously produced rightward shifts of SRIF concentration-effect curves, with an estimated pKp value of 7.6, confirming that it was acting as a partial agonist. BIM-23056, but not BIM-23027, potently antagonized SRIF-induced [3H]-InsPx accumulation, with an estimated pKB value of 7.4. BIM-23056 did not antagonize [3H]-InsPx accumulation induced by uridine 5?-triphosphate (UTP). SRIF- but not UTP-induced [3H]-InsPx accumulation was inhibited by increasing concentrations of pertussis toxin (0.01–100?ng?ml?1), indicating the involvement of pertussis toxin-sensitive G-proteins. These findings show that the human recombinant sst5 receptor, when stably expressed in CHO-K1 cells, is able to mediate activation of phosphoinositide metabolism in a pertussis toxin-sensitive manner. In this system L-362,855 behaved as a partial agonist while BIM-23056 was a specific antagonist. These agents should provide useful tools for functionally characterizing endogenous SRIF receptors. PMID:9146892

  18. Targeting Phosphoinositide-3-Kinase-? with Theophylline Reverses Corticosteroid Insensitivity in Chronic Obstructive Pulmonary Disease

    PubMed Central

    To, Yasuo; Ito, Kazuhiro; Kizawa, Yasuo; Failla, Marco; Ito, Misako; Kusama, Tadashi; Elliott, W. Mark; Hogg, James C.; Adcock, Ian M.; Barnes, Peter J.

    2010-01-01

    Rationale: Patients with chronic obstructive pulmonary disease (COPD) show a poor response to corticosteroids. This has been linked to a reduction of histone deacetylase-2 as a result of oxidative stress and is reversed by theophylline. Objectives: To determine the role of phosphoinositide-3-kinase-delta (PI3K-?) on the development of corticosteroid insensitivity in COPD and under oxidative stress, and as a target for theophylline. Methods: Corticosteroid sensitivity was determined as the 50% inhibitory concentration of dexamethasone on tumor necrosis factor-?–induced interleukin-8 release in peripheral blood mononuclear cells from patients with COPD (n = 17) and compared with that of nonsmoking (n = 8) and smoking (n = 7) control subjects. The effect of theophylline and a selective PI3K-? inhibitor (IC87114) on restoration of corticosteroid sensitivity was confirmed in cigarette smoke–exposed mice. Measurements and Main Results: Peripheral blood mononuclear cells of COPD (50% inhibitory concentration of dexamethasone: 156.8 ± 32.6 nM) were less corticosteroid sensitive than those of nonsmoking (41.2 ± 10.5 nM; P = 0.018) and smoking control subjects (47.5 ± 19.6 nM; P = 0.031). Corticosteroid insensitivity and reduced histone deacetylase-2 activity after oxidative stress were reversed by a non-selective PI3K inhibitor (LY294002) and low concentrations of theophylline. Theophylline was a potent selective inhibitor of oxidant-activated PI3K-?, which was up-regulated in peripheral lung tissue of patients with COPD. Furthermore, cells with knock-down of PI3K-? failed to develop corticosteroid insensitivity with oxidative stress. Both theophylline and IC87114, combined with dexamethasone, inhibited corticosteroid-insensitive lung inflammation in cigarette–smoke-exposed mice in vivo. Conclusions: Inhibition of oxidative stress dependent PI3K-? activation by a selective inhibitor or theophylline provides a novel approach to reversing corticosteroid insensitivity in COPD. PMID:20224070

  19. Changes in phosphoinositide turnover, Ca sup 2+ mobilization, and protein phosphorylation in platelets from NIDDM patients

    SciTech Connect

    Ishii, H.; Umeda, F.; Hashimoto, T.; Nawata, H. (Kyushu Univ., Fukuoka (Japan))

    1990-12-01

    Enhanced platelet functions have been demonstrated in patients with non-insulin-dependent diabetes mellitus (NIDDM). This study evaluated abnormalities in platelet signal transduction in diabetic patients, including turnover of phosphoinositides, mobilization of intracellular Ca2+, and phosphorylation of 20,000- and 47,000-Mr proteins (P20 and P47). Washed platelets were obtained from 6 patients with NIDDM whose platelet aggregation rates were abnormally elevated (DM-A group), 11 NIDDM patients with normal platelet aggregation rates (DM-B group), and 8 age-matched healthy control subjects. The mass and specific radioactivity of phosphatidylinositol 4,5-bisphosphate (PIP2), phosphatidylinositol 4-phosphate (PIP), phosphatidylinositol (PI), and phosphatidic acid (PA) in 32P-labeled platelets were not different among the three groups. Hydrolysis of PIP2, PIP, and PI; accumulation of PA; and phosphorylation of P20 in platelets stimulated by 0.05 U/ml thrombin were significantly increased in the DM-A group compared with the control or DM-B group. There was no difference in P47 phosphorylation among the three groups. On the contrary, P20 and P47 phosphorylation induced by 50 nM of 12-O-tetradecanoylphorbol-13-acetate, an activator of protein kinase C, was significantly decreased in the DM-A group. Additionally, the intracellular free Ca2+ concentration (( Ca2+)i) was measured with the fluorescent Ca2+ indicator fura 2. Although the basal (Ca2+)i value was similar in the three groups, the rise in (Ca2+)i induced by 0.05 U/ml thrombin in the presence and the absence of extracellular Ca2+ was significantly higher in the DM-A group than the other groups.

  20. Role of phosphoinositide 3-kinase in the pathogenesis of acute pancreatitis

    PubMed Central

    Lupia, Enrico; Pigozzi, Luca; Goffi, Alberto; Hirsch, Emilio; Montrucchio, Giuseppe

    2014-01-01

    A large body of experimental and clinical data supports the notion that inflammation in acute pancreatitis has a crucial role in the pathogenesis of local and systemic damage and is a major determinant of clinical severity. Thus, research has recently focused on molecules that can regulate the inflammatory processes, such as phosphoinositide 3-kinases (PI3Ks), a family of lipid and protein kinases involved in intracellular signal transduction. Studies using genetic ablation or pharmacologic inhibitors of different PI3K isoforms, in particular the class I PI3K? and PI3K?, have contributed to a greater understanding of the roles of these kinases in the modulation of inflammatory and immune responses. Recent data suggest that PI3Ks are also involved in the pathogenesis of acute pancreatitis. Activation of the PI3K signaling pathway, and in particular of the class IB PI3K? isoform, has a significant role in those events which are necessary for the initiation of acute pancreatic injury, namely calcium signaling alteration, trypsinogen activation, and nuclear factor-?B transcription. Moreover, PI3K? is instrumental in modulating acinar cell apoptosis, and regulating local neutrophil infiltration and systemic inflammatory responses during the course of experimental acute pancreatitis. The availability of PI3K inhibitors selective for specific isoforms may provide new valuable therapeutic strategies to improve the clinical course of this disease. This article presents a brief summary of PI3K structure and function, and highlights recent advances that implicate PI3Ks in the pathogenesis of acute pancreatitis. PMID:25386068

  1. Phosphoinositide 3-kinase gamma: a key modulator in inflammation and allergy.

    PubMed

    Wymann, M P; Björklöf, K; Calvez, R; Finan, P; Thomast, M; Trifilieff, A; Barbier, M; Altruda, F; Hirsch, E; Laffargue, M

    2003-02-01

    Chronic inflammation and allergy involve the activation of tissue-resident cells and, later on, the invasion of effector cells. We have previously shown that the loss of phosphoinositide 3-kinase (PI3K) gamma impairs chemokine-dependent migration of neutrophils and macrophages both in vitro and in vivo. On the other hand, PI3K gamma is not required either during phagocytic processes or in the activation of bactericidal activities like granule secretion and particle-mediated respiratory burst in neutrophils. Tissue mast cells are key regulators in allergy and inflammation and release histamine upon clustering of their IgE receptors. We have demonstrated that murine mast cell responses are exacerbated in vitro and in vivo by autocrine signals, and require functional PI3K gamma. Adenosine, acting through the A(3) adenosine receptor, as well as other agonists of G(alpha i)-coupled receptors, transiently increased PtdIns(3,4,5) P (3) exclusively via PI3K gamma. PI3K gamma-derived PtdIns(3,4,5) P (3) was instrumental for initiation of a sustained influx of external Ca(2+) and degranulation. Mice that lacked PI3K gamma did not form oedema when challenged by passive systemic anaphylaxis. PI3K gamma thus relays inflammatory signals through various GPCRs, and is thus central to mast cell function. Taken together, this suggests that pharmaceutical targeting of PI3K gamma might alleviate inflammation at both early and late stages of the allergic response. PMID:12546701

  2. Targeting melanoma with dual phosphoinositide 3-kinase/mammalian target of rapamycin inhibitors.

    PubMed

    Marone, Romina; Erhart, Dominik; Mertz, Ann C; Bohnacker, Thomas; Schnell, Christian; Cmiljanovic, Vladimir; Stauffer, Frédéric; Garcia-Echeverria, Carlos; Giese, Bernd; Maira, Sauveur-Michel; Wymann, Matthias P

    2009-04-01

    Phosphoinositide 3-kinase (PI3K)/protein kinase B/Akt and Ras/mitogen-activated protein kinase pathways are often constitutively activated in melanoma and have thus been considered as promising drug targets. Exposure of melanoma cells to NVP-BAG956, NVP-BBD130, and NVP-BEZ235, a series of novel, potent, and stable dual PI3K/mammalian target of rapamycin (mTOR) inhibitors, resulted in complete G1 growth arrest, reduction of cyclin D1, and increased levels of p27(KIP1), but negligible apoptosis. In contrast, treatment of melanoma with the pan-class I PI3K inhibitor ZSTK474 or the mTORC1 inhibitor rapamycin resulted only in minor reduction of cell proliferation. In a syngeneic B16 mouse melanoma tumor model, orally administered NVP-BBD130 and NVP-BEZ235 efficiently attenuated tumor growth at primary and lymph node metastatic sites with no obvious toxicity. Metastatic melanoma in inhibitor-treated mice displayed reduced numbers of proliferating and significantly smaller tumor cells. In addition, neovascularization was blocked and tumoral necrosis increased when compared with vehicle-treated mice. In conclusion, compounds targeting PI3K and mTOR simultaneously were advantageous to attenuate melanoma growth and they develop their potential by targeting tumor growth directly, and indirectly via their interference with angiogenesis. Based on the above results, NVP-BEZ235, which has entered phase I/II clinical trials in patients with advanced solid tumors, has a potential in metastatic melanoma therapy. PMID:19372588

  3. IQGAP Proteins Reveal an Atypical Phosphoinositide (aPI) Binding Domain with a Pseudo C2 Domain Fold*

    PubMed Central

    Dixon, Miles J.; Gray, Alexander; Schenning, Martijn; Agacan, Mark; Tempel, Wolfram; Tong, Yufeng; Nedyalkova, Lyudmila; Park, Hee-Won; Leslie, Nicholas R.; van Aalten, Daan M. F.; Downes, C. Peter; Batty, Ian H.

    2012-01-01

    Class I phosphoinositide (PI) 3-kinases act through effector proteins whose 3-PI selectivity is mediated by a limited repertoire of structurally defined, lipid recognition domains. We describe here the lipid preferences and crystal structure of a new class of PI binding modules exemplified by select IQGAPs (IQ motif containing GTPase-activating proteins) known to coordinate cellular signaling events and cytoskeletal dynamics. This module is defined by a C-terminal 105–107 amino acid region of which IQGAP1 and -2, but not IQGAP3, binds preferentially to phosphatidylinositol 3,4,5-trisphosphate (PtdInsP3). The binding affinity for PtdInsP3, together with other, secondary target-recognition characteristics, are comparable with those of the pleckstrin homology domain of cytohesin-3 (general receptor for phosphoinositides 1), an established PtdInsP3 effector protein. Importantly, the IQGAP1 C-terminal domain and the cytohesin-3 pleckstrin homology domain, each tagged with enhanced green fluorescent protein, were both re-localized from the cytosol to the cell periphery following the activation of PI 3-kinase in Swiss 3T3 fibroblasts, consistent with their common, selective recognition of endogenous 3-PI(s). The crystal structure of the C-terminal IQGAP2 PI binding module reveals unexpected topological similarity to an integral fold of C2 domains, including a putative basic binding pocket. We propose that this module integrates select IQGAP proteins with PI 3-kinase signaling and constitutes a novel, atypical phosphoinositide binding domain that may represent the first of a larger group, each perhaps structurally unique but collectively dissimilar from the known PI recognition modules. PMID:22493426

  4. Phosphoinositides Are Involved in Control of the Glucose-Dependent Growth Resumption That Follows the Transition Phase in Streptomyces lividans?

    PubMed Central

    Chouayekh, H.; Nothaft, H.; Delaunay, S.; Linder, M.; Payrastre, B.; Seghezzi, N.; Titgemeyer, F.; Virolle, M. J.

    2007-01-01

    The interruption of the sblA gene of Streptomyces lividans was previously shown to lead to relief of glucose repression of the normally strongly glucose-repressed ?-amylase gene. In addition to this relief, an early entry into stationary phase was observed when cells were grown in a minimal medium containing glucose as the main carbon source. In this study, we established that this mutant does not resume growth after the transition phase when cultured in the complex glucose-rich liquid medium R2YE and sporulates much earlier than the wild-type strain when plated on solid R2YE. These phenotypic differences, which were abolished when glucose was omitted from the R2YE medium, correlated with a reduced glucose uptake ability of the sblA mutant strain. sblA was shown to encode a bifunctional enzyme possessing phospholipase C-like and phosphoinositide phosphatase activities. The cleavage of phosphoinositides by SblA seems necessary to trigger the glucose-dependent renewed growth that follows the transition phase. The transient expression of sblA that takes place just before the transition phase is consistent with a regulatory role for this gene during the late stages of growth. The tight temporal control of sblA expression was shown to depend on two operator sites. One, located just upstream of the ?35 promoter region, likely constitutes a repressor binding site. The other, located 170 bp downstream of the GTG sblA translational start codon, may be involved in the regulation of the degradation of the sblA transcript. This study suggests that phosphoinositides constitute important regulatory molecules in Streptomyces, as they do in eukaryotes. PMID:17122350

  5. IQGAP Proteins Reveal an Atypical Phosphoinositide (aPI) Binding Domain with a Pseudo C2 Domain Fold

    SciTech Connect

    Dixon, Miles J.; Gray, Alexander; Schenning, Martijn; Agacan, Mark; Tempel, Wolfram; Tong, Yufeng; Nedyalkova, Lyudmila; Park, Hee-Won; Leslie, Nicholas R.; van Aalten, Daan M.F.; Downes, C. Peter; Batty, Ian H. (Toronto); (Dundee)

    2012-10-16

    Class I phosphoinositide (PI) 3-kinases act through effector proteins whose 3-PI selectivity is mediated by a limited repertoire of structurally defined, lipid recognition domains. We describe here the lipid preferences and crystal structure of a new class of PI binding modules exemplified by select IQGAPs (IQ motif containing GTPase-activating proteins) known to coordinate cellular signaling events and cytoskeletal dynamics. This module is defined by a C-terminal 105-107 amino acid region of which IQGAP1 and -2, but not IQGAP3, binds preferentially to phosphatidylinositol 3,4,5-trisphosphate (PtdInsP3). The binding affinity for PtdInsP3, together with other, secondary target-recognition characteristics, are comparable with those of the pleckstrin homology domain of cytohesin-3 (general receptor for phosphoinositides 1), an established PtdInsP3 effector protein. Importantly, the IQGAP1 C-terminal domain and the cytohesin-3 pleckstrin homology domain, each tagged with enhanced green fluorescent protein, were both re-localized from the cytosol to the cell periphery following the activation of PI 3-kinase in Swiss 3T3 fibroblasts, consistent with their common, selective recognition of endogenous 3-PI(s). The crystal structure of the C-terminal IQGAP2 PI binding module reveals unexpected topological similarity to an integral fold of C2 domains, including a putative basic binding pocket. We propose that this module integrates select IQGAP proteins with PI 3-kinase signaling and constitutes a novel, atypical phosphoinositide binding domain that may represent the first of a larger group, each perhaps structurally unique but collectively dissimilar from the known PI recognition modules.

  6. NHE3 Activity Is Dependent on Direct Phosphoinositide Binding at the N Terminus of Its Intracellular Cytosolic Region*

    PubMed Central

    Mohan, Sachin; Tse, Chung Ming; Gabelli, Sandra B.; Sarker, Rafiquel; Cha, Boyoung; Fahie, Kamau; Nadella, Mythili; Zachos, Nicholas C.; Tu-Sekine, Becky; Raben, Daniel; Amzel, L. Mario; Donowitz, Mark

    2010-01-01

    The small intestinal BB Na+/H+ antiporter NHE3 accounts for the majority of intestinal sodium and water absorption. It is highly regulated with both postprandial inhibition and stimulation sequentially occurring. Phosphatidylinositide 4,5-bisphosphate (PI(4,5)P2) and phosphatidylinositide 3,4,5-trisphosphate (PI(3,4,5)P3) binding is involved with regulation of multiple transporters. We tested the hypothesis that phosphoinositides bind NHE3 under basal conditions and are necessary for its acute regulation. His6 proteins were made from the NHE3 C-terminal region divided into four parts as follows: F1 (amino acids 475–589), F2 (amino acids 590–667), F3 (amino acids 668–747), and F4 (amino acids 748–832) and purified by a nickel column. Mutations were made in the F1 region of NHE3 and cloned in pet30a and pcDNA3.1 vectors. PI(4,5)P2 and PI(3,4,5)P3 bound only to the NHE3 F1 fusion protein (amino acids 475–589) on liposomal pulldown assays. Mutations were made in the putative lipid binding region of the F1 domain and studied for alterations in lipid binding and Na+/H+ exchange as follows: Y501A/R503A/K505A; F509A/R511A/R512A; R511L/R512L; R520/FR527F; and R551L/R552L. Our results indicate the following. 1) The F1 domain of the NHE3 C terminus has phosphoinositide binding regions. 2) Mutations of these regions alter PI(4,5)P2 and PI(3,4,5)P3 binding and basal NHE3 activity. 3) The magnitude of serum stimulation of NHE3 correlates with PI(4,5)P2 and PI(3,4,5)P3 binding of NHE3. 4) Wortmannin inhibition of PI3K did not correlate with PI(4,5)P2 or PI(3,4,5)P3 binding of NHE3. Two functionally distinct phosphoinositide binding regions (Tyr501–Arg512 and Arg520–Arg552) are present in the NHE3 F1 domain; both regions are important for serum stimulation, but they display differences in phosphoinositide binding, and the latter but not the former alters NHE3 surface expression. PMID:20736165

  7. Effect of aging on alpha-1 adrenergic stimulation of phosphoinositide hydrolysis in various regions of rat brain

    SciTech Connect

    Burnett, D.M.; Bowyer, J.F.; Masserano, J.M.; Zahniser, N.R. (Univ. of Colorado Health Sciences Center, Denver (USA))

    1990-12-01

    The effects of aging were examined on the ability of alpha-1 adrenergic receptor agonists to stimulate phosphoinositide hydrolysis in three brain regions. Tissue minces of thalamus, cerebral cortex and hippocampus from 3-, 18- and 28-month-old male Fischer 344 rats were prelabeled with ({sup 3}H)myoinositol. Exposure of these prelabeled minces to phenylephrine and (-)-norepinephrine revealed that accumulation of ({sup 3}H)inositol phosphates was selectively reduced by 20 to 30% in the thalamus and cerebral cortex of the oldest age group. Analysis of concentration-response and competition binding curves indicated that this decrease was due to diminished agonist efficacy rather than diminished receptor affinity. The reduction in responsiveness to phenylephrine and (-)-norepinephrine in the cerebral cortex and the lack of any changes in the hippocampus parallel previously reported changes in the density of alpha-1 adrenergic receptors with aging. These data indicate that the ability of alpha-1 adrenergic receptor agonists to stimulate phosphoinositide hydrolysis is reduced in some, but not all, brain regions of aged Fischer 344 rats.

  8. Phosphoinositide phosphatase SHIP-1 regulates apoptosis induced by edelfosine, Fas ligation and DNA damage in mouse lymphoma cells.

    PubMed

    Alderliesten, Maaike C; Klarenbeek, Jeffrey B; van der Luit, Arnold H; van Lummel, Menno; Jones, David R; Zerp, Shuraila; Divecha, Nullin; Verheij, Marcel; van Blitterswijk, Wim J

    2011-11-15

    S49 mouse lymphoma cells undergo apoptosis in response to the ALP (alkyl-lysophospholipid) edelfosine (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine), FasL (Fas ligand) and DNA damage. S49 cells made resistant to ALP (S49(AR)) are defective in sphingomyelin synthesis and ALP uptake, and also have acquired resistance to FasL and DNA damage. However, these cells can be re-sensitized following prolonged culturing in the absence of ALP. The resistant cells show sustained ERK (extracellular-signal-regulated kinase)/Akt activity, consistent with enhanced survival signalling. In search of a common mediator of the observed cross-resistance, we found that S49(AR) cells lacked the PtdIns(3,4,5)P(3) phosphatase SHIP-1 [SH2 (Src homology 2)-domain-containing inositol phosphatase 1], a known regulator of the Akt survival pathway. Re-sensitization of the S49(AR) cells restored SHIP-1 expression as well as phosphoinositide and sphingomyelin levels. Knockdown of SHIP-1 mimicked the S49(AR) phenotype in terms of apoptosis cross-resistance, sphingomyelin deficiency and altered phosphoinositide levels. Collectively, the results of the present study suggest that SHIP-1 collaborates with sphingomyelin synthase to regulate lymphoma cell death irrespective of the nature of the apoptotic stimulus. PMID:21793801

  9. Adenosine triphosphate stimulates phosphoinositide metabolism, mobilizes intracellular calcium, and inhibits terminal differentiation of human epidermal keratinocytes.

    PubMed Central

    Pillai, S; Bikle, D D

    1992-01-01

    During wound healing, release of ATP from platelets potentially exposes the epidermis to concentrations of ATP known to alter cellular functions mediated via changes in inositol trisphosphate (IP3) and intracellular calcium (Cai) levels. Therefore, we determined whether keratinocytes respond to ATP with a rise in Cai and IP3 and whether such increases are accompanied by a change in their proliferation and differentiation. Changes in Cai were measured in Indo-1-loaded neonatal human foreskin keratinocytes after stimulation with extracellular ATP. Extracellular ATP evoked a transient and acute increase in Cai of keratinocytes both in the presence and in the absence of extracellular calcium. ATP also induced the phosphoinositide turnover of keratinocytes, consistent with its effect in releasing calcium from intracellular sources. ATP did not permeabilize keratinocytes, nor did it promote Ca influx into the cells. The half-maximal effect of ATP was at 10 microM, and saturation was observed at 30-100 microM. UTP, ITP, and ATP gamma S were as effective as ATP in releasing Cai from intracellular stores and competed with ATP for their response, whereas AMP and adenosine were ineffective, suggesting the specificity of P2 purinergic receptors in mediating the ATP response in keratinocytes. Single cell measurements revealed heterogeneity in the calcium response to ATP. This heterogeneity did not appear to be due to differences in the initial Cai response but to subsequent removal of increased Cai by these cells. ATP inhibited terminal differentiation of keratinocytes as measured by [35S]methionine incorporation into cornified envelopes and modestly stimulated incorporation of [3H]thymidine into DNA. Chelation of Cai by bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid reduced basal Cai, blocked the Cai response to ATP, inhibited the basal rate of DNA synthesis, and blocked the ATP-induced increase in DNA synthesis. We conclude that extracellular ATP may be an important physiological regulator of epidermal growth and differentiation acting via IP3 and Cai. Images PMID:1321844

  10. Pharmacologic Profiling of Phosphoinositide 3-Kinase Inhibitors as Mitigators of Ionizing Radiation–Induced Cell Death

    PubMed Central

    Sharlow, Elizabeth R.; Epperly, Michael W.; Lira, Ana; Leimgruber, Stephanie; Skoda, Erin M.; Wipf, Peter; Greenberger, Joel S.

    2013-01-01

    Ionizing radiation (IR) induces genotoxic stress that triggers adaptive cellular responses, such as activation of the phosphoinositide 3-kinase (PI3K)/Akt signaling cascade. Pluripotent cells are the most important population affected by IR because they are required for cellular replenishment. Despite the clear danger to large population centers, we still lack safe and effective therapies to abrogate the life-threatening effects of any accidental or intentional IR exposure. Therefore, we computationally analyzed the chemical structural similarity of previously published small molecules that, when given after IR, mitigate cell death and found a chemical cluster that was populated with PI3K inhibitors. Subsequently, we evaluated structurally diverse PI3K inhibitors. It is remarkable that 9 of 14 PI3K inhibitors mitigated ?IR-induced death in pluripotent NCCIT cells as measured by caspase 3/7 activation. A single intraperitoneal dose of LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one], administered to mice at 4 or 24 hours, or PX-867 [(4S,4aR,5R,6aS,9aR,Z)-11-hydroxy-4-(methoxymethyl)-4a,6a-dimethyl-2,7,10-trioxo-1-(pyrrolidin-1-ylmethylene)-1,2,4,4a,5,6,6a,7,8,9,9a,10-dodecahydroindeno[4,5-H]isochromen-5-yl acetate (CID24798773)], administered 4 hours after a lethal dose of ?IR, statistically significantly (P < 0.02) enhanced in vivo survival. Because cell cycle checkpoints are important regulators of cell survival after IR, we examined cell cycle distribution in NCCIT cells after ?IR and PI3K inhibitor treatment. LY294002 and PX-867 treatment of nonirradiated cells produced a marked decrease in S phase cells with a concomitant increase in the G1 population. In irradiated cells, LY294002 and PX-867 treatment also decreased S phase and increased the G1 and G2 populations. Treatment with LY294002 or PX-867 decreased ?IR-induced DNA damage as measured by ?H2AX, suggesting reduced DNA damage. These results indicate pharmacologic inhibition of PI3K after IR abrogated cell death. PMID:24068833

  11. Tanshinone IIA induces mitochondria dependent apoptosis in prostate cancer cells in association with an inhibition of phosphoinositide 3-kinase/AKT pathway.

    PubMed

    Won, Suk-Hyun; Lee, Hyo-Jeong; Jeong, Soo-Jin; Lee, Hyo-Jung; Lee, Eun-Ok; Jung, Deok-Beom; Shin, Ji-Min; Kwon, Tae-Rin; Yun, Sun-Mi; Lee, Min-Ho; Choi, Seung-Hoon; Lü, Junxuan; Kim, Sung-Hoon

    2010-01-01

    Tanshinone IIA (Tan IIA; 14,16-epoxy-20-nor-5(10),6,8,13,15-abietapentaene-11,12-dione), a phytochemical derived from the roots of Salvia miltiorrhiza BUNGE, has been reported to posses anti-angiogenic, anti-oxidant, anti-inflammatory and apoptotic activities. However, the cancer growth inhibitory/cytocidal effects and molecular mechanisms in prostate cancer cells have not been well studied. In the present study, we demonstrate that Tan IIA significantly decreased the viable cell number of LNCaP (phosphate and tensin homolog (PTEN) mutant, high AKT, wild type p53) prostate cancer cells more sensitively than against the PC-3 (PTEN null, high AKT, p53 null) prostate cancer cells. Tan IIA significantly increased TdT-mediated dUTP nick-end labeling (TUNEL) positive index and sub-G1 DNA contents of treated cells, consistent with apoptosis. Tan IIA treatment led to cleavage activation of pro-caspases-9 and 3, but not pro-caspase-8, and cleavage of poly (ADP ribose) polymerase (PARP), a caspase-3 substrate. Additionally, Tan IIA treatment induced cytochrome c release from the mitochondria into the cytosol and reduced mitochondrial membrane potential and suppressed the expression of mitochondria protective Bcl-2 family protein Mcl-1(L). Tan IIA reduced the expression of phosphoinositide 3-kinase (PI3K) p85 subunit, and the phosphorylation of AKT and mammalian target of rapamycin (mTOR) in a concentration-dependent manner. Moreover, the combination of Tan IIA and LY294002, a specific PI3K inhibitor, enhanced PARP cleavage of LNCaP and PC-3, but not in MDA-MB-231 breast cancer cells which do not contain detectable active AKT. The findings suggest that Tan IIA-induced apoptosis involves mitochondria intrinsic caspase activation cascade and an inhibition of the PI3K/AKT survival pathway. PMID:21048307

  12. The Forkhead Transcription Factor FOXO3a Increases Phosphoinositide-3 Kinase/Akt Activity in Drug-Resistant Leukemic Cells through Induction of PIK3CA Expression?

    PubMed Central

    Hui, Rosaline C.-Y.; Gomes, Ana R.; Constantinidou, Demetra; Costa, Joana R.; Karadedou, Christina T.; Fernandez de Mattos, Silvia; Wymann, Matthias P.; Brosens, Jan J.; Schulze, Almut; Lam, Eric W.-F.

    2008-01-01

    The phosphoinositide-3 kinase (PI3K)/Akt signal pathway plays a key role in the tumorigenesis of many cancers and in the subsequent development of drug resistance. Using the K562 chronic myelogenous leukemia (CML) cell line and the doxorubicin-resistant derivative lines KD30 and KD225 as models, we observed that enhanced PI3K/Akt activity and the acquisition of chemoresistance correlated unexpectedly with the increased expression and nuclear accumulation of FOXO3a. Moreover, we found that the induction of FOXO3a activity in naďve K562 cells was sufficient to enhance PI3K/Akt activity and to confer resistance to the cytotoxic effects of doxorubicin. Conversely, the knockdown of endogenous FOXO3a expression reduced PI3K/Akt activity and sensitized these cells to doxorubicin. Further chromatin immunoprecipitation and promoter mutation analyses demonstrated that FOXO3a regulates the expression of the PI3K catalytic subunit p110? through the activation of a promoter region proximal to a novel untranslated exon upstream from the reported transcription start site of the p110? gene PIK3CA. As was the case for FOXO3a, the expression or knockdown of p110? was sufficient to amplify or reduce PI3K/Akt activity, respectively. Thus, our results suggest that the chronic activation of FOXO3a by doxorubicin in CML cells can enhance survival through a feedback mechanism that involves enhanced p110? expression and hyperactivation of the PI3K/Akt pathway. PMID:18644865

  13. The forkhead transcription factor FOXO3a increases phosphoinositide-3 kinase/Akt activity in drug-resistant leukemic cells through induction of PIK3CA expression.

    PubMed

    Hui, Rosaline C-Y; Gomes, Ana R; Constantinidou, Demetra; Costa, Joana R; Karadedou, Christina T; Fernandez de Mattos, Silvia; Wymann, Matthias P; Brosens, Jan J; Schulze, Almut; Lam, Eric W-F

    2008-10-01

    The phosphoinositide-3 kinase (PI3K)/Akt signal pathway plays a key role in the tumorigenesis of many cancers and in the subsequent development of drug resistance. Using the K562 chronic myelogenous leukemia (CML) cell line and the doxorubicin-resistant derivative lines KD30 and KD225 as models, we observed that enhanced PI3K/Akt activity and the acquisition of chemoresistance correlated unexpectedly with the increased expression and nuclear accumulation of FOXO3a. Moreover, we found that the induction of FOXO3a activity in naďve K562 cells was sufficient to enhance PI3K/Akt activity and to confer resistance to the cytotoxic effects of doxorubicin. Conversely, the knockdown of endogenous FOXO3a expression reduced PI3K/Akt activity and sensitized these cells to doxorubicin. Further chromatin immunoprecipitation and promoter mutation analyses demonstrated that FOXO3a regulates the expression of the PI3K catalytic subunit p110alpha through the activation of a promoter region proximal to a novel untranslated exon upstream from the reported transcription start site of the p110alpha gene PIK3CA. As was the case for FOXO3a, the expression or knockdown of p110alpha was sufficient to amplify or reduce PI3K/Akt activity, respectively. Thus, our results suggest that the chronic activation of FOXO3a by doxorubicin in CML cells can enhance survival through a feedback mechanism that involves enhanced p110alpha expression and hyperactivation of the PI3K/Akt pathway. PMID:18644865

  14. Studies on the metabolism of metallothionein and alkaline phosphatase of adult rat primary hepatocyte cultures: role of fetal calf serum and agonists of the phosphoinositide cascade

    Microsoft Academic Search

    K. Krfimer; A. Markwitan; J. Pallauf

    1993-01-01

    Summary Adult rat primary hepatocytes maintained in DMEM\\/F12 (Ham) media were used as a model system for studying the role of fetal calf serum (FCS) and agonists of the phosphoinositide cascade in the metabolism of metallothionein (MT) and alkaline phosphatase (ALP). Experiments were performed both after a 24 h preincubation with FCS and with bovine serum albumin (BSA). Hepatocytes were

  15. Response to platelet-activating factor in human platelets stored and aged in plasma. Decrease in aggregation, phosphoinositide turnover, and receptor affinity

    SciTech Connect

    Shukla, S.D.; Morrison, W.J.; Klachko, D.M.

    1989-07-01

    Human platelet concentrates were stored in polyolefin bags at 22 to 24 degrees C on a horizontal shaker for up to 8 days. At different intervals, aliquots of platelet-rich plasma (PRP) were removed aseptically and five variables, i.e., platelet counts, morphology, platelet-activating factor (PAF)-stimulated aggregation, phosphoinositide turnover, and (3H)PAF binding to platelet receptors, were studied. The number of platelets did not change during the 8 days of storage. Scanning electron microscopy of the platelets revealed a gradual morphologic change from biconcave flat discs to irregular, crenated forms. The PAF-induced aggregation of platelets declined with time of storage. A decrease to 50 percent of the Day 1 aggregatory response to PAF was evident on Day 2, and there was a further decline to about 20 percent by Day 6. Similarly, PAF receptor-coupled phosphoinositide turnover, as monitored by 32P incorporation into individual phosphoinositides, decreased dramatically with storage. After 2 to 3 days of storage, the phosphoinositide turnover was reduced to 50 percent of the original response, and it continued to decline to about 25 percent of original response by Day 5 or 6. The binding of (3H)PAF to washed human platelets indicated subtle changes between Days 2 and 4, which became more noticeable by Day 6. These results have raised the possibility of changes in the number of the receptors and/or their affinity for the ligand during storage. We conclude that although the number of platelets was maintained during storage for 8 days, a general deterioration of their responses to PAF occurred at the levels of cell surface receptor, transmembrane signaling (phosphoinositide turnover), and response (aggregation).

  16. Structure-Based Design of Potent and Selective 3-Phosphoinositide-Dependent Kinase-1 (PDK1) Inhibitors

    SciTech Connect

    Medina, Jesus R.; Becker, Christopher J.; Blackledge, Charles W.; Duquenne, Celine; Feng, Yanhong; Grant, Seth W.; Heerding, Dirk; Li, William H.; Miller, William H.; Romeril, Stuart P.; Scherzer, Daryl; Shu, Arthur; Bobko, Mark A.; Chadderton, Antony R.; Dumble, Melissa; Gardiner, Christine M.; Gilbert, Seth; Liu, Qi; Rabindran, Sridhar K.; Sudakin, Valery; Xiang, Hong; Brady, Pat G.; Campobasso, Nino; Ward, Paris; Axten, Jeffrey M. (GSKPA)

    2014-10-02

    Phosphoinositide-dependent protein kinase-1(PDK1) is a master regulator of the AGC family of kinases and an integral component of the PI3K/AKT/mTOR pathway. As this pathway is among the most commonly deregulated across all cancers, a selective inhibitor of PDK1 might have utility as an anticancer agent. Herein we describe our lead optimization of compound 1 toward highly potent and selective PDK1 inhibitors via a structure-based design strategy. The most potent and selective inhibitors demonstrated submicromolar activity as measured by inhibition of phosphorylation of PDK1 substrates as well as antiproliferative activity against a subset of AML cell lines. In addition, reduction of phosphorylation of PDK1 substrates was demonstrated in vivo in mice bearing OCl-AML2 xenografts. These observations demonstrate the utility of these molecules as tools to further delineate the biology of PDK1 and the potential pharmacological uses of a PDK1 inhibitor.

  17. The phosphoinositide dependent-phospholipase C pathway differentially controls the basal expression of DREB1 and DREB2genes

    PubMed Central

    Ruelland, Eric; Djafi, Nabila; Zachowski, Alain

    2013-01-01

    We recently showed that—in Arabidopsis thaliana suspension cells—phosphoinositide dependent-phospholipase C (PI-PLC) and diacylglycerol kinase (DGK) negatively regulated the basal expression of most DREB2 genes. DREB2 genes encode transcription factors that bind to Drought Responsive Elements (DRE). Those elements are also bound by DREB1 factors. While DREB2 factors are mostly involved in drought and heat responses, DREB1s are induced in the response to chilling. We here show that the pharmacological inhibition of PI-PLC or DGK leads to the basal induction of DREB1 genes. However, the induction is much less marked for the DREB1 genes than that of DREB2A, a member of the DREB2 family. This illustrates that DREB1 and DREB2 genes, while having the same targets, are not submitted to the same transcription regulation, and that lipid signaling might in part explain these differences in the regulation of the DREB genes.

  18. Expression of 3-phosphoinositide-dependent protein kinase 1 in colorectal cancer as a potential therapeutic target.

    PubMed

    Xu, Zhenglei; Liao, Bihong; Zhang, Ru; Yao, Jun; Shi, Ruiyue; Wang, Lisheng

    2015-07-01

    3-Phosphoinositide-dependent protein kinase 1 (PDK1) is centrally involved in cancer progression, including proliferation, apoptosis and invasion. However, its expression pattern and possible cellular functions in human colorectal cancer remain unclear. In the present study, we show that PDK1 expression is up-regulated at both mRNA and protein levels in colorectal cancer clinical specimens and cell lines. Transient knockdown of PDK1 suppresses cellular growth, induces cellular apoptosis and causes abnormal cell cycle distribution. Meanwhile, decreased PDK1 level is closely associated with reduced Akt/cyclin D1 activity. Activating AKT activity and reintroducing cyclin D1 expression significantly compromised the oncogenic activity induced by PDK1. Together, our findings elucidate a key role for PDK1 in colorectal cellular functions trigged by the Akt/cyclin D1 pathway, thus providing a novel insight of PDK1 in colorectal carcinogenesis. PMID:26055151

  19. Dominant-Activating, Germline Mutations in Phosphoinositide 3-Kinase p110? Cause T Cell Senescence and Human Immunodeficiency

    PubMed Central

    Lucas, Carrie L.; Kuehn, Hye Sun; Zhao, Fang; Niemela, Julie E.; Deenick, Elissa K.; Palendira, Umaimainthan; Avery, Danielle T.; Moens, Leen; Cannons, Jennifer L.; Biancalana, Matthew; Stoddard, Jennifer; Ouyang, Weiming; Frucht, David L.; Rao, V. Koneti; Atkinson, T. Prescott; Agharahimi, Anahita; Hussey, Ashleigh A.; Folio, Les R.; Olivier, Kenneth N.; Fleisher, Thomas A.; Pittaluga, Stefania; Holland, Steven M.; Cohen, Jeffrey I.; Oliviera, Joao B.; Tangye, Stuart G.; Schwartzberg, Pamela L.; Lenardo, Michael J.; Uzel, Gulbu

    2014-01-01

    The p110? subunit of phosphoinositide 3-kinase (PI(3)K) is selectively expressed in leukocytes and is critical for lymphocyte biology. Here we report three different germline, heterozygous, gain-of-function mutations in the PIK3CD gene encoding p110? in fourteen patients from seven families. These patients presented with sinopulmonary infections, lymphadenopathy, nodular lymphoid hyperplasia and CMV and/or EBV viremia. Strikingly, naďve and central memory T cells were severely deficient, while senescent effector T cells were over-represented. In vitro, patient T cells exhibited increased phosphorylation of Akt and hyperactivation of mTOR, enhanced glucose uptake and terminal effector differentiation. Importantly, treatment with rapamycin to inhibit mTOR activity in vivo partially restored naďve T cells, largely rescued the in vitro T cell defects, and improved clinical course. PMID:24165795

  20. 3-Phosphoinositide-dependent protein kinase-1/Akt signalling and inhibition in a canine prostate carcinoma cell line.

    PubMed

    Alvarez, F J; Murahari, S; Couto, C G; Rosol, T J; Kulp, S K; Chen, C-S; Kisseberth, W C

    2007-03-01

    Deregulation of the 3-phosphoinositide-dependent protein kinase-1 (PDK-1)/Akt signalling pathway is associated with prostate cancer development and progression. Inhibition of PDK-1/Akt signalling can be achieved using structurally optimized celecoxib derivatives such as OSU-03012. In this study, we treated the novel canine prostate cancer cell line, Ace-1, with OSU-03012 or dimethyl sulphoxide in vitro. We found that Akt was constitutively phosphorylated in the canine prostate cancer cell line Ace-1 and that there was a dose-dependent decrease in cell viability, and Akt and glycogen synthase kinase-3beta phosphorylation, in response to OSU-03012 treatment. This was accompanied by a dose-dependent increase in apoptosis. These data suggest that Akt signalling pathway inhibition is a potential strategy for the treatment of dogs with prostate cancer and that canine prostate cancer is a relevant large animal model for evaluating Akt pathway inhibitors such as OSU-03012 for use in people. PMID:19754801

  1. Phosphoinositide 3-Kinase p110? activity: Key Role in Metabolism and Mammary Gland Cancer but not Development #

    PubMed Central

    Ciraolo, Elisa; Iezzi, Manuela; Marone, Romina; Marengo, Stefano; Curcio, Claudia; Costa, Carlotta; Azzolino, Ornella; Gonella, Cristiano; Rubinetto, Cristina; Wu, Haiyan; Dastrů, Walter; Martin, Erica L.; Silengo, Lorenzo; Altruda, Fiorella; Turco, Emilia; Lanzetti, Letizia; Musiani, Piero; Rückle, Thomas; Rommel, Christian; Backer, Jonathan M.; Forni, Guido; Wymann, Matthias P.; Hirsch, Emilio

    2009-01-01

    The phosphoinositide 3-kinase (PI3K) pathway crucially controls metabolism and cell growth. Although different PI3K catalytic subunits are known to play distinct roles, the specific in vivo function of p110? (the product of the PIK3CB gene) is not clear. Here, we show that mouse mutants expressing a catalytically inactive PIK3CBK805R mutant survived to adulthood but showed growth retardation and developed mild insulin resistance with age. Pharmacological and genetic analyses of p110? function revealed that p110? catalytic activity is required for PI3K signaling downstream of heterotrimeric guanine nucleotide-binding (G protein)-coupled receptors as well as to sustain long term insulin signaling. In addition, PIK3CBK805R mice were protected in a model of ERBB2-driven tumor development. These findings indicate an unexpected role for p110? catalytic activity in diabetes and cancer, opening potential new avenues for therapeutic intervention. PMID:18780892

  2. A novel class of PTEN protein in Arabidopsis displays unusual phosphoinositide phosphatase activity and efficiently binds phosphatidic acid.

    PubMed

    Pribat, Anne; Sormani, Rodnay; Rousseau-Gueutin, Mathieu; Julkowska, Magdalena M; Testerink, Christa; Joubčs, Jerôme; Castroviejo, Michel; Laguerre, Michel; Meyer, Christian; Germain, Véronique; Rothan, Christophe

    2012-01-01

    PTEN (phosphatase and tensin homologue deleted on chromosome ten) proteins are dual phosphatases with both protein and phosphoinositide phosphatase activity. They modulate signalling pathways controlling growth, metabolism and apoptosis in animals and are implied in several human diseases. In the present paper we describe a novel class of PTEN pro-teins in plants, termed PTEN2, which comprises the AtPTEN (Arabidopsis PTEN) 2a and AtPTEN2b proteins in Arabidopsis. Both display low in vitro tyrosine phosphatase activity. In addition, AtPTEN2a actively dephosphorylates in vitro the 3' phosphate group of PI3P (phosphatidylinositol 3-phosphate), PI(3,4)P2 (phosphatidylinositol 3,4-bisphosphate) and PI(3,5)P2 (phosphatidylinositol 3,5-bisphosphate). In contrast with animal PTENs, PI(3,4,5)P3 (phosphatidylinositol 3,4,5-trisphosphate) is a poor substrate. Site-directed mutagenesis of AtPTEN2a and molecular modelling of protein-phosphoinositide interactions indicated that substitutions at the PTEN2 core catalytic site of the Lys267 and Gly268 residues found in animals, which are critical for animal PTEN activity, by Met267 and Ala268 found in the eudicot PTEN2 are responsible for changes in substrate specificity. Remarkably, the AtPTEN2a protein also displays strong binding activity for PA (phosphatidic acid), a major lipid second messenger in plants. Promoter::GUS (?-glucuronidase) fusion, transcript and protein analyses further showed the transcriptional regulation of the ubiquitously expressed AtPTEN2a and AtPTEN2b by salt and osmotic stress. The results of the present study suggest a function for this novel class of plant PTEN proteins as an effector of lipid signalling in plants. PMID:21864294

  3. Phosphoinositides and phosphatidic acid regulate pollen tube growth and reorientation through modulation of [Ca2+]c and membrane secretion.

    PubMed

    Monteiro, David; Liu, Qunlu; Lisboa, Saskia; Scherer, G E F; Quader, Hartmut; Malhó, Rui

    2005-06-01

    The maintenance of a calcium gradient and vesicle secretion in the apex of pollen tubes is essential for growth. It is shown here that phosphatidylinositol-4,5-bisphosphate (PIP2) and D-myo-inositol-1,4,5-trisphosphate (IP3), together with phosphatidic acid (PA), play a vital role in the regulation of these processes. Changes in the intracellular concentration of both PIP2 and IP3 (induced by photolysis of caged-probes), modified growth and caused reorientation of the growth axis. However, measurements of cytosolic free calcium ([Ca2+]c) and apical secretion revealed significant differences between the photo-release of PIP2 or IP3. When released in the first 50 mum of the pollen tube, PIP2 led to transient growth perturbation, [Ca2+]c increases, and inhibition of apical secretion. By contrast, a concentration of IP3 which caused a [Ca2+]c transient of similar magnitude, stimulated apical secretion and caused severe growth perturbation. Furthermore, the [Ca2+]c transient induced by IP3 was spatially different causing a pronounced elevation in the sub-apical region. These observations suggest different targets for the two phosphoinositides. One of the targets is suggested to be PA, a product of PIP2 hydrolysis via phospholipase C (PLC) or phospholipase D (PLD) activity. It was found that antagonists of PA accumulation (e.g. butan-1-ol) and inhibitors of PLC and PLD reversibly halted polarity. Reduction of PA levels caused the dissipation of the [Ca2+]c gradient and inhibited apical plasma membrane recycling. It was also found to cause abolition of the apical zonation. These data suggest that phosphoinositides and phospholipids regulate tip growth through a multiple pathway system involving regulation of [Ca2+]c levels, endo/exocytosis, and vesicular trafficking. PMID:15837704

  4. Solution structure of the C-terminal SH2 domain of the p85? regulatory subunit of phosphoinositide 3-kinase1

    Microsoft Academic Search

    Gregg Siegal; Ben Davis; Sřren M. Kristensen; Andrew Sankar; Jeffrey Linacre; Robert C. Stein; George Panayotou; Michael D. Waterfield; Paul C. Driscoll

    1998-01-01

    Heterodimeric class IA phosphoinositide 3-kinase (PI 3-kinase) plays a crucial role in a variety of cellular signalling events downstream of a number of cell-surface receptor tyrosine kinases. Activation of the enzyme is effected in part by the binding of two Src homology-2 domains (SH2) of the 85 kDa regulatory subunit to specific phosphotyro- sine-containing peptide motifs within activated cytoplasmic receptor

  5. Phosphoinositide 3-kinase ? controls autonomic regulation of the mouse heart through G i-independent downregulation of cAMP level

    Microsoft Academic Search

    Giuseppe Alloatti; Andrea Marcantoni; Renzo Levi; Maria Pia Gallo; Lorenzo Del Sorbo; Enrico Patrucco; Laura Barberis; Daniela Malan; Ornella Azzolino; Matthias Wymann; Emilio Hirsch; Giuseppe Montrucchio

    2005-01-01

    Cardiac ?-adrenergic and the muscarinic receptors control contractility and heart rate by triggering multiple signaling events involving downstream targets like the phosphoinositide 3-kinase ? (PI3K?). We thus investigated whether the lack of PI3K? could play a role in the autonomic regulation of the mouse heart. Contractility and ICaL of mutant cardiac preparations appeared increased in basal conditions and after ?-adrenergic

  6. The Forkhead Transcription Factor FOXO3a Increases Phosphoinositide3 Kinase\\/Akt Activity in Drug-Resistant Leukemic Cells through Induction of PIK3CA Expression

    Microsoft Academic Search

    R. C.-Y. Hui; A. R. Gomes; D. Constantinidou; J. R. Costa; C. T. Karadedou; S. Fernandez de Mattos; M. P. Wymann; J. J. Brosens; A. Schulze; E. W.-F. Lam

    2008-01-01

    The phosphoinositide-3 kinase (PI3K)\\/Akt signal pathway plays a key role in the tumorigenesis of many cancers and in the subsequent development of drug resistance. Using the K562 chronic myelogenous leukemia (CML) cell line and the doxorubicin-resistant derivative lines KD30 and KD225 as models, we observed that enhanced PI3K\\/Akt activity and the acquisition of chemoresistance correlated unexpectedly with the increased expression

  7. Modulation of phosphoinositide–protein kinase C signal transduction by omega-3 fatty acids: Implications for the pathophysiology and treatment of recurrent neuropsychiatric illness

    Microsoft Academic Search

    Robert K. McNamara; Michelle Ostrander; William Abplanalp; Neil M. Richtand; Stephen C. Benoit; Debbie J. Clegg

    2006-01-01

    The phosphoinositide (PI)–protein kinase C (PKC) signal transduction pathway is initiated by pre- and postsynaptic G?q-coupled receptors, and regulates several clinically relevant neurochemical events, including neurotransmitter release efficacy, monoamine receptor function and trafficking, monoamine transporter function and trafficking, axonal myelination, and gene expression. Mounting evidence for PI–PKC signaling hyperactivity in the peripheral (platelets) and central (premortem and postmortem brain) tissues

  8. Enhanced Ca2+-dependent activation of phosphoinositide 3-kinase class II? isoform-Rho axis in blood vessels of spontaneously hypertensive rats.

    PubMed

    Seok, Young Mi; Azam, Mohammed Ali; Okamoto, Yasuo; Sato, Atsushi; Yoshioka, Kazuaki; Maeda, Masataka; Kim, Inkyeom; Takuwa, Yoh

    2010-11-01

    Rho-mediated inhibition of myosin light chain (MLC) phosphatase (MLCP), together with Ca(2+)-dependent MLC kinase activation, constitutes the major signaling mechanisms for vascular smooth muscle contraction. We recently unveiled the involvement of Ca(2+)-induced, phosphoinositide 3-kinase (PI3K) class II? isoform (PI3K-C2?)-dependent Rho activation and resultant Rho kinase-dependent MLCP suppression in membrane depolarization- and receptor agonist-induced contraction. It is unknown whether Ca(2+)- and PI3K-C2?-dependent regulation of MLCP is altered in vascular smooth muscle of hypertensive animals and is involved in hypertension. Therefore, we studied the role of the Ca(2+)-PI3K-C2?-Rho-MLCP pathway in spontaneously hypertensive rats (SHRs). PI3K-C2? was readily detected in various vascular beds of Wistar-Kyoto rats and activated by high KCl. High KCl also stimulated vascular Rho activity and phosphorylation of the MLCP regulatory subunit MYPT1 at Thr(853) in a PI3K inhibitor wortmannin-sensitive manner. In mesenteric and other vessels of SHRs at the hypertensive but not the prehypertensive stage, the activity of PI3K-C2? but not class I PI3K p110? was elevated with concomitant rises of Rho activity and Thr(853)-phosphorylation of MYPT1, as compared with normotensive controls. Infusion of the Ca(2+) channel antagonist nicardipine reduced blood pressure with suppression of vascular activity of PI3K-C2?-Rho and phosphorylation of MYPT1 in hypertensive SHRs. Infusion of wortmannin lowered blood pressure with inhibition of PI3K-C2?-Rho activities and MYPT1 phosphorylation in hypertensive SHRs. These observations suggest that an increased activity of the Ca(2+)-PI3K-C2?-Rho signaling pathway with resultant augmented MLCP suppression contributes to hypertension in SHRs. The Ca(2+)- and PI3K-C2?-dependent Rho stimulation in vascular smooth muscle may be a novel, promising target for treating hypertension. PMID:20921425

  9. Phosphoinositide metabolism links cGMP-dependent protein kinase G to essential Ca˛? signals at key decision points in the life cycle of malaria parasites.

    PubMed

    Brochet, Mathieu; Collins, Mark O; Smith, Terry K; Thompson, Eloise; Sebastian, Sarah; Volkmann, Katrin; Schwach, Frank; Chappell, Lia; Gomes, Ana Rita; Berriman, Matthew; Rayner, Julian C; Baker, David A; Choudhary, Jyoti; Billker, Oliver

    2014-03-01

    Many critical events in the Plasmodium life cycle rely on the controlled release of Ca˛? from intracellular stores to activate stage-specific Ca˛?-dependent protein kinases. Using the motility of Plasmodium berghei ookinetes as a signalling paradigm, we show that the cyclic guanosine monophosphate (cGMP)-dependent protein kinase, PKG, maintains the elevated level of cytosolic Ca˛? required for gliding motility. We find that the same PKG-dependent pathway operates upstream of the Ca˛? signals that mediate activation of P. berghei gametocytes in the mosquito and egress of Plasmodium falciparum merozoites from infected human erythrocytes. Perturbations of PKG signalling in gliding ookinetes have a marked impact on the phosphoproteome, with a significant enrichment of in vivo regulated sites in multiple pathways including vesicular trafficking and phosphoinositide metabolism. A global analysis of cellular phospholipids demonstrates that in gliding ookinetes PKG controls phosphoinositide biosynthesis, possibly through the subcellular localisation or activity of lipid kinases. Similarly, phosphoinositide metabolism links PKG to egress of P. falciparum merozoites, where inhibition of PKG blocks hydrolysis of phosphatidylinostitol (4,5)-bisphosphate. In the face of an increasing complexity of signalling through multiple Ca˛? effectors, PKG emerges as a unifying factor to control multiple cellular Ca˛? signals essential for malaria parasite development and transmission. PMID:24594931

  10. Regulation of the phosphoinositide pathway in cultured Sertoli cells from immature rats: effects of follicle-stimulating hormone and fluoride

    SciTech Connect

    Quirk, S.M.; Reichert, L.E. Jr.

    1988-07-01

    Many hormones elicit effects on target cells by stimulating the enzyme phospholipase-C, which catalyzes the hydrolysis of phosphoinositides to the intracellular second messengers diacylglycerol and inositol phosphates. The present study examined the roles of FSH and guanine nucleotide-binding proteins (G-proteins) in regulating the hydrolysis of phosphoinositides in Sertoli cells. Sertoli cell cultures prepared from 16- to 18-day-old rats were incubated for 24 h with myo-(2-3H) inositol to label endogenous phospholipids. Treatment of cells from 0.5-20 min with preparations of ovine FSH ranging in potency from 1-60 times that of NIH FSH S1 did not affect accumulation of inositol phosphates. Levels of total (3H)inositol phosphates ((3H)inositol mono-, di-, and triphosphates (IP, IP2, and IP3)) in FSH-treated cultures was 75-120% the levels in control cultures over the various time intervals studied. Addition of testosterone and the combination of testosterone plus retinoic acid, agents that have been shown to potentiate effects of FSH in other systems, did not affect accumulation of inositol phosphates in response to FSH. In contrast to the lack of effect on accumulation of inositol phosphates, FSH stimulated 4- to 11-fold increases in estradiol secretion over 24 h of culture, indicating that Sertoli cells were viable and responsive to FSH. AIF4- has been shown to activate G-proteins involved in regulation of adenylate cyclase activity. In the present study, AIF4- induced 4- to 5-fold increases in IP, IP2, and IP3 in experiments wherein FSH had no effect. Pretreatment of Sertoli cells with pertussis toxin (100 and 1000 ng/ml) for 24 h inhibited fluoride-induced generation of IP, IP2, and IP3 by 24-51%. Similar treatment with cholera toxin had no effect on basal or fluoride-induced generation of IP2 or IP3, but increased fluoride-induced generation of IP by 20-34%.

  11. The Shigella flexneri type three secretion system effector IpgD inhibits T cell migration by manipulating host phosphoinositide metabolism.

    PubMed

    Konradt, Christoph; Frigimelica, Elisabetta; Nothelfer, Katharina; Puhar, Andrea; Salgado-Pabon, Wilmara; di Bartolo, Vincenzo; Scott-Algara, Daniel; Rodrigues, Cristina D; Sansonetti, Philippe J; Phalipon, Armelle

    2011-04-21

    Shigella, the Gram-negative enteroinvasive bacterium that causes shigellosis, relies on its type III secretion system (TTSS) and injected effectors to modulate host cell functions. However, consequences of the interaction between Shigella and lymphocytes have not been investigated. We show that Shigella invades activated human CD4(+) T lymphocytes. Invasion requires a functional TTSS and results in inhibition of chemokine-induced T cell migration, an effect mediated by the TTSS effector IpgD, a phosphoinositide 4-phosphatase. Remarkably, IpgD injection into bystander T cells can occur in the absence of cell invasion. Upon IpgD-mediated hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP(2)), the pool of PIP(2) at the plasma membrane is reduced, leading to dephosphorylation of the ERM proteins and their inability to relocalize at one T cell pole upon chemokine stimulus, likely affecting the formation of the polarized edge required for cell migration. These results reveal a bacterial TTSS effector-mediated strategy to impair T cell function. PMID:21501826

  12. A method to control phosphoinositides and to analyze PTEN function in living cells using voltage sensitive phosphatases

    PubMed Central

    Mavrantoni, Angeliki; Thallmair, Veronika; Leitner, Michael G.; Schreiber, Daniela N.; Oliver, Dominik; Halaszovich, Christian R.

    2015-01-01

    Voltage sensitive phosphatases (VSPs), including engineered voltage sensitive PTEN, are excellent tools to rapidly and reversibly alter the phosphoinositide (PI) content of the plasma membrane in vivo and study the tumor suppressor PTEN. However, widespread adoption of these tools is hampered by the requirement for electrophysiological instrumentation to control the activity of VSPs. Additionally, monitoring and quantifying the PI changes in living cells requires sophisticated microscopy equipment and image analysis. Here we present methods that bypass these obstacles. First, we explore technically simple means for activation of VSPs via extracellularly applied agents or light. Secondly, we characterize methods to monitor PI(4,5)P2 and PI(3,4,5)P3 levels using fluorescence microscopy or photometry in conjunction with translocation or FRET based PI probes, respectively. We then demonstrate the application of these techniques by characterizing the effect of known PTEN mutations on its enzymatic activity, analyzing the effect of PTEN inhibitors, and detecting in real time rapid inhibition of protein kinase B following depletion of PI(3,4,5)P3. Thus, we established an approach that does not only allow for rapidly manipulating and monitoring PI(4,5)P2 and PI(3,4,5)P3 levels in a population of cells, but also facilitates the study of PTEN mutants and pharmacological targeting in mammalian cells. PMID:25873899

  13. A Novel Function of eIF2? Kinases as Inducers of the Phosphoinositide-3 Kinase Signaling Pathway

    PubMed Central

    Kazemi, Shirin; Mounir, Zineb; Baltzis, Dionissios; Raven, Jennifer F.; Wang, Shuo; Krishnamoorthy, Jothi-Latha; Pluquet, Olivier; Pelletier, Jerry

    2007-01-01

    Phosphoinositide-3 kinase (PI3K) plays an important role in signal transduction in response to a wide range of cellular stimuli involved in cellular processes that promote cell proliferation and survival. Phosphorylation of the ? subunit of the eukaryotic translation initiation factor eIF2 at Ser51 takes place in response to various types of environmental stress and is essential for regulation of translation initiation. Herein, we show that a conditionally active form of the eIF2? kinase PKR acts upstream of PI3K and turns on the Akt/PKB-FRAP/mTOR pathway leading to S6 and 4E-BP1 phosphorylation. Also, induction of PI3K signaling antagonizes the apoptotic and protein synthesis inhibitory effects of the conditionally active PKR. Furthermore, induction of the PI3K pathway is impaired in PKR?/? or PERK?/? mouse embryonic fibroblasts (MEFs) in response to various stimuli that activate each eIF2? kinase. Mechanistically, PI3K signaling activation is indirect and requires the inhibition of protein synthesis by eIF2? phosphorylation as demonstrated by the inactivation of endogenous eIF2? by small interfering RNA or utilization of MEFs bearing the eIF2? Ser51Ala mutation. Our data reveal a novel property of eIF2? kinases as activators of PI3K signaling and cell survival. PMID:17596516

  14. Disruption of Phosphoinositide-Specific Phospholipases C?1 Contributes to Extracellular Matrix Synthesis of Human Osteoarthritis Chondrocytes

    PubMed Central

    Zeng, Guoqing; Cui, Xu; Liu, Zejun; Zhao, Honghai; Zheng, Xinpeng; Zhang, Bing; Xia, Chun

    2014-01-01

    Osteoarthritis (OA) is a degenerative joint disease characterized by articular cartilage degradation including extracellular matrix (ECM) degradation and cell loss. It is known that phosphoinositide-specific phospholipase ?1 (PLC?1) can trigger several signaling pathways to regulate cell metabolism. However, whether this kinase is expressive and active in human OA chondrocytes and its role in the pathological progression of OA have not been investigated. The current study was designed to investigate the PLC?1 expression in human OA cartilage, and whether PLC?1 was involved in the ECM synthesis had been further explored using cultured human OA chondrocytes. Our results indicated that PLC?1 was highly expressed in human OA chondrocytes. In our further study using the cultured human OA chondrocytes, the results demonstrated that the disruption of PLC?1 by its inhibitor, U73122, and siRNA contributed to the ECM synthesis of human OA chondrocytes through regulating the expression of ECM-related signaling molecules, including MMP-13, Col II, TIMP1, Sox-9, and AGG. Furthermore, PLC?1/IP3/Ca(2+)/CaMK II signaling axis regulated the ECM synthesis of human chondrocytes through triggering mTOR/P70S6K/S6 pathway. In summary, our results suggested that PLC-?1 activities played an important role in the ECM synthesis of human OA chondrocytes, and may serve as a therapeutic target for treating OA. PMID:25073093

  15. Role of calcium ions in rapid effects of L-thyroxine on phosphoinositide metabolism in rat liver cells.

    PubMed

    Krasilnikova, O A; Kavok, N S; Babenko, N A

    2003-07-01

    The role of calcium ions in the L-thyroxine-induced initiation of hydrolysis of phosphatidylinositol 4,5-bisphosphate (PtdInsP2) and also the course of releasing individual fractions of inositol phosphates and diacylglycerides (DAG) were studied in liver cells during early stages of the hormone effect. L-Thyroxine stimulated a rapid hydrolysis in hepatocytes of PtdInsP2 labeled with [14C]linoleic acid and [3H]inositol mediated by phosphoinositide-specific phospholipase C. This was associated with accumulation of [14C]DAG, total inositol phosphates, [3H]inositol 1,4,5-trisphosphate (Ins1,4,5P3) and [3H]inositol 1,4-bisphosphate (Ins1,4P2). Elimination of calcium ions from the incubation medium of hepatocytes did not abolish the effect of thyroxine on the accumulation of [14C]DAG and total [3H]inositol phosphates. Preincubation of liver cells with TMB-8 increased the stimulatory effect of L-thyroxine on the accumulation of [14C]DAG. During the incubation of hepatocytes in the presence of the hormone the content of 14C-labeled fatty acids did not change. The L-thyroxine-induced accumulation of [3H]Ins1,4,5P3 and [3H]Ins1,4P2 did not depend on the presence of calcium ions in the incubation medium of the cells. PMID:12946260

  16. Expression Analysis of a Stress-Related Phosphoinositide-Specific Phospholipase C Gene in Wheat (Triticum aestivum L.)

    PubMed Central

    Wu, Lizhu; Hou, Mingyu; Dou, Shijuan; Pan, Yanyun

    2014-01-01

    Plant phosphoinositide-specific phospholipases C (PI-PLCs) function in several essential plant processes associated with either development or environmental stress. In this report, we examined the expression patterns of TaPLC1 under drought and high salinity stress at the transcriptional and post-transcriptional levels. TaPLC1 mRNA was expressed in all wheat organs examined. U73122 and edelfosine, the PLC inhibitor, impaired seedling growth and enhanced seedling sensitivity to drought and high salinity stress. Though TaPLC1 expression in wheat was lowest at the seedling stage, it was strongly induced under conditions of stress. When 6-day-old wheat seedlings were treated with 200 mM NaCl or 20% (w/v) PEG 6000 for 6 or 12 h, respectively, the TaPLC1 transcript level increased by 16-fold compared to the control. Western blotting showed that the TaPLC protein concentration was also maintained at a high level from 24 to 48 h during stress treatment. Together, our results indicate the possible biological functions of TaPLC1 in regulating seedling growth and the response to drought and salinity stress. PMID:25121594

  17. Phosphoinositide 3-kinase p110beta activity: key role in metabolism and mammary gland cancer but not development.

    PubMed

    Ciraolo, Elisa; Iezzi, Manuela; Marone, Romina; Marengo, Stefano; Curcio, Claudia; Costa, Carlotta; Azzolino, Ornella; Gonella, Cristiano; Rubinetto, Cristina; Wu, Haiyan; Dastrů, Walter; Martin, Erica L; Silengo, Lorenzo; Altruda, Fiorella; Turco, Emilia; Lanzetti, Letizia; Musiani, Piero; Rückle, Thomas; Rommel, Christian; Backer, Jonathan M; Forni, Guido; Wymann, Matthias P; Hirsch, Emilio

    2008-01-01

    The phosphoinositide 3-kinase (PI3K) pathway crucially controls metabolism and cell growth. Although different PI3K catalytic subunits are known to play distinct roles, the specific in vivo function of p110beta (the product of the PIK3CB gene) is not clear. Here, we show that mouse mutants expressing a catalytically inactive PIK3CB(K805R) mutant survived to adulthood but showed growth retardation and developed mild insulin resistance with age. Pharmacological and genetic analyses of p110beta function revealed that p110beta catalytic activity is required for PI3K signaling downstream of heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors as well as to sustain long-term insulin signaling. In addition, PIK3CB(K805R) mice were protected in a model of ERBB2-driven tumor development. These findings indicate an unexpected role for p110beta catalytic activity in diabetes and cancer, opening potential avenues for therapeutic intervention. PMID:18780892

  18. Involvement of Class II Phosphoinositide 3-Kinase ?-Isoform in Antigen-Induced Degranulation in RBL-2H3 Cells

    PubMed Central

    Nigorikawa, Kiyomi; Hazeki, Kaoru; Guo, Ying; Hazeki, Osamu

    2014-01-01

    In this study, we present findings that suggest that PI3K-C2?, a member of the class II phosphoinositide 3-kinase (PI3K) subfamily, regulates the process of Fc?RI-triggered degranulation. RBL-2H3 cells were transfected with shRNA targeting PI3K-C2?. The knockdown impaired the Fc?RI-induced release of a lysosome enzyme, ?-hexosaminidase, without affecting the intracellular Ca2+ mobilization. The release of mRFP-tagged neuropeptide-Y, a reporter for the regulated exocytosis, was also decreased in the PI3K-C2?-deficient cells. The release was increased significantly by the expression of the siRNA-resistant version of PI3K-C2?. In wild-type cells, Fc?RI stimulation induced the formation of large vesicles, which were associated with CD63, a marker protein of secretory granules. On the vesicles, the existence of PI3K-C2? and PtdIns(3,4)P2 was observed. These results indicated that PI3K-C2? and its product PtdIns(3,4)P2 may play roles in the secretory process. PMID:25357130

  19. Spatial and Temporal Regulation of 3-Phosphoinositides by PI 3Kinase and PTEN Mediates Chemotaxis

    Microsoft Academic Search

    Satoru Funamoto; Ruedi Meili; Susan Lee; Lisa Parry; Richard A. Firtel

    2002-01-01

    We have investigated the mechanisms of leading edge formation in chemotaxing Dictyostelium cells. We demonstrate that while phosphatidylinositol 3-kinase (PI3K) transiently translocates to the plasma membrane in response to chemoattractant stimulation and to the leading edge in chemotaxing cells, PTEN, a negative regulator of PI3K pathways, exhibits a reciprocal pattern of localization. By uniformly localizing PI3K along the plasma membrane,

  20. The Phosphoinositide 3-Kinase Isoform PI3K? Regulates Osteoclast-Mediated Bone Resorption in Humans and Mice

    PubMed Central

    Gy?ri, Dávid; Csete, Dániel; Benk?, Szilvia; Kulkarni, Suhasini; Mandl, Péter; Dobó-Nagy, Csaba; Vanhaesebroeck, Bart; Stephens, Len; Hawkins, Phillip T; Mócsai, Attila

    2014-01-01

    Objective While phosphoinositide 3-kinases (PI3Ks) are involved in various intracellular signal transduction processes, the specific functions of the different PI3K isoforms are poorly understood. We have previously shown that the PI3K? isoform is required for arthritis development in the K/BxN serum–transfer model. Since osteoclasts play a critical role in pathologic bone loss during inflammatory arthritis and other diseases, we undertook this study to test the role of PI3K? in osteoclast development and function using a combined genetic and pharmacologic approach. Methods The role of PI3K? in primary human and murine osteoclast cultures was tested with the PI3K?-selective inhibitor TGX221 and by using PI3K??/? mice. The trabecular bone architecture of PI3K??/? mice was evaluated using micro–computed tomography and histomorphometric analyses. Results The expression of PI3K? was strongly and specifically up-regulated during in vitro osteoclast differentiation. In vitro development of large multinucleated osteoclasts from human or murine progenitors and their resorption capacity were strongly reduced by the PI3K? inhibitor TGX221 or by the genetic deficiency of PI3K?. This was likely due to defective cytoskeletal reorganization and vesicular trafficking, since PI3K??/? mouse multinucleated cells failed to form actin rings and retained intracellular acidic vesicles and cathepsin K. In contrast, osteoclast-specific gene expression and the survival and apoptosis of osteoclasts were not affected. PI3K??/? mice had significantly increased trabecular bone volume and showed abnormal osteoclast morphology with defective resorption pit formation. Conclusion PI3K? plays an important role in osteoclast development and function and is required for in vivo bone homeostasis. PMID:24719382

  1. Suppression of the {alpha}-isoform of class II phosphoinositide 3-kinase gene expression leads to apoptotic cell death

    SciTech Connect

    Kang, Shinhae [Technology Innovation Center, Cheju National University, Jeju, Jeju 690-756 (Korea, Republic of); Song, Jihoon [Department of Life Science, Cheju National University, Jeju, Jeju 690-756 (Korea, Republic of); Kang, Jihoon [Department of Medicine, Cheju National University, Jeju, Jeju 690-756 (Korea, Republic of); Kang, Heekyoung [Department of Medicine, Cheju National University, Jeju, Jeju 690-756 (Korea, Republic of); Lee, Daeho [Department of Medicine, Cheju National University, Jeju, Jeju 690-756 (Korea, Republic of); Lee, Youngki [Department of Medicine, Cheju National University, Jeju, Jeju 690-756 (Korea, Republic of); Park, Deokbae [Department of Medicine, Cheju National University, Jeju, Jeju 690-756 (Korea, Republic of)]. E-mail: parkdb@cheju.ac.kr

    2005-04-01

    Phosphoinositide 3-kinases (PI3Ks) have known to be key enzymes activating intracellular signaling molecules when a number of growth factors bind to their cell surface receptors. PI3Ks are divided into three classes (I, II, and III) and enzymes of each class have different tissue-specificities and physiological functions. Class II PI3Ks consist of three isoforms ({alpha}, {beta}, {gamma}). Although the {alpha}-isoform (PI3K-C2{alpha}) is considered ubiquitous and preferentially activated by insulin rather than the {beta}-isoform, the physiological significance of PI3K-C2{alpha} is poorly understood. The present study aimed to determine whether PI3K-C2{alpha} is associated with the suppression of apoptotic cell death. Different sense- and antisense oligonucleotides (ODNs) were synthesized based on the sequence of C2 domain of PI3K-C2{alpha} gene. Transfection of CHO-IR cells with two different antisense ODNs clearly reduced the protein content as well as mRNA levels of PI3K-C2{alpha} whereas neither the nonspecific mock- nor sense ODNs affected. The decrease of PI3K-C2{alpha} gene expression was paralleled by cellular changes indicating apoptotic cell death such as nuclear condensation, formation of apoptotic bodies, and DNA fragmentation. PI3K-C2{alpha} mRNA levels were also reduced when cells were incubated in growth factor-deficient medium. Supplementing growth factors (serum or insulin) into medium lead to an increase of PI3K-C2{alpha} mRNA levels. This finding strongly suggests that PI3K-C2{alpha} is a crucial survival factor.

  2. How Cells Integrate Complex Stimuli: The Effect of Feedback from Phosphoinositides and Cell Shape on Cell Polarization and Motility

    PubMed Central

    Edelstein-Keshet, Leah

    2012-01-01

    To regulate shape changes, motility and chemotaxis in eukaryotic cells, signal transduction pathways channel extracellular stimuli to the reorganization of the actin cytoskeleton. The complexity of such networks makes it difficult to understand the roles of individual components, let alone their interactions and multiple feedbacks within a given layer and between layers of signalling. Even more challenging is the question of if and how the shape of the cell affects and is affected by this internal spatiotemporal reorganization. Here we build on our previous 2D cell motility model where signalling from the Rho family GTPases (Cdc42, Rac, and Rho) was shown to organize the cell polarization, actin reorganization, shape change, and motility in simple gradients. We extend this work in two ways: First, we investigate the effects of the feedback between the phosphoinositides (PIs) , and Rho family GTPases. We show how that feedback increases heights and breadths of zones of Cdc42 activity, facilitating global communication between competing cell “fronts”. This hastens the commitment to a single lamellipodium initiated in response to multiple, complex, or rapidly changing stimuli. Second, we show how cell shape feeds back on internal distribution of GTPases. Constraints on chemical isocline curvature imposed by boundary conditions results in the fact that dynamic cell shape leads to faster biochemical redistribution when the cell is repolarized. Cells with frozen cytoskeleton, and static shapes, consequently respond more slowly to reorienting stimuli than cells with dynamic shape changes, the degree of the shape-induced effects being proportional to the extent of cell deformation. We explain these concepts in the context of several in silico experiments using our 2D computational cell model. PMID:22396633

  3. The Arabidopsis DREB2 genetic pathway is constitutively repressed by basal phosphoinositide-dependent phospholipase C coupled to diacylglycerol kinase

    PubMed Central

    Djafi, Nabila; Vergnolle, Chantal; Cantrel, Catherine; Wietrzyński, Wojciech; Delage, Elise; Cochet, Françoise; Puyaubert, Juliette; Soubigou-Taconnat, Ludivine; Gey, Delphine; Collin, Sylvie; Balzergue, Sandrine; Zachowski, Alain; Ruelland, Eric

    2013-01-01

    Phosphoinositide-dependent phospholipases C (PI-PLCs) are activated in response to various stimuli. They utilize substrates provided by type III-Phosphatidylinositol-4 kinases (PI4KIII) to produce inositol triphosphate and diacylglycerol (DAG) that is phosphorylated into phosphatidic acid (PA) by DAG-kinases (DGKs). The roles of PI4KIIIs, PI-PLCs, and DGKs in basal signaling are poorly understood. We investigated the control of gene expression by basal PI-PLC pathway in Arabidopsis thaliana suspension cells. A transcriptome-wide analysis allowed the identification of genes whose expression was altered by edelfosine, 30 ?M wortmannin, or R59022, inhibitors of PI-PLCs, PI4KIIIs, and DGKs, respectively. We found that a gene responsive to one of these molecules is more likely to be similarly regulated by the other two inhibitors. The common action of these agents is to inhibit PA formation, showing that basal PI-PLCs act, in part, on gene expression through their coupling to DGKs. Amongst the genes up-regulated in presence of the inhibitors, were some DREB2 genes, in suspension cells and in seedlings. The DREB2 genes encode transcription factors with major roles in responses to environmental stresses, including dehydration. They bind to C-repeat motifs, known as Drought-Responsive Elements that are indeed enriched in the promoters of genes up-regulated by PI-PLC pathway inhibitors. PA can also be produced by phospholipases D (PLDs). We show that the DREB2 genes that are up-regulated by PI-PLC inhibitors are positively or negatively regulated, or indifferent, to PLD basal activity. Our data show that the DREB2 genetic pathway is constitutively repressed in resting conditions and that DGK coupled to PI-PLC is active in this process, in suspension cells and seedlings. We discuss how this basal negative regulation of DREB2 genes is compatible with their stress-triggered positive regulation. PMID:23964284

  4. Increasing Plasma Membrane Phosphatidylinositol(4,5)Bisphosphate Biosynthesis Increases Phosphoinositide Metabolism in Nicotiana tabacum[W][OA

    PubMed Central

    Im, Yang Ju; Perera, Imara Y.; Brglez, Irena; Davis, Amanda J.; Stevenson-Paulik, Jill; Phillippy, Brian Q.; Johannes, Eva; Allen, Nina S.; Boss, Wendy F.

    2007-01-01

    A genetic approach was used to increase phosphatidylinositol(4,5)bisphosphate [PtdIns(4,5)P2] biosynthesis and test the hypothesis that PtdInsP kinase (PIPK) is flux limiting in the plant phosphoinositide (PI) pathway. Expressing human PIPKI? in tobacco (Nicotiana tabacum) cells increased plasma membrane PtdIns(4,5)P2 100-fold. In vivo studies revealed that the rate of 32Pi incorporation into whole-cell PtdIns(4,5)P2 increased >12-fold, and the ratio of [3H]PtdInsP2 to [3H]PtdInsP increased 6-fold, but PtdInsP levels did not decrease, indicating that PtdInsP biosynthesis was not limiting. Both [3H]inositol trisphosphate and [3H]inositol hexakisphosphate increased 3-and 1.5-fold, respectively, in the transgenic lines after 18 h of labeling. The inositol(1,4,5)trisphosphate [Ins(1,4,5)P3] binding assay showed that total cellular Ins(1,4,5)P3/g fresh weight was >40-fold higher in transgenic tobacco lines; however, even with this high steady state level of Ins(1,4,5)P3, the pathway was not saturated. Stimulating transgenic cells with hyperosmotic stress led to another 2-fold increase, suggesting that the transgenic cells were in a constant state of PI stimulation. Furthermore, expressing Hs PIPKI? increased sugar use and oxygen uptake. Our results demonstrate that PIPK is flux limiting and that this high rate of PI metabolism increased the energy demands in these cells. PMID:17496116

  5. Up-regulation of phosphoinositide metabolism in tobacco cells constitutively expressing the human type I inositol polyphosphate 5-phosphatase

    NASA Technical Reports Server (NTRS)

    Perera, Imara Y.; Love, John; Heilmann, Ingo; Thompson, William F.; Boss, Wendy F.; Brown, C. S. (Principal Investigator)

    2002-01-01

    To evaluate the impact of suppressing inositol 1,4,5-trisphosphate (InsP(3)) in plants, tobacco (Nicotiana tabacum) cells were transformed with the human type I inositol polyphosphate 5-phosphatase (InsP 5-ptase), an enzyme which specifically hydrolyzes InsP(3). The transgenic cell lines showed a 12- to 25-fold increase in InsP 5-ptase activity in vitro and a 60% to 80% reduction in basal InsP(3) compared with wild-type cells. Stimulation with Mas-7, a synthetic analog of the wasp venom peptide mastoparan, resulted in an approximately 2-fold increase in InsP(3) in both wild-type and transgenic cells. However, even with stimulation, InsP(3) levels in the transgenic cells did not reach wild-type basal values, suggesting that InsP(3) signaling is compromised. Analysis of whole-cell lipids indicated that phosphatidylinositol 4,5-bisphosphate (PtdInsP(2)), the lipid precursor of InsP(3), was greatly reduced in the transgenic cells. In vitro assays of enzymes involved in PtdInsP(2) metabolism showed that the activity of the PtdInsP(2)-hydrolyzing enzyme phospholipase C was not significantly altered in the transgenic cells. In contrast, the activity of the plasma membrane PtdInsP 5 kinase was increased by approximately 3-fold in the transgenic cells. In vivo labeling studies revealed a greater incorporation of (32)P into PtdInsP(2) in the transgenic cells compared with the wild type, indicating that the rate of PtdInsP(2) synthesis was increased. These studies show that the constitutive expression of the human type I InsP 5-ptase in tobacco cells leads to an up-regulation of the phosphoinositide pathway and highlight the importance of PtdInsP(2) synthesis as a regulatory step in this system.

  6. PTEN blocks insulin-mediated ETS-2 phosphorylation through MAP kinase, independently of the phosphoinositide 3-kinase pathway.

    PubMed

    Weng, Liang-Ping; Brown, Jessica L; Baker, Kim M; Ostrowski, Michael C; Eng, Charis

    2002-07-15

    The tumor suppressor PTEN possesses lipid and protein phosphatase activities. It has been well established that the lipid phosphatase activity is essential for its tumor-suppressive function via the phosphoinositide 3-kinase (PI3K) and Akt pathways. The precise role of the protein phosphatase activity is still unclear. In the current study, we demonstrate that overexpression of wild-type PTEN in the MCF-7 breast cancer line results in phosphatase activity-dependent decreases in the phosphorylation of ETS-2, which is a transcription factor whose DNA-binding ability is controlled by phosphorylation. Exposure of MCF-7 cells to insulin, insulin-like growth factor 1 (IGF-1) or epidermal growth factor (EGF) can lead to the phosphorylation of ETS-2, Akt and ERK1/2. The MEK inhibitor PD590089 abrogates insulin-stimulated phosphorylation of ETS-2. In contrast, the PI3K inhibitor LY492002 has no effect on insulin-stimulated phosphorylation of ETS-2, despite the fact that it diminishes insulin-stimulated phosphorylation of Akt. Interestingly, overexpression of PTEN in MCF-7 leads to blockade of insulin-stimulated, but not EGF-stimulated, phosphorylation of ERK, accompanied by dramatic decreases in ETS-2 phosphorylation. We further show that the relationship of PTEN and ETS-2 has functional significance by demonstrating that PTEN abrogates activation of the uPA Ras-responsive enhancer, a target of ETS-2 action, in a phosphatase-dependent manner, irrespective of the presence or absence of insulin. Our observations, therefore, suggest that PTEN blocks insulin-stimulated ETS-2 phosphorylation through inhibition of the ERK members of the MAP kinase family independently of PI3K, and that the PTEN effect on the phosphorylation status of ETS-2 may be mediated through PTEN's protein phosphatase activity. PMID:12095911

  7. Differential signaling by regulatory subunits of phosphoinositide-3-kinase influences cell survival in INS-1E insulinoma cells.

    PubMed

    Schrader, J; Niebel, P; Rossi, A; Archontidou-Aprin, E; Hörsch, D

    2015-02-01

    Class 1A phosphoinositide 3-kinase (PI3K) is essential for beta-cell growth and survival. Although PI3K has been studied extensively in diabetes the effect of alternatively spliced isoforms of the catalytic subunit p85? on beta cell proliferation and survival remains to be defined.We examined expression and signaling of alternatively spliced PI3K regulatory subunits p85?, p55? and p50? in insulinoma cells (INS-1E), an insulin-producing beta cell line. PI3K regulatory isoforms were knocked down by siRNA transfection or overexpressed by adenoviral gene delivery.Knockdown of p85? elevated PI3K activation determined by Akt phosphorylation at baseline and after stimulation with growth factors. In contrast, Akt phosphorylation was inhibited by overexpression of all isoforms of p85?. Correspondingly, p55? and p85? overexpression decreased downstream kinase GSK-3 phosphorylation as well, whereas p50? overexpression resulted in an activation of GSK-3. Moreover, overexpression of p50? and p85? lead to retinoblastoma protein hyperphosphorylation and S-phase entry. Upon challenge of INS-1E cells with a cytotoxic cytokine cocktail, levels of p85? were reduced and p50? was upregulated. Selective overexpression of p50? prevented cytokine induced apoptosis in INS-1E cells.In conclusion, signalling of p50?, p55? and p85? is similar at the level of Akt, but differentially influence downstream GSK-3 activation and cell cycle entry. PI3K isoform p50? induction by cytokines provides a link between regeneration and cell survival under cytotoxic stress in insulin-producing pancreatic beta-cells. PMID:25393342

  8. Leptin promotes invasiveness of kidney and colonic epithelial cells via phosphoinositide 3-kinase-, rho-, and rac-dependent signaling pathways.

    PubMed

    Attoub, S; Noe, V; Pirola, L; Bruyneel, E; Chastre, E; Mareel, M; Wymann, M P; Gespach, C

    2000-11-01

    Leptin plays a key role regulating food intake, body weight and fat mass. These critical parameters are associated with an increased risk for digestive and mammary gland cancer in the Western population. Here we determined whether leptin contributes to the invasive phenotype of colonic and kidney epithelial cells at various stages of the neoplastic progression. First, leptin potently (EC50 = 10-30 ng/ml) induces invasion of collagen gels by premalignant familial adenomatous colonic cells PC/AA/C1 and nontumorigenic MDCK kidney epithelial cells, their src-transformed counterparts, and the human adenocarcinoma colonic cells LoVo and HCT-8/S11. Leptin and its Ob-Rb receptors were consistently identified by RT-PCR and immunoblotting in these cell lines, as well as in human colonic epithelial crypts, polyps, colonic tumor resections, and adjacent mucosa. Leptin-induced invasion was effectively blocked by pharmacological inhibitors of several downstream signaling pathways involved in cell transformation, namely, JAK2 tyrosine kinase (AG490), phosphoinositide PI3'-kinase (wortmannin and LY294002), mTOR kinase (rapamycin), and protein kinases C (GF109203X, Gö6976). Accordingly, leptin induces transient elevation of the PI3'-kinase lipid products in JAK2 immunoprecipitates prepared from parental MDCK cells. The leptin effect on invasion was potentiated by the activated form of the small GTPase RhoA and was abrogated by dominant negative mutants of RhoA, Rac1, and the p110alpha of PI3'-K. Our data indicate that leptin may exert a local and beneficial effect on migration of normal colonic epithelial cells and reparation of the inflamed or wounded digestive mucosa. We also emphasize a new role for leptin, linking the nutritional and body fat status to digestive cancer susceptibility by stimulating the invasive capacity of colonic epithelial cells at early stages of neoplasia. This finding has potential clinical implications for colon cancer progression and management of obesity. PMID:11053255

  9. The p110? structure: mechanisms for selectivity and potency of new PI(3)K inhibitors

    Microsoft Academic Search

    Alex Berndt; Simon Miller; Olusegun Williams; Daniel D Le; Benjamin T Houseman; Joseph I Pacold; Fabrice Gorrec; Wai-Ching Hon; Pingda Ren; Yi Liu; Christian Rommel; Pascale Gaillard; Thomas Rückle; Matthias K Schwarz; Kevan M Shokat; Jeffrey P Shaw; Roger L Williams

    2010-01-01

    Deregulation of the phosphoinositide-3-OH kinase (PI(3)K) pathway has been implicated in numerous pathologies including cancer, diabetes, thrombosis, rheumatoid arthritis and asthma. Recently, small-molecule and ATP-competitive PI(3)K inhibitors with a wide range of selectivities have entered clinical development. In order to understand the mechanisms underlying the isoform selectivity of these inhibitors, we developed a new expression strategy that enabled us to

  10. The Growth Arrest-Specific Gene Product Gas6 Promotes the Survival of Human Oligodendrocytes via a Phosphatidylinositol 3-Kinase-Dependent Pathway

    Microsoft Academic Search

    Sai Latha Shankar; Kathleen O'Guin; Michael Cammer; F. Arthur McMorris; Trevor N. Stitt; Ross S. Basch; Brian Varnum; Bridget Shafit-Zagardo

    2003-01-01

    Microarray analysis revealed that transcripts for the Axl and Mer receptor tyrosine kinases are expressed at high levels in O4- immunopanned oligodendrocytes isolated from second trimester human fetal spinal cord. In humans the sole known ligand for the Axl\\/Rse\\/Mer kinases is growth arrest-specific gene 6 (Gas6), which in the CNS is secreted by neurons and endothelial cells. We hypothe- sized

  11. Requirement of Phosphatidylinositol 3-Kinase-Dependent Pathway and Src for Gas6-Axl Mitogenic and Survival Activities in NIH 3T3 Fibroblasts

    Microsoft Academic Search

    SANDRO GORUPPI; ELISABETTA RUARO; BRIAN VARNUM; CLAUDIO SCHNEIDER

    1997-01-01

    Gas6 is a secreted protein previously identified as the ligand of the Axl receptor tyrosine kinase. We have shown that Gas6 is able to induce cell cycle reentry of serum-starved NIH 3T3 cells and to efficiently prevent apoptosis after complete growth factor removal, a survival effect uncoupled from Gas6-induced mitogenesis. Here we report that the mitogenic effect of Gas6 requires

  12. Glucagon-like peptide-2-stimulated protein synthesis through the PI 3-kinase-dependent Akt-mTOR signaling pathway

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Glucagon-like peptide-2 (GLP-2) is a nutrient-responsive neuropeptide that exerts diverse actions in the gastrointestinal tract, including enhancing mucosal cell survival and proliferation. GLP-2 stimulates mucosal growth in vivo with an increased rate of protein synthesis. However, it was unclear w...

  13. A phosphatidylinositol transfer protein integrates phosphoinositide signaling with lipid droplet metabolism to regulate a developmental program of nutrient stress–induced membrane biogenesis

    PubMed Central

    Ren, Jihui; Pei-Chen Lin, Coney; Pathak, Manish C.; Temple, Brenda R. S.; Nile, Aaron H.; Mousley, Carl J.; Duncan, Mara C.; Eckert, Debra M.; Leiker, Thomas J.; Ivanova, Pavlina T.; Myers, David S.; Murphy, Robert C.; Brown, H. Alex; Verdaasdonk, Jolien; Bloom, Kerry S.; Ortlund, Eric A.; Neiman, Aaron M.; Bankaitis, Vytas A.

    2014-01-01

    Lipid droplet (LD) utilization is an important cellular activity that regulates energy balance and release of lipid second messengers. Because fatty acids exhibit both beneficial and toxic properties, their release from LDs must be controlled. Here we demonstrate that yeast Sfh3, an unusual Sec14-like phosphatidylinositol transfer protein, is an LD-associated protein that inhibits lipid mobilization from these particles. We further document a complex biochemical diversification of LDs during sporulation in which Sfh3 and select other LD proteins redistribute into discrete LD subpopulations. The data show that Sfh3 modulates the efficiency with which a neutral lipid hydrolase-rich LD subclass is consumed during biogenesis of specialized membrane envelopes that package replicated haploid meiotic genomes. These results present novel insights into the interface between phosphoinositide signaling and developmental regulation of LD metabolism and unveil meiosis-specific aspects of Sfh3 (and phosphoinositide) biology that are invisible to contemporary haploid-centric cell biological, proteomic, and functional genomics approaches. PMID:24403601

  14. Wortmannin inactivates phosphoinositide 3-kinase by covalent modification of Lys-802, a residue involved in the phosphate transfer reaction.

    PubMed Central

    Wymann, M P; Bulgarelli-Leva, G; Zvelebil, M J; Pirola, L; Vanhaesebroeck, B; Waterfield, M D; Panayotou, G

    1996-01-01

    Wortmannin at nanomolar concentrations is a potent and specific inhibitor of phosphoinositide (PI) 3-kinase and has been used extensively to demonstrate the role of this enzyme in diverse signal transduction processes. At higher concentrations, wortmannin inhibits the ataxia telangiectasia gene (ATM)-related DNA-dependent protein kinase (DNA-PKcs). We report here the identification of the site of interaction of wortmannin on the catalytic subunit of PI 3-kinase, p110alpha. At physiological pH (6.5 to 8) wortmannin reacted specifically with p110alpha. Phosphatidylinositol-4,5-diphosphate, ATP, and ATP analogs [adenine and 5'-(4-fluorosulfonylbenzoyl)adenine] competed effectively with wortmannin, while substances containing nucleophilic amino acid side chain functions had no effect at the same concentrations. This suggests that the wortmannin target site is localized in proximity to the substrate-binding site and that residues involved in wortmannin binding have an increased nucleophilicity because of their protein environment. Proteolytic fragments of wortmannin-treated, recombinant p110alpha were mapped with anti-wortmannin and anti-p110alpha peptide antibodies, thus limiting the target site within a 10-kDa fragment, colocalizing with the ATP-binding site. Site-directed mutagenesis of all candidate residues within this region showed that only the conservative Lys-802-to-Arg mutation abolished wortmannin binding. Inhibition of PI 3-kinase occurs, therefore, by the formation of an enamine following the attack of Lys-802 on the furan ring (at C-20) of wortmannin. The Lys-802-to-Arg mutant was also unable to bind FSBA and was catalytically inactive in lipid and protein kinase assays, indicating a crucial role for Lys-802 in the phosphotransfer reaction. In contrast, an Arg-916-to-Pro mutation abolished the catalytic activity whereas covalent wortmannin binding remained intact. Our results provide the basis for the design of novel and specific inhibitors of an enzyme family, including PI kinases and ATM-related genes, that play a central role in many physiological processes. PMID:8657148

  15. The Phosphoinositide 3-Kinase p110? Isoform Regulates Leukemia Inhibitory Factor Receptor Expression via c-Myc and miR-125b to Promote Cell Proliferation in Medulloblastoma

    PubMed Central

    von Bueren, André O.; ?wiek, Paulina; Rehrauer, Hubert; Djonov, Valentin; Anderle, Pascale; Arcaro, Alexandre

    2015-01-01

    Medulloblastoma (MB) is the most common malignant brain tumor in childhood and represents the main cause of cancer-related death in this age group. The phosphoinositide 3-kinase (PI3K) pathway has been shown to play an important role in the regulation of medulloblastoma cell survival and proliferation, but the molecular mechanisms and downstream effectors underlying PI3K signaling still remain elusive. The impact of RNA interference (RNAi)-mediated silencing of PI3K isoforms p110? and p110? on global gene expression was investigated by DNA microarray analysis in medulloblastoma cell lines. A subset of genes with selectively altered expression upon p110? silencing in comparison to silencing of the closely related p110? isoform was revealed. Among these genes, the leukemia inhibitory factor receptor ? (LIFR ?) was validated as a novel p110? target in medulloblastoma. A network involving c-Myc and miR-125b was shown to be involved in the control of LIFR? expression downstream of p110?. Targeting the LIFR? by RNAi, or by using neutralizing reagents impaired medulloblastoma cell proliferation in vitro and induced a tumor volume reduction in vivo. An analysis of primary tumors revealed that LIFR? and p110? expression were elevated in the sonic hedgehog (SHH) subgroup of medulloblastoma, indicating its clinical relevance. Together, these data reveal a novel molecular signaling network, in which PI3K isoform p110? controls the expression of LIFR? via c-Myc and miR-125b to promote MB cell proliferation. PMID:25915540

  16. Anti-tumor peptide AP25 decreases cyclin D1 expression and inhibits MGC-803 proliferation via phospho-extracellular signal-regulated kinase-, Src-, c-Jun N-terminal kinase- and phosphoinositide 3-kinase-associated pathways.

    PubMed

    Hu, Jialiang; Cheng, Tao; Zhang, Lijun; Sun, Beicheng; Deng, Lei; Xu, Hanmei

    2015-09-01

    The anti-tumor peptide AP25 is a prototype integrin antagonist, which exhibits anti?angiogenic and anti?tumor activity. The molecular mechanisms by which AP25 inhibits the growth of the MGC?803 gastric carcinoma cell line were investigated in the present study. K?ras specific RNA interference by lentiviral infection was successfully induced in MGC?803 cells [MGC?803 short hairpin (sh)RNA group] and the expression levels of K?ras, phosphorylated extracellular signal?regulated kinase (p-ERK) and cyclin D1 were observed to be markedly decreased. By contrast, AP25 caused cell cycle arrest of intact MGC?803 cells and decreased p?ERK and cyclin D1 expression levels. Of note, 0.4?3.2 µM AP25 no longer inhibited MGC?803 shRNA growth, indicating that AP25, at such concentrations, exerts its effect mainly through the Ras/Raf/mitogen-activated protein kinase kinase/ERK pathway, whereas at 25 µM, AP25 was able to inhibit MGC?803 shRNA growth. Chemical inhibitors of Src, c?Jun N?terminal kinase (JNK) and phosphoinositide 3?kinase (PI3K) were used to confirm that 25 µM AP25 inhibited growth of cells in the MGC?803 shRNA group and activated intracellular signaling pathways with Src, JNK and PI3K as key enzymes. In conclusion, the present study revealed the signal transduction pathways activated by AP25 at low (0.4?3.2 µM) or high (25 µM) concentrations. It also confirmed that integrins, when interacting with the freely moving ligand AP25 instead of immobilized extracellular matrix glycoproteins, are able to initiate cell signaling via similar pathways as in the latter case but with a reversed effect, to inhibit cell growth. PMID:26063313

  17. Involvement of p21racA, phosphoinositide 3-kinase, and vacuolar ATPase in phagocytosis of bacteria and erythrocytes by Entamoeba histolytica: suggestive evidence for coincidental evolution of amebic invasiveness.

    PubMed Central

    Ghosh, S K; Samuelson, J

    1997-01-01

    Trophozoites of Entamoeba histolytica, the protozoan parasite that causes amebic dysentery, phagocytose bacteria in the colonic lumen and erythrocytes (RBC) in host tissues. Because tissue invasion is an evolutionary dead end, it is likely that amebic pathogenicity is coincidentally selected, i.e., the same methods used to kill bacteria in the colonic lumen are used by parasites to damage host cells and cause disease. In support of this idea, the amebic lectin and pore-forming peptide are involved in binding and killing, respectively, bacteria and host epithelial cells. Here amebic phagocytosis of bacteria, RBC, and mucin-coated beads was disrupted by overexpression of E. histolytica p21(racA-V12), a ras-family protein involved in selection of sites of actin polymerization, which had been mutated to eliminate its GTPase activity. p21(racA-V12) transformants were also defective in capping and cytokinesis, while pinocytosis of fluorescent dextrans was not affected. Wortmannin, a fungal inhibitor of phosphoinositide 3-kinase, markedly inhibited phagocytosis of bacteria, RBC, and mucin-coated beads by wild-type amebae. In contrast to p21(racA-V12) overexpression, wortmannin abolished amebic pinocytosis of dextrans but had no inhibitory effects on capping. Inhibition of amebic vacuolar acidification by bafilomycin also decreased bacterial and RBC uptake. These results, which demonstrate similarities between mechanisms of phagocytosis of bacteria and RBC by amebae and macrophages, support the idea of coincidental selection of amebic genes encoding proteins that mediate destruction of host cells. PMID:9317033

  18. Sann-Joong-Kuey-Jian-Tang induces autophagy in HepG2 cells via regulation of the phosphoinositide-3 kinase/Akt/mammalian target of rapamycin and p38 mitogen-activated protein kinase pathways

    PubMed Central

    CHUANG, WAN-LING; SU, CHIN-CHENG; LIN, PING-YI; LIN, CHI-CHEN; CHEN, YAO-LI

    2015-01-01

    Sann-Joong-Kuey-Jian-Tang (SJKJT), a traditional Chinese medicine, was previously reported to induce autophagy and inhibit the proliferation of the human HepG2 hepatocellular carcinoma cell line via an extrinsic pathway. In the present study, the effects of SJKJT-induced autophagy and the cytotoxic mechanisms mediating these effects were investigated in HepG2 cells. The cytotoxicity of SJKJT in the HepG2 cells was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The results demonstrated that the half-maximal inhibitory concentration of SJKJT was 2.91 mg/ml at 24 h, 1.64 mg/ml at 48 h and 1.26 mg/ml at 72 h. The results of confocal fluorescence microscopy indicated that SJKJT resulted in the accumulation of green fluorescent protein-LC3 and vacuolation of the cytoplasm. Flow cytometric analysis revealed the accumulation of acidic vesicular organelles. Furthermore, western blot analysis, used to determine the expression levels of autophagy-associated proteins, demonstrated that the HepG2 cells treated with SJKJT exhibited LC3B-I/LC3B-II conversion, increased expression levels of Beclin, Atg-3 and Atg-5 and reduced expression levels of p62 and decreased signaling of the phosphoinositide-3 kinase/Akt/mammalian target of rapamycin and the p38 mitogen-activated protein kinase pathways. Taken together, these findings may assist in the development of novel chemotherapeutic agents for the treatment of malignant types of liver cancer. PMID:25847489

  19. Sann-Joong-Kuey-Jian-Tang induces autophagy in HepG2 cells via regulation of the phosphoinositide-3 kinase/Akt/mammalian target of rapamycin and p38 mitogen-activated protein kinase pathways.

    PubMed

    Chuang, Wan-Ling; Su, Chin-Cheng; Lin, Ping-Yi; Lin, Chi-Chen; Chen, Yao-Li

    2015-08-01

    Sann-Joong-Kuey-Jian?Tang (SJKJT), a traditional Chinese medicine, was previously reported to induce autophagy and inhibit the proliferation of the human HepG2 hepatocellular carcinoma cell line via an extrinsic pathway. In the present study, the effects of SJKJT?induced autophagy and the cytotoxic mechanisms mediating these effects were investigated in HepG2 cells. The cytotoxicity of SJKJT in the HepG2 cells was evaluated using a 3?(4,5?dimethylthiazol?2?yl)?2,5?diphenyltetrazolium bromide assay. The results demonstrated that the half?maximal inhibitory concentration of SJKJT was 2.91 mg/ml at 24 h, 1.64 mg/ml at 48 h and 1.26 mg/ml at 72 h. The results of confocal fluorescence microscopy indicated that SJKJT resulted in the accumulation of green fluorescent protein?LC3 and vacuolation of the cytoplasm. Flow cytometric analysis revealed the accumulation of acidic vesicular organelles. Furthermore, western blot analysis, used to determine the expression levels of autophagy?associated proteins, demonstrated that the HepG2 cells treated with SJKJT exhibited LC3B??/LC3B?? conversion, increased expression levels of Beclin, Atg?3 and Atg?5 and reduced expression levels of p62 and decreased signaling of the phosphoinositide?3 kinase/Akt/mammalian target of rapamycin and the p38 mitogen?activated protein kinase pathways. Taken together, these findings may assist in the development of novel chemotherapeutic agents for the treatment of malignant types of liver cancer. PMID:25847489

  20. A retinoic acid receptor ? agonist (CD2019) overcomes inhibition of axonal outgrowth via phosphoinositide 3-kinase signalling in the injured adult spinal cord

    PubMed Central

    Agudo, Marta; Yip, Ping; Davies, Meirion; Bradbury, Elizabeth; Doherty, Patrick; McMahon, Stephen; Maden, Malcolm; Corcoran, Jonathan P.T.

    2010-01-01

    After spinal cord injury in the adult mammal, axons do not normally regrow and this commonly leads to paralysis. Retinoic acid (RA) can stimulate neurite outgrowth in vitro of both the embryonic central and peripheral nervous system, via activation of the retinoic acid receptor (RAR) ?2. We show here that regions of the adult CNS, including the cerebellum and cerebral cortex, express RAR?2. We show that when cerebellar neurons are grown in the presence of myelin-associated glycoprotein (MAG) which inhibits neurite outgrowth, RAR? can be activated in a dose dependent manner by a RAR? agonist (CD2019) and neurite outgrowth can occur via phosphoinositide 3-kinase (PI3K) signalling. In a model of spinal cord injury CD2019 also acts through PI3K signalling to induce axonal outgrowth of descending corticospinal fibres and promote functional recovery. Our data suggest that RAR? agonists may be of therapeutic potential for human spinal cord injuries. PMID:19800972

  1. PDGF-induced receptor phosphorylation and phosphoinositide hydrolysis are unaffected by protein kinase C activation in mouse swiss 3T3 and human skin fibroblasts

    SciTech Connect

    Sturani, E.; Vicentini, L.M.; Zippel, R.; Toschi, L.; Pandiella-Alonso, A.; Comoglio, P.M.; Meldolesi, J.

    1986-05-29

    Short (1-10 min) pretreatment of intact cells with activators of protein kinase C (e.g. phorbol-12 myristate, 13-acetate, PMA) affects the activity of a variety of surface receptors (for growth factors, hormones and neurotransmitters), with inhibition of transmembrane signal generation. In two types of fibroblasts it is demonstrated that the PDGF receptor is unaffected by PMA. Exposure to PMA at concentrations up to 100 nM for 10 min failed to inhibit either one of the agonist-induced, receptor-coupled responses of PDGF: the autophosphorylation of receptor molecules at tyrosine residues, and the hydrolysis of membrane polyphosphoinositides. In contrast, the EGF receptor autophosphorylation (in A 431 cells) and the bombesin-induced phosphoinositide hydrolysis were readily inhibited by PMA.

  2. Lipid kinase and protein kinase activities of G-protein-coupled phosphoinositide 3-kinase gamma: structure-activity analysis and interactions with wortmannin.

    PubMed Central

    Stoyanova, S; Bulgarelli-Leva, G; Kirsch, C; Hanck, T; Klinger, R; Wetzker, R; Wymann, M P

    1997-01-01

    Signalling via seven transmembrane helix receptors can lead to a massive increase in cellular PtdIns(3,4,5)P3, which is critical for the induction of various cell responses and is likely to be produced by a trimeric G-protein-sensitive phosphoinositide 3-kinase (PI3Kgamma). We show here that PI3Kgamma is a bifunctional lipid kinase and protein kinase, and that both activities are inhibited by wortmannin at concentrations equal to those affecting the p85/p110alpha heterodimeric PI3K (IC50 approx. 2 nM). The binding of wortmannin to PI3Kgamma, as detected by anti-wortmannin antisera, closely followed the inhibition of the kinase activities. Truncation of more than the 98 N-terminal amino acid residues from PI3Kgamma produced proteins that were inactive in wortmannin binding and kinase assays. This suggests that regions apart from the core catalytic domain are important in catalysis and inhibitor interaction. The covalent reaction of wortmannin with PI3Kgamma was prevented by preincubation with phosphoinositides, ATP and its analogues adenine and 5'-(4-fluorosulphonylbenzoyl)adenine. Proteolytic analysis of wortmannin-prelabelled PI3Kgamma revealed candidate wortmannin-binding peptides around Lys-799. Replacement of Lys-799 by Arg through site-directed mutagenesis aborted the covalent reaction with wortmannin and the lipid kinase and protein kinase activities completely. The above illustrates that Lys-799 is crucial to the phosphate transfer reaction and wortmannin reactivity. Parallel inhibition of the PI3Kgamma-associated protein kinase and lipid kinase by wortmannin and by the Lys-799-->Arg mutation reveals that both activities are inherent in the PI3Kgamma polypeptide. PMID:9182708

  3. Phospholipase C Signaling Involvement in Macrotubule Assembly and Activation of the Mechanism Regulating Protoplast Volume in Plasmolyzed Root Cells of Triticum turgidum

    Microsoft Academic Search

    George Komis; Basil Galatis; Hartmut Quader; Dia Galanopoulou; Panagiotis Apostolakos

    2008-01-01

    The role of phosphoinositide-specific phospholipase C (PI-PLC) signaling in the macrotubule-dependent protoplast volume regulation in plasmolyzed root cells of Triticum turgidum was investigated. At the onset of hyperosmotic stress, PI-PLC activation was documented. Inhibition of PI-PLC activity by U73122 blocked tubulin macrotubule formation in plasmolyzed cells and their protoplast volume regulatory mechanism. In neomycin-treated plasmolyzed cells, macrotubule formation and protoplast

  4. Amplification of Chromosome 1q Genes Encoding the Phosphoinositide Signalling Enzymes PI4KB, AKT3, PIP5K1A and PI3KC2B in Breast Cancer

    PubMed Central

    Waugh, Mark G.

    2014-01-01

    Little is known about the possible oncogenic roles of genes encoding for the phosphatidylinositol 4-kinases, a family of enzymes that regulate an early step in phosphoinositide signalling. To address this issue, the mutational status of all four human phosphatidylinositol 4-kinases genes was analyzed across 852 breast cancer samples using the COSMIC data resource. Point mutations in the phosphatidylinositol 4-kinase genes were uncommon and appeared in less than 1% of the patient samples however, 62% of the tumours had increases in gene copy number for PI4KB which encodes the phosphatidylinositol 4-kinase IIIbeta isozyme. Extending this analysis to subsequent enzymes in the phosphoinositide signalling cascades revealed that the only PIP5K1A, PI3KC2B and AKT3 genes exhibited similar patterns of gene copy number variation. By comparison, gene copy number increases for established oncogenes such as EGFR and HER2/Neu were only evident in 20% of the samples. The PI4KB, PIP5K1A, PI3KC2B and AKT3 genes are related in that they all localize to chromosome 1q which is often structurally and numerically abnormal in breast cancer. These results demonstrate that a gene quartet encoding a potential phosphoinositide signalling pathway is amplified in a subset of breast cancers. PMID:25368680

  5. Ca2+ influx and phosphoinositide signalling are essential for the establishment and maintenance of cell polarity in monospores from the red alga Porphyra yezoensis

    PubMed Central

    Saga, Naotsune; Mikami, Koji

    2009-01-01

    The asymmetrical distribution of F-actin directed by cell polarity has been observed during the migration of monospores from the red alga Porphyra yezoensis. The significance of Ca2+ influx and phosphoinositide signalling during the formation of cell polarity in migrating monospores was analysed pharmacologically. The results indicate that the inhibition of the establishment of cell polarity, as judged by the ability of F-actin to localize asymmetrically, cell wall synthesis, and development into germlings, occurred when monospores were treated with inhibitors of the Ca2+ permeable channel, phospholipase C (PLC), diacylglycerol kinase, and inositol-1,4,5-trisphosphate receptor. Moreover, it was also found that light triggered the establishment of cell polarity via photosynthetic activity but not its direction, indicating that the Ca2+ influx and PLC activation required for the establishment of cell polarity are light dependent. By contrast, inhibition of phospholipase D (PLD) prevented the migration of monospores but not the asymmetrical localization of F-actin. Taken together, these findings suggest that there is functional diversity between the PLC and PLD signalling systems in terms of the formation of cell polarity; the former being critical for the light-dependent establishment of cell polarity and the latter playing a role in the maintenance of established cell polarity. PMID:19531546

  6. ROLE OF PHOSPHOINOSITIDE 3-KINASE ALPHA, PKC AND L-TYPE CALCIUM CHANNELS IN MEDIATING THE COMPLEX ACTIONS OF ANGIOTENSIN II ON MOUSE CARDIAC CONTRACTILITY

    PubMed Central

    Liang, Wenbin; Oudit, Gavin Y.; Patel, Mikin M.; Shah, Ajay M.; Woodgett, James R.; Tsushima, Robert G.; Ward, Michael E.; Backx, Peter H.

    2014-01-01

    Although angiotensin II (AngII) plays an important role in heart disease associated with pump dysfunction, its direct effects on cardiac pump function remain controversial. We found that, following AngII infusion, the developed pressure and +dP/dtmax in isolated Langendorff-perfused mouse hearts showed a complex temporal response, with a rapid transient decrease followed by an increase above baseline. Similar time-dependent changes in cell shortening and L-type Ca2+ currents were observed in isolated ventricular myocytes. Previous studies have established that AngII signaling involves phosphoinositide 3-kinases (PI3Ks). Dominant-negative inhibition of PI3K? in the myocardium selectively eliminated the rapid negative inotropic action of AngII while the loss of PI3K? had no effect on the response to AngII. Consistent with a link between PI3K? and PKC, PKC inhibition (with GF 109203X) reduced the negative inotropic effects of AngII by ~50%. Although both PI3K? and PKC activities are associated with glycogen synthase kinase-3? (GSK3?) and NADPH oxidase, genetic ablation of either GSK3? or p47phox (an essential subunit of NOX2-NADPH oxidase activity) had no effect on AngII’s inotropic actions. Our results establish that AngII has complex temporal effects on contractility and L-type Ca2+ channels in normal mouse myocardium with the negative inotropic effects requiring PI3K? and PKC activities. PMID:20696985

  7. Phosphorylation-independent dual-site binding of the FHA domain of KIF13 mediates phosphoinositide transport via centaurin [alpha]1

    SciTech Connect

    Tong, Yufeng; Tempel, Wolfram; Wang, Hui; Yamada, Kaori; Shen, Limin; Senisterra, Guillermo A.; MacKenzie, Farrell; Chishti, Athar H.; Park, Hee-Won (Toronto); (UICM)

    2011-11-07

    Phosphatidylinositol 3,4,5-triphosphate (PIP3) plays a key role in neuronal polarization and axon formation. PIP3-containing vesicles are transported to axon tips by the kinesin KIF13B via an adaptor protein, centaurin {alpha}1 (CENTA1). KIF13B interacts with CENTA1 through its forkhead-associated (FHA) domain. We solved the crystal structures of CENTA1 in ligand-free, KIF13B-FHA domain-bound, and PIP3 head group (IP4)-bound conformations, and the CENTA1/KIF13B-FHA/IP4 ternary complex. The first pleckstrin homology (PH) domain of CENTA1 specifically binds to PIP3, while the second binds to both PIP3 and phosphatidylinositol 3,4-biphosphate (PI(3,4)P2). The FHA domain of KIF13B interacts with the PH1 domain of one CENTA1 molecule and the ArfGAP domain of a second CENTA1 molecule in a threonine phosphorylation-independent fashion. We propose that full-length KIF13B and CENTA1 form heterotetramers that can bind four phosphoinositide molecules in the vesicle and transport it along the microtubule.

  8. Phosphoinositide 3-kinase gamma controls autonomic regulation of the mouse heart through Gi-independent downregulation of cAMP level.

    PubMed

    Alloatti, Giuseppe; Marcantoni, Andrea; Levi, Renzo; Gallo, Maria Pia; Del Sorbo, Lorenzo; Patrucco, Enrico; Barberis, Laura; Malan, Daniela; Azzolino, Ornella; Wymann, Matthias; Hirsch, Emilio; Montrucchio, Giuseppe

    2005-01-01

    Cardiac beta-adrenergic and the muscarinic receptors control contractility and heart rate by triggering multiple signaling events involving downstream targets like the phosphoinositide 3-kinase gamma (PI3Kgamma). We thus investigated whether the lack of PI3Kgamma could play a role in the autonomic regulation of the mouse heart. Contractility and ICaL of mutant cardiac preparations appeared increased in basal conditions and after beta-adrenergic stimulation. However, basal and beta-adrenergic stimulated heart rate were normal. Conversely, muscarinic inhibition of heart rate was reduced without alteration of the Gbetagamma-dependent stimulation of IK,ACh current. In addition, muscarinic-mediated anti-adrenergic effect on papillary muscle contractility and ICaL was significantly depressed. Consistently, cAMP level of PI3Kgamma-null ventricles was always higher than wild-type controls. Thus, PI3Kgamma controls the cardiac function by reducing cAMP concentration independently of Gi-mediated signaling. PMID:15620702

  9. Phosphatidic acid and phosphoinositides facilitate liposome association of Yas3p and potentiate derepression of ARE1 (alkane-responsive element one)-mediated transcription control.

    PubMed

    Kobayashi, Satoshi; Hirakawa, Kiyoshi; Horiuchi, Hiroyuki; Fukuda, Ryouichi; Ohta, Akinori

    2013-12-01

    In the n-alkane assimilating yeast Yarrowia lipolytica, the expression of ALK1, encoding a cytochrome P450 that catalyzes terminal mono-oxygenation of n-alkanes, is induced by n-alkanes. The transcription of ALK1 is regulated by a heterocomplex that comprises the basic helix-loop-helix transcription activators, Yas1p and Yas2p, and binds to alkane-responsive element 1 (ARE1) in the ALK1 promoter. An Opi1 family transcription repressor, Yas3p, represses transcription by binding to Yas2p. Yas3p localizes in the nucleus when Y. lipolytica is grown on glucose but localizes to the endoplasmic reticulum (ER) upon the addition of n-alkanes. In this study, we showed that recombinant Yas3p binds to the acidic phospholipids, phosphatidic acid (PA) and phosphoinositides (PIPs), in vitro. The ARE1-mediated transcription was enhanced in vivo in mutants defective in an ortholog of the Saccharomyces cerevisiae gene PAH1, encoding PA phosphatase, and in an ortholog of SAC1, encoding PIP phosphatase in the ER. Truncation mutation analyses for Yas3p revealed two regions that bound to PA and PIPs. These results suggest that the interaction with acidic phospholipids is important for the n-alkane-induced association of Yas3p with the ER membrane. PMID:24120453

  10. Expression pattern of class I phosphoinositide 3-kinase and distribution of its product, phosphatidylinositol-3,4,5-trisphosphate, during Drosophila embryogenesis.

    PubMed

    Xi, Xin; Tatei, Kazuaki; Kihara, Yumiko; Izumi, Takashi

    2014-07-01

    The class I phosphoinositide 3-kinase (PI3K) can be activated by a large variety of extracellular stimuli and is responsible for generating phosphatidylinositol-3,4,5-trisphosphate (PI(3,4,5)P(3)) from phosphatidylinositol-4,5-bisphosphate at the plasma membrane. The expression pattern of the class I PI3K and distribution of PI(3,4,5)P(3), visualized by its specific binding protein, GRP1-PH, were examined during Drosophila embryogenesis. We found that the RNA of Pi3K21B, encoding the Drosophila p60 regulatory subunit of the class I PI3Ks, was expressed maternally and expressed primarily in pole cells after cellularization until completion of germ band elongation. The RNA of Pi3K92E, encoding the Drosophila p110 catalytic subunit of the class I PI3Ks, was also expressed maternally. During gastrulation, its transcript level became lower and was slightly enriched in invaginating cells. Both Pi3K21B and Pi3K92E were expressed ubiquitously after germ band elongation and persisted during germ band shortening. PI(3,4,5)P(3) was distributed at the apical region of the invaginating cells during gastrulation. These findings suggest a possible involvement of class I PI3K and PI(3,4,5)P(3) in the regulation of invagination during gastrulation. PMID:24928809

  11. The inhibition of phosphoinositide synthesis and muscarinic-receptor-mediated phospholipase C activity by Li+ as secondary, selective, consequences of inositol depletion in 1321N1 cells.

    PubMed Central

    Batty, I H; Downes, C P

    1994-01-01

    Conditions are described for culture of 1321N1 cells under which cellular inositol is decreased from approximately 20 mM to < 0.5 mM but phosphoinositide concentrations are unaffected. The effects of the muscarinic-receptor agonist carbachol (1 mM) and/or LiCl (10 mM) on phosphoinositide turnover in these or in inositol-replete cells was examined after steady-state [3H]inositol labelling of phospholipid pools. In both inositol-replete and -depleted cells, carbachol stimulated similar initial (0-15 min) rates of phospholipase C (PLC) activity, in the presence of Li+. Subsequently (> 30-60 min) stimulated PLC activity and [3H]PtdIns concentrations declined dramatically only in depleted cells. In inositol-depleted cells, carbachol alone evoked increased concentrations of [3H]inositol, [3H]InsP1, [3H]InsP2, [3H]InsP3 and [3H]InsP4, which were largely sustained over 90 min, and concentrations of [3H]PtdIns, [3H]PtdInsP and [3H]PtdInsP2 were decreased only to approximately 82, 84 and 93% of control respectively. In the presence of Li+ in these cells, the stimulated rise in [3H]inositol was prevented and, although accumulation of [3H]InsP1, [3H]InsP2 and [3H]InsP3 was initially (0-30 min) potentiated, rates of accumulation of [3H]InsP1 and concentrations of [3H]polyphosphates later (> 30-60 min) declined, and concentrations of [3H]PtdIns, [3H]PtdInsP and [3H]PtdInsP2 were decreased respectively to approximately 39, 48 and 81% of control. After 60 min in the presence of both carbachol and Li+, stimulated PLC activity was decreased by approximately 70% compared with the initial rate in depleted cells. This decreased PLC activity was reflected by changes in the stimulated concentrations of [3H]Ins(1,3,4)P3 but not of [3H]Ins(1,4,5)P3, but effects of Li+ on the latter may have been obscured by the demonstrated, concomitant and equal stimulated accumulation of [3H]inositol 1:2cyclic,4,5-trisphosphate. These data suggest that receptor-mediated PLC activity is selectively impaired by Li+ as a secondary consequence of inositol monophosphatase inhibition in cells which are highly dependent on inositol re-cycling, but imply that, although Li+ attenuation of PLC activity correlates closely with parameters indicative of limiting inositol supply, it is not readily attributed to decreased PtdInsP2 availability. The potential for complex regulation of PLC and PtdIns synthase is discussed. PMID:8110190

  12. Agonist-induced desensitization of ATP receptor-mediated phosphoinositide turnover in C6 glioma cells: comparison with the negative-feedback regulation by protein kinase C.

    PubMed

    Lin, W W; Chuang, D M

    1993-07-01

    In C6 glioma cells, ATP increased 3H-inositol phosphate (IP) accumulation in a dose-dependent manner. Preincubation of cells with ATP (100 microM or 1 mM) resulted in a time-dependent loss of the ability of ATP to stimulate phosphoinositide (PI) hydrolysis. The agonist-induced desensitization of ATP-stimulated PI hydrolysis developed rapidly, and appeared to be independent on the activation of protein kinase C (PKC). Thus, PKC inhibitors (staurosporine, H-7 and polymyxin B), depletion of PKC and diacylglycerol (DG) kinase inhibitors (R59002, R59949) had no effect on the homologous desensitization. ATP-induced PI breakdown was inhibited by a 10 min pretreatment with the PKC activator, phorbol 12-myristate 13-acetate (PMA) or octylindolactam V, with a comparable IC50 of 5 nM, but was unaffected by the biologically inactive 4-alpha-phorbol 12,13-didecanoate (4 alpha-PDD). The inhibition caused by PMA and octylindolactam V was completely prevented by staurosporine (1 microM) and partially prevented by H-7 (300 microM), H-8 (300 microM) and polymyxin B (300 micrograms/ml). In addition, PKC activator-induced inhibition was unchanged after ATP pretreatment, but disappeared after PKC depletion. The IP formation elicited by NaF was inhibited by PMA and octylindolactam V with a comparable IC50 value of 7.5 nM while was unchanged after ATP pretreatment. These results indicate that ATP receptors are present in the C6 glioma cells, and that these receptors are coupled to PI turnover and undergo homologous desensitization. The agonist-induced desensitization, unlike the negative-feedback regulation caused by PMA and octylindolactam V, does not seem to involve PKC activation. PMID:8396484

  13. Dual Phosphoinositide 3-Kinase/Mammalian Target of Rapamycin Inhibitor NVP-BEZ235 Has a Therapeutic Potential and Sensitizes Cisplatin in Nasopharyngeal Carcinoma

    PubMed Central

    Deng, Rong; Wu, Xiao-Qi; Qin, Juan; Feng, Gong-Kan; Zhu, Xiao-Feng

    2013-01-01

    Phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin inhibitor (mTOR) pathway is often constitutively activated in human tumor cells and thus has been considered as a promising drug target. To ascertain a therapeutical approach of nasopharyngeal carcinoma (NPC), we hypothesized NVP-BEZ235, a novel and potent imidazo[4,5-c] quinolone derivative, that dually inhibits both PI3K and mTOR kinases activities, had antitumor activity in NPC. Expectedly, we found that NVP-BEZ235 selectively inhibited proliferation of NPC cells rather than normal nasopharyngeal cells using MTT assay. In NPC cell lines, with the extended exposure, NVP-BEZ235 selectively inhibited proliferation of NPC cells harboring PIK3CA mutation, compared to cells with wild-type PIK3CA. Furthermore, exposure of NPC cells to NVP-BEZ235 resulted in G1 growth arrest by Propidium iodide uptake assay, reduction of cyclin D1and CDK4, and increased levels of P27 and P21 by Western blotting, but negligible apoptosis. Moreover, we found that cisplatin (CDDP) activated PI3K/AKT and mTORC1 pathways and NVP-BEZ235 alleviated the activation by CDDP through dually targeting PI3K and mTOR kinases. Also, NVP-BEZ235 combining with CDDP synergistically inhibited proliferation and induced apoptosis in NPC cells. In CNE2 and HONE1 nude mice xenograft models, orally NVP-BEZ235 efficiently attenuated tumor growth with no obvious toxicity. In combination with NVP-BEZ235 and CDDP, there was dramatic synergy in shrinking tumor volumes and inducing apoptosis through increasing Noxa, Bax and decreasing Mcl-1, Bcl-2. Based on the above results, NVP-BEZ235, which has entered phase I/II clinical trials in patients with advanced solid tumors, has a potential as a monotherapy or in combination with CDDP for NPC treatment. PMID:23533654

  14. Genomic organization and complete cDNA sequence of the human phosphoinositide-specific phospholipase C {beta}3 gene (PLCB3)

    SciTech Connect

    Lagercrantz, J.; Carson, E.; Phelan, C. [Karolinska Hospital, Stockholm (Sweden)] [and others] [Karolinska Hospital, Stockholm (Sweden); and others

    1995-04-10

    We have characterized the complete cDNA sequence, genomic structure, and expression of the human phosphoinositide-specific phospholipase C {beta}3 (PLC {beta}3) gene (gene symbol PLCB3). PLC {beta}3 plays an important role in initiating receptor-mediated signal transduction. Activation of PLC takes place in many cells as a response to stimulation by hormones, growth factors, neurotransmitters, and other ligands. The partial cDNA sequence of PLC {beta}3, previously published, was extended with 876 bp in the 5{prime} direction, giving a transcript of 4400 bp and a total open reading frame of 1234 amino acids. This was in accordance with expression analysis by Northern blotting that revealed a single 4.4-kb transcript in all tissues tested. Genomic data were obtained by sequencing plasmid subclones of a cosmid that contained the whole gene. The size of the complete transcription unit was estimated to be on the order of 15 kb. The gene contains 31 exons, with all splice donor and acceptor sites conforming to the GT/AG rule. No exon exceeds 571 bp in length, and the shortest exon spans only 36 bp. More than half of the introns are smaller than 200 bp, with the smallest being only 79 bp long. The transcription initiation site was determined to be within an 8-bp cluster 328-321 bp upstream of the translation initiation site. The 5{prime} flanking region is highly GC rich, with multiple CpG doublets, and contains multiple binding sites for Sp1. Lacking typical transcriptional regulatory sequences such as TATA and CAAT boxes, the putative promoter region conforms to the group of housekeeping promoters. 28 refs., 4 figs., 1 tab.

  15. Synergistic inhibition of colon carcinoma cell growth by Hedgehog-Gli1 inhibitor arsenic trioxide and phosphoinositide 3-kinase inhibitor LY294002

    PubMed Central

    Cai, Xinyi; Yu, Kun; Zhang, Lijuan; Li, Yunfeng; Li, Qiang; Yang, Zhibin; Shen, Tao; Duan, Lincan; Xiong, Wei; Wang, Weiya

    2015-01-01

    The Hedgehog (Hh) signaling pathway not only plays important roles in embryogenesis and adult tissue homeostasis, but also in tumorigenesis. Aberrant Hh pathway activation has been reported in a variety of malignant tumors including colon carcinoma. Here, we sought to investigate the regulation of the Hh pathway transcription factor Gli1 by arsenic trioxide and phosphoinositide 3-kinase (PI3K) inhibitor LY294002 in colon carcinoma cells. We transfected cells with siGli1 and observed a significant reduction of Gli1 expression in HCT116 and HT29 cells, which was confirmed by quantitative real-time polymerase chain reaction and Western blots. Knocking down endogenous Gli1 reduced colon carcinoma cell viability through inducing cell apoptosis. Similarly, knocking down Gli2 using short interfering RNA impaired colon carcinoma cell growth in vitro. To elucidate the regulation of Gli1 expression, we found that both Gli inhibitor arsenic trioxide and PI3K inhibitor LY294002 significantly reduced Gli1 protein expression and colon carcinoma cell proliferation. Arsenic trioxide treatment also reduced Gli1 downstream target gene expression, such as Bcl2 and CCND1. More importantly, the inhibition of Hedgehog-Gli1 by arsenic trioxide showed synergistic anticancer effect with the PI3K inhibitor LY294002 in colon carcinoma cells. Our findings suggest that the Hh pathway transcription factor Gli1 is involved in the regulation of colon carcinoma cell viability. Inhibition of Hedgehog-Gli1 expression by arsenic trioxide and PI3K inhibitor synergistically reduces colon cancer cell proliferation, indicating that they could be used as an effective anti-colon cancer combination therapy. PMID:25945059

  16. Activation loop sequences confer substrate specificity to phosphoinositide 3-kinase alpha (PI3Kalpha ). Functions of lipid kinase-deficient PI3Kalpha in signaling.

    PubMed

    Pirola, L; Zvelebil, M J; Bulgarelli-Leva, G; Van Obberghen, E; Waterfield, M D; Wymann, M P

    2001-06-15

    Phosphoinositide 3-kinases (PI3Ks) are dual specificity lipid and protein kinases. While the lipid-dependent PI3K downstream signaling is well characterized, little is known about PI3K protein kinase signaling and structural determinants of lipid substrate specificity across the various PI3K classes. Here we show that sequences C-terminal to the PI3K ATP-binding site determine the lipid substrate specificity of the class IA PI3Kalpha (p85/p110alpha). Transfer of such activation loop sequences from class II PI3Ks, class III PI3Ks, and a related mammalian target of rapamycin (FRAP) into p110alpha turns the lipid substrate specificity of the resulting hybrid protein into that of the donor protein, while leaving the protein kinase activity unaffected. All resulting hybrids lacked the ability to produce phosphatidylinositol 3,4,5-trisphosphate in intact cells. Amino acid substitutions and structure modeling showed that two conserved positively charged (Lys and Arg) residues in the activation loop are crucial for the functionality of class I PI3Ks as phosphatidylinositol 4,5-bisphosphate kinases. By transient transfecion of 293 cells, we show that p110alpha hybrids, although unable to support lipid-dependent PI3K signaling, such as activation of protein kinase B/Akt and p70(S6k), retain the capability to associate with and phosphorylate insulin receptor substrate-1, with the same specificity and higher efficacy than wild type PI3Kalpha. Our data lay the basis for the understanding of the class I PI3K substrate selectivity and for the use of PI3Kalpha hybrids to dissect PI3Kalpha function as lipid and protein kinase. PMID:11278889

  17. The Na+/H+ Exchanger-3 (NHE3) Activity Requires Ezrin Binding to Phosphoinositide and Its Phosphorylation

    PubMed Central

    Babich, Victor; Di Sole, Francesca

    2015-01-01

    Na+/H+ exchanger-3 (NHE3) plays an essential role in maintaining sodium and fluid homeostasis in the intestine and kidney epithelium. Thus, NHE3 is highly regulated and its function depends on binding to multiple regulatory proteins. Ezrin complexed with NHE3 affects its activity via not well-defined mechanisms. This study investigates mechanisms by which ezrin regulates NHE3 activity in epithelial Opossum Kidney cells. Ezrin is activated sequentially by phosphatidylinositol-4,5-bisphosphate (PIP2) binding and phosphorylation of threonine 567. Expression of ezrin lacking PIP2 binding sites inhibited NHE3 activity (-40%) indicating that ezrin binding to PIP2 is required for preserving NHE3 activity. Expression of a phosphomimetic ezrin mutated at the PIP2 binding region was sufficient not only to reverse NHE3 activity to control levels but also to increase its activity (+80%) similar to that of the expression of ezrin carrying the phosphomimetic mutation alone. Calcineurin Homologous Protein-1 (CHP1) is part, with ezrin, of the NHE3 regulatory complex. CHP1-mediated activation of NHE3 activity was blocked by expression of an ezrin variant that could not be phosphorylated but not by an ezrin variant unable to bind PIP2. Thus, for NHE3 activity under baseline conditions not only ezrin phosphorylation, but also ezrin spatial-temporal targeting on the plasma membrane via PIP2 binding is required; however, phosphorylation of ezrin appears to overcome the control of NHE3 transport. CHP1 action on NHE3 activity is not contingent on ezrin binding to PIP2 but rather on ezrin phosphorylation. These findings are important in understanding the interrelation and dynamics of a CHP1-ezrin-NHE3 regulatory complex. PMID:26042733

  18. Biomaterials differentially regulate Src kinases and phosphoinositide 3-kinase-? in polymorphonuclear leukocyte primary and tertiary granule release.

    PubMed

    Cohen, Hannah Caitlin; Frost, Dustin C; Lieberthal, Tyler Jacob; Li, Lingjun; Kao, W John

    2015-05-01

    In the foreign body response, infiltrating PMNs exocytose granule subsets to influence subsequent downstream inflammatory and wound healing events. In previous studies, we found that PMNs cultured on poly(ethylene glycol) (PEG)-containing hydrogels (i.e., PEG and gelatin + PEG hydrogels) had enhanced primary granule release, yet similar tertiary granule release compared with PMNs cultured on polydimethylsiloxane or tissue culture polystyrene. PMN primary granules contain microbicidal proteins and proteases, which can potentially injure bystander cells, degrade the extracellular matrix, and promote inflammation. Here, we sought to understand the mechanism of the enhanced primary granule release from PMNs on PEG hydrogels. We found that primary granule release from PMNs on PEG hydrogels was adhesion mediated and involved Src family kinases and PI3K-?. The addition of gelatin to PEG hydrogels did not further enhance PMN primary granule release. Using stable-isotope dimethyl labeling-based shotgun proteomics, we identified many serum proteins - including Ig gamma constant chain region proteins and alpha-1-acid glycoprotein 1 - that were absorbed/adsorbed in higher quantities on PEG hydrogels than on TCPS, and may be involved in mediating PMN primary granule release. Ultimately, this mechanistic knowledge can be used to direct inflammation and wound healing following biomaterial implantation to promote a more favorable healing response. PMID:25736495

  19. Alpha 1-adrenergic receptor-mediated phosphoinositide hydrolysis and prostaglandin E2 formation in Madin-Darby canine kidney cells. Possible parallel activation of phospholipase C and phospholipase A2

    SciTech Connect

    Slivka, S.R.; Insel, P.A.

    1987-03-25

    alpha 1-Adrenergic receptors mediate two effects on phospholipid metabolism in Madin-Darby canine kidney (MDCK-D1) cells: hydrolysis of phosphoinositides and arachidonic acid release with generation of prostaglandin E2 (PGE2). The similarity in concentration dependence for the agonist (-)-epinephrine in eliciting these two responses implies that they are mediated by a single population of alpha 1-adrenergic receptors. However, we find that the kinetics of the two responses are quite different, PGE2 production occurring more rapidly and transiently than the hydrolysis of phosphoinositides. The antibiotic neomycin selectively decreases alpha 1-receptor-mediated phosphatidylinositol 4,5-bisphosphate hydrolysis without decreasing alpha 1-receptor-mediated arachidonic acid release and PGE2 generation. In addition, receptor-mediated inositol trisphosphate formation is independent of extracellular calcium, whereas release of labeled arachidonic acid is largely calcium-dependent. Moreover, based on studies obtained with labeled arachidonic acid, receptor-mediated generation of arachidonic acid cannot be accounted for by breakdown of phosphatidylinositol monophosphate, phosphatidylinositol bisphosphate, or phosphatidic acid. Further studies indicate that epinephrine produces changes in formation or turnover of several classes of membrane phospholipids in MDCK cells. We conclude that alpha 1-adrenergic receptors in MDCK cells appear to regulate phospholipid metabolism by the parallel activation of phospholipase C and phospholipase A2. This parallel activation of phospholipases contrasts with models described in other systems which imply sequential activation of phospholipase C and diacylglycerol lipase or phospholipase A2.

  20. The Essential Phosphoinositide Kinase MSS-4 Is Required for Polar Hyphal Morphogenesis, Localizing to Sites of Growth and Cell Fusion in Neurospora crassa

    PubMed Central

    Stenzel, Irene; Hempel, Franziska; Seiler, Stephan; Heilmann, Ingo

    2012-01-01

    Fungal hyphae and plant pollen tubes are among the most highly polarized cells known and pose extraordinary requirements on their cell polarity machinery. Cellular morphogenesis is driven through the phospholipid-dependent organization at the apical plasma membrane. We characterized the contribution of phosphoinositides (PIs) in hyphal growth of the filamentous ascomycete Neurospora crassa. MSS-4 is an essential gene and its deletion resulted in spherically growing cells that ultimately lyse. Two conditional mss-4-mutants exhibited altered hyphal morphology and aberrant branching at restrictive conditions that were complemented by expression of wild type MSS-4. Recombinant MSS-4 was characterized as a phosphatidylinositolmonophosphate-kinase phosphorylating phosphatidylinositol 4-phosphate (PtdIns4P) to phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2). PtdIns3P was also used as a substrate. Sequencing of two conditional mss-4 alleles identified a single substitution of a highly conserved Y750 to N. The biochemical characterization of recombinant protein variants revealed Y750 as critical for PI4P 5-kinase activity of MSS-4 and of plant PI4P 5-kinases. The conditional growth defects of mss-4 mutants were caused by severely reduced activity of MSS-4(Y750N), enabling the formation of only trace amounts of PtdIns(4,5)P2. In N. crassa hyphae, PtdIns(4,5)P2 localized predominantly in the plasma membrane of hyphae and along septa. Fluorescence-tagged MSS-4 formed a subapical collar at hyphal tips, localized to constricting septa and accumulated at contact points of fusing N. crassa germlings, indicating MSS-4 is responsible for the formation of relevant pools of PtdIns(4,5)P2 that control polar and directional growth and septation. N. crassa MSS-4 differs from yeast, plant and mammalian PI4P 5-kinases by containing additional protein domains. The N-terminal domain of N. crassa MSS-4 was required for correct membrane association. The data presented for N. crassa MSS-4 and its roles in hyphal growth are discussed with a comparative perspective on PI-control of polar tip growth in different organismic kingdoms. PMID:23272106

  1. Inhibition of 5-hydroxytryptamine-induced phosphoinositide hydrolysis and Ca2+ mobilization in canine cultured tracheal smooth muscle cells by phorbol ester.

    PubMed

    Yang, C M; Fen, L W; Tsao, H L; Chiu, C T

    1997-07-01

    1. Regulation of the increase in inositol-1,4,5-trisphosphate (IP3) production and intracellular Ca2+ concentration ([Ca2+]i by protein kinase C (PKC) was investigated in canine cultured tracheal smooth muscle cells (TSMCs). Stimulation of TSMCs by 5-hydroxytryptamine (5-HT) caused an initial transient [Ca2+]i peak followed by a sustained elevation of [Ca2+]i in a concentration-dependent manner. 2. Pretreatment of TSMCs with phorbol 12-myristate 13-acetate (PMA, 1 microM) for 30 min blocked the 5-HT-induced IP3 formation and Ca2+ mobilization. This inhibition was reduced after the cells had been incubated with PMA for 8 h, and within 48 h the 5-HT-induced Ca2+ mobilization reached the same extent as control cells. 3. The concentration of PMA that gave half-maximal inhibition of 5-HT-induced increase in [Ca2+]i was 4 nM. Pretreatment of TSMCs with staurosporine (1 microM) of GF109203X (0.1 microM), PKC inhibitors, inhibited the ability of PMA to attenuate 5-HT-induced responses, suggesting that the inhibitory effect of PMA was mediated through the activation of PKC. 4. In parallel with the effect of PMA on 5-HT-induced IP3 formation and Ca2+ mobilization, the translocation and down-regulation of PKC isozymes were determined by Western blot analysis in TSMCs. Analysis of cell extracts by Western blotting with antibodies against different PKC isozymes revealed that TSMCs expressed PKC-alpha, beta I, beta II, delta, epsilon, theta and zeta. With PMA treatment of the cells for various times, translocation of PKC-alpha, beta I, beta II, delta, epsilon, and theta from the cytosol to the membrane was seen after 5 min, 30 min, 2 h, and 4 h treatment. However, 24 h treatment caused a partial down-regulation of these PKC isozymes PKC-zeta was not significantly translocated and down-regulated at any of the times tested. 5. In conclusion, these results suggest that activation of PKC may inhibit the receptor-mediated phosphoinositide hydrolysis and consequently attenuate the [Ca2+]i increase or inhibit both responses independently. The translocation of PKC-alpha, beta I, beta II, delta, epsilon, and theta induced by PMA caused an attenuation of 5-HT-stimulated IP3 accumulation and Ca2+ mobilization in TSMCs. PMID:9222540

  2. Melatonin-mediated inhibition of Purkinje neuron P-type Ca˛? channels in vitro induces neuronal hyperexcitability through the phosphatidylinositol 3-kinase-dependent protein kinase C delta pathway.

    PubMed

    Zhang, Yuan; Li, Hua; Pu, Yicheng; Gong, Shan; Liu, Chunfeng; Jiang, Xinghong; Tao, Jin

    2015-04-01

    Although melatonin receptors are widely expressed in the mammalian central nervous system and peripheral tissues, there are limited data regarding the functions of melatonin in cerebellar Purkinje cells. Here, we identified a novel functional role of melatonin in modulating P-type Ca(2+) channels and action-potential firing in rat Purkinje neurons. Melatonin at 0.1 ?m reversibly decreased peak currents (I(Ba)) by 32.9%. This effect was melatonin receptor 1 (MT(R1)) dependent and was associated with a hyperpolarizing shift in the voltage dependence of inactivation. Pertussis toxin pretreatment, intracellular application of QEHA peptide, and a selective antibody raised against the G? subunit prevented the inhibitory effects of melatonin. Pretreatment with phosphatidylinositol 3-kinase (PI3K) inhibitors abolished the melatonin-induced decrease in I(Ba). Surprisingly, melatonin responses were not regulated by Akt, a common downstream target of PI3K. Melatonin treatment significantly increased protein kinase C (PKC) activity 2.1-fold. Antagonists of PKC, but not of protein kinase A, abolished the melatonin-induced decrease in I(Ba). Melatonin application increased the membrane abundance of PKC?, and PKC? inhibition (either pharmacologically or genetically) abolished the melatonin-induced IBa response. Functionally, melatonin increased spontaneous action-potential firing by 53.0%; knockdown of MT(R1) and blockade of P-type channels abolished this effect. Thus, our results suggest that melatonin inhibits P-type channels through MT(R1) activation, which is coupled sequentially to the ?? subunits of G(i/o)-protein and to downstream PI3K-dependent PKC? signaling. This likely contributes to its physiological functions, including spontaneous firing of cerebellar Purkinje neurons. PMID:25707622

  3. Physical process Mechanical mechanisms

    E-print Network

    Berlin,Technische Universität

    1 Physical process Generation · Mechanical mechanisms F = m·a · Electric/Magnetic mechanisms F = B·i·l · Fluid dynamic/Hydraulic mechanisms q, p, ij · Thermal/Optical #12;2 Source unit and source mechanisms ­ Monopoles......volume fluctuations ­ Dipoles ......pressure fluctuations

  4. Shear stress stimulates phosphorylation of eNOS at Ser(635) by a protein kinase A-dependent mechanism

    NASA Technical Reports Server (NTRS)

    Boo, Yong Chool; Hwang, Jinah; Sykes, Michelle; Michell, Belinda J.; Kemp, Bruce E.; Lum, Hazel; Jo, Hanjoong

    2002-01-01

    Shear stress stimulates nitric oxide (NO) production by phosphorylating endothelial NO synthase (eNOS) at Ser(1179) in a phosphoinositide-3-kinase (PI3K)- and protein kinase A (PKA)-dependent manner. The eNOS has additional potential phosphorylation sites, including Ser(116), Thr(497), and Ser(635). Here, we studied these potential phosphorylation sites in response to shear, vascular endothelial growth factor (VEGF), and 8-bromocAMP (8-BRcAMP) in bovine aortic endothelial cells (BAEC). All three stimuli induced phosphorylation of eNOS at Ser(635), which was consistently slower than that at Ser(1179). Thr(497) was rapidly dephosphorylated by 8-BRcAMP but not by shear and VEGF. None of the stimuli phosphorylated Ser(116). Whereas shear-stimulated Ser(635) phosphorylation was not affected by phosphoinositide-3-kinase inhibitors wortmannin and LY-294002, it was blocked by either treating the cells with a PKA inhibitor H89 or infecting them with a recombinant adenovirus-expressing PKA inhibitor. These results suggest that shear stress stimulates eNOS by two different mechanisms: 1) PKA- and PI3K-dependent and 2) PKA-dependent but PI3K-independent pathways. Phosphorylation of Ser(635) may play an important role in chronic regulation of eNOS in response to mechanical and humoral stimuli.

  5. Paclitaxel induces calcium oscillations via an inositol 1,4,5-trisphosphate receptor and neuronal calcium sensor 1-dependent mechanism

    PubMed Central

    Boehmerle, Wolfgang; Splittgerber, Ute; Lazarus, Michael B.; McKenzie, Kathleen M.; Johnston, David G.; Austin, David J.; Ehrlich, Barbara E.

    2006-01-01

    Taxol (Paclitaxel) is an important natural product for the treatment of solid tumors. Despite a well documented tubulin-stabilizing effect, many side effects of taxol therapy cannot be explained by cytoskeletal mechanisms. In the present study submicromolar concentrations of taxol, mimicking concentrations found in patients, induced cytosolic calcium (Ca2+) oscillations in a human neuronal cell line. These oscillations were independent of extracellular and mitochondrial Ca2+ but dependent on intact signaling via the phosphoinositide signaling pathway. We identified a taxol binding protein, neuronal Ca2+ sensor 1 (NCS-1), a Ca2+ binding protein that interacts with the inositol 1,4,5-trisphosphate receptor from a human brain cDNA phage display library. Taxol increased binding of NCS-1 to the inositol 1,4,5-trisphosphate receptor. Short hairpin RNA-mediated knockdown of NCS-1 in the same cell line abrogated the response to taxol but not to other agonists stimulating the phosphoinositide signaling pathway. These findings are important for studies involving taxol as a research tool in cell biology and may help to devise new strategies for the management of side effects induced by taxol therapy. PMID:17114292

  6. Efficacy, safety, pharmacokinetics and pharmacodynamics of SAR245409 (voxtalisib, XL765), an orally administered phosphoinositide 3-kinase/mammalian target of rapamycin inhibitor: a phase 1 expansion cohort in patients with relapsed or refractory lymphoma.

    PubMed

    Papadopoulos, Kyriakos P; Egile, Coumaran; Ruiz-Soto, Rodrigo; Jiang, Jason; Shi, Weiliang; Bentzien, Frauke; Rasco, Drew; Abrisqueta, Pau; Vose, Julie M; Tabernero, Josep

    2015-06-01

    The maximum tolerated dose of SAR245409 (voxtalisib), a pan-class I phosphoinositide 3-kinase (PI3K) and mammalian target of rapamycin (mTOR) inhibitor, was determined in a phase 1 dose-escalation study in advanced solid tumors. We report safety, pharmacokinetics (PK), pharmacodynamics and preliminary efficacy of SAR245409 capsules 50 mg twice daily in an expansion cohort of 16 patients with relapsed/refractory lymphoma. The most common treatment-related adverse events (AEs) were nausea (31.3%) and diarrhea (25.0%). The most common grade 3/4 treatment-related AE was increased alanine aminotransferase (12.5%). PK results were consistent with solid tumors, confirming a relatively short steady-state half-life (mean 4.61 h). Among 12 evaluable patients, one complete response and two partial responses were achieved in patients with and without PI3K/mTOR pathway alterations. In a patient with mantle cell lymphoma achieving PR, SAR245409 was associated with significant inhibition of PI3K/mTOR and extracellular signal-related kinase (ERK) pathways. Preliminary efficacy warrants further evaluation of SAR245409 in lymphoma. PMID:25300944

  7. Wnt1 Inducible Signaling Pathway Protein 1 (WISP1) Blocks Neurodegeneration through Phosphoinositide 3 Kinase/Akt1 and Apoptotic Mitochondrial Signaling Involving Bad, Bax, Bim, and Bcl-xL

    PubMed Central

    Wang, Shaohui; Chong, Zhao Zhong; Shang, Yan Chen; Maiese, Kenneth

    2012-01-01

    Wnt1 inducible signaling pathway protein 1 (WISP1) is a member of the CCN family of proteins that determine cell growth, cell differentiation, immune system activation, and cell survival in tissues ranging from the cardiovascular-pulmonary system to the reproductive system. Yet, little is known of the role of WISP1 as a neuroprotective entity in the nervous system. Here we demonstrate that WISP1 is present in primary hippocampal neurons during oxidant stress with oxygen-glucose deprivation (OGD). WISP1 expression is significantly enhanced during OGD exposure by the cysteine-rich glycosylated protein Wnt1. Similar to the neuroprotective capabilities known for Wnt1 and its signaling pathways, WISP1 averts neuronal cell injury and apoptotic degeneration during oxidative stress exposure. WISP1 requires activation of phosphoinositide 3-kinase (PI 3-K) and Akt1 pathways to promote neuronal cell survival, since blockade of these pathways abrogates cellular protection. Furthermore, WISP1 through PI 3-K and Akt1 phosphorylates Bad and GSK-3?, minimizes expression of the Bim/Bax complex while increasing the expression of Bcl-xL/Bax complex, and prevents mitochondrial membrane permeability, cytochrome c release, and caspase 3 activation in the presence of oxidant stress. These studies provide novel considerations for the development of WISP1 as an effective and robust therapeutic target not only for neurodegenerative disorders, but also for disease entities throughout the body. PMID:22272766

  8. CD28 Costimulation: A Source of Vav-1 for TCR Signaling with the Help of SLP-76?

    NSDL National Science Digital Library

    Frederique Michel (Institut Pasteur; Molecular Immunology Unit, Department of Immunology REV)

    2002-08-06

    T cells require dual stimulation to become activated. When T cells encounter antigen-presenting cells, both the T cell receptor (TCR) and the CD28 coreceptor are ligated and activated. Michel and Acuto discuss how the adaptor SLP-76, which is recruited to the activated TCR complex, and the Rho family guanosine triphosphatase exchanger Vav-1, which is recruited by the CD28 receptor and TCR, may form a macromolecular complex that results in T cells activation. Vav-1 may serve as a central integrator between CD28 signaling and TCR signaling through its indirect effects on phosphoinositide 3-kinase-dependent signaling.

  9. Ghrelin inhibit cell apoptosis in pancreatic ? cell line HIT-T15 via mitogen-activated protein kinase\\/phosphoinositide 3-kinase pathways

    Microsoft Academic Search

    Yuwei Zhang; Binwu Ying; LiXin Shi; Hong Fan; Dongmei Yang; Dan Xu; Yonggang Wei; Xiaobo Hu; Yonggang Zhang; Xiaohong Zhang; Tao Wang; Daishun Liu; Liyang Dou; Guo Chen; Fei Jiang; Fuqiang Wen

    2007-01-01

    Ghrelin stimulates growth hormone (GH) release and induces positive energy balances. Previous studies have reported that ghrelin inhibits apoptosis in several cell types but the precise underlying protective mechanisms in pancreatic ? cells are poorly understood. Therefore, we investigated which pathway was related with its anti-apoptotic effect in pancreatic ? cells. Exposure of HIT-T15 cells to ghrelin caused a rapid

  10. Atg21 Is a Phosphoinositide Binding Protein Required for Efficient Lipidation and Localization of Atg8 during Uptake of Aminopeptidase I by Selective Autophagy

    Microsoft Academic Search

    P. E. Stromhaug; Fulvio Reggiori; Ju Guan; Chao-Wen Wang; Daniel J. Klionsky

    2004-01-01

    Delivery of proteins and organelles to the vacuole by autophagy and the cytoplasm to vacuole targeting (Cvt) pathway involves novel rearrangements of membrane resulting in the formation of vesicles that fuse with the vacuole. The mechanism of vesicle formation and the origin of the membrane are complex issues still to be resolved. Atg18 and Atg21 are proteins essential to vesicle

  11. Phosphoinositide-specific Phospholipase C ? 1b (PI-PLC?1b) Interactome: Affinity Purification-Mass Spectrometry Analysis of PI-PLC?1b with Nuclear Protein*

    PubMed Central

    Piazzi, Manuela; Blalock, William L.; Bavelloni, Alberto; Faenza, Irene; D'Angelo, Antonietta; Maraldi, Nadir M.; Cocco, Lucio

    2013-01-01

    Two isoforms of inositide-dependent phospholipase C ?1 (PI-PLC?1) are generated by alternative splicing (PLC?1a and PLC?1b). Both isoforms are present within the nucleus, but in contrast to PLC?1a, the vast majority of PLC?1b is nuclear. In mouse erythroid leukemia cells, PI-PLC?1 is involved in the regulation of cell division and the balance between cell proliferation and differentiation. It has been demonstrated that nuclear localization is crucial for the enzymatic function of PI-PLC?1, although the mechanism by which this nuclear import occurs has never been fully characterized. The aim of this study was to characterize both the mechanism of nuclear localization and the molecular function of nuclear PI-PLC?1 by identifying its interactome in Friend's erythroleukemia isolated nuclei, utilizing a procedure that coupled immuno-affinity purification with tandem mass spectrometry analysis. Using this procedure, 160 proteins were demonstrated to be in association with PI-PLC?1b, some of which have been previously characterized, such as the splicing factor SRp20 (Srsf3) and Lamin B (Lmnb1). Co-immunoprecipitation analysis of selected proteins confirmed the data obtained via mass spectrometry. Of particular interest was the identification of the nuclear import proteins Kpna2, Kpna4, Kpnb1, Ran, and Rangap1, as well as factors involved in hematological malignancies and several anti-apoptotic proteins. These data give new insight into possible mechanisms of nuclear trafficking and functioning of this critical signaling molecule. PMID:23665500

  12. A Potent Inhibitor of Phosphoinositide 3-Kinase (PI3K) and Mitogen Activated Protein (MAP) Kinase Signalling, Quercetin (3, 3', 4', 5, 7-Pentahydroxyflavone) Promotes Cell Death in Ultraviolet (UV)-B-Irradiated B16F10 Melanoma Cells

    PubMed Central

    Rafiq, Rather A.; Quadri, Afnan; Nazir, Lone A.; Peerzada, Kaiser; Ganai, Bashir A.; Tasduq, Sheikh A.

    2015-01-01

    Ultraviolet (UV) radiation–induced skin damage contributes strongly to the formation of melanoma, a highly lethal form of skin cancer. Quercetin (Qu), the most widely consumed dietary bioflavonoid and well known inhibitor of phosphoinositide 3-kinase (PI3K) and mitogen activated protein (MAP) kinase signalling, has been reported to be chemopreventive in several forms of non-melanoma skin cancers. Here, we report that the treatment of ultraviolet (UV)-B-irradiated B16F10 melanoma cells with quercetin resulted in a dose dependent reduction in cell viability and increased apoptosis. The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (??M). Promotion of UVB-induced cell death by quercetin was further revealed by cleavage of chromosomal DNA, caspase activation, poly (ADP) ribose polymerase (PARP) cleavage, and an increase in sub-G1 cells. Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-?B nuclear translocation. Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF). Overall, these results suggest the possibility of using quercetin in combination with UVB as a possible treatment option for melanoma in future. PMID:26148186

  13. Cadmium?induced autophagy promotes survival of rat cerebral cortical neurons by activating class III phosphoinositide 3?kinase/beclin?1/B?cell lymphoma 2 signaling pathways.

    PubMed

    Wang, Qi-Wen; Wang, Yi; Wang, Tao; Zhang, Kang-Bao; Jiang, Cheng-Yang; Hu, Fei-Fei; Yuan, Yan; Bian, Jian-Chun; Liu, Xue-Zhong; Gu, Jian-Hong; Liu, Zong-Ping

    2015-08-01

    Autophagy is an evolutionarily conserved response that can be activated in response to heavy metal. Thus, the present study investigated the effect of autophagy on neurotoxic damage caused by cadmium (Cd) in rat cerebral cortical neurons. The results indicated that the viability of cortical neurons treated with Cd was markedly decreased in a dose? and time?dependent manner. The present study provided evidence that cortical neurons treated with Cd underwent autophagy: The conversion of microtubule?associated protein 1A/1B?light chain 3 (LC3) to LC3?II, an increase in the punctate distribution of endogenous LC3?II and the presence of autophagosomes were identified. Combined treatment with Cd and chloroquine, an autophagy inhibitor, reduced the amount of autophagocytosis and cell activity, whereas rapamycin, an autophagy inducer, reduced Cd?mediated cytotoxicity. Furthermore, it was found that beclin?1 and class III phosphoinositide 3 kinase (PI3K) levels were increased, while levels of the anti?apoptotic protein B?cell lymphoma 2 (Bcl?2) were decreased after Cd treatment. LY294002, a specific inhibitor of PI3K, prevented the decline in Bcl?2 production and the increase in levels of beclin?1, class III PI3K and autophagy following Cd treatment. In conclusion, the results of the present study suggested that Cd can induce cytoprotective autophagy by activating the class III PI3K/beclin?1/Bcl?2 signaling pathway, and that the autophagy pathway can serve as a sensitive biomarker for nervous system injury after exposure to Cd. PMID:25955216

  14. The Toll-like Receptor 9 Ligand, CpG Oligodeoxynucleotide, Attenuates Cardiac Dysfunction in Polymicrobial Sepsis, Involving Activation of Both Phosphoinositide 3 Kinase/Akt and Extracellular-Signal-Related Kinase Signaling

    PubMed Central

    Gao, Ming; Ha, Tuanzhu; Zhang, Xia; Wang, Xiaohui; Liu, Li; Kalbfleisch, John; Singh, Krishna; Williams, David; Li, Chuanfu

    2013-01-01

    Background.?Toll-like receptors (TLRs) play a role in the pathophysiology of sepsis and multiple organ failure. This study examined the effect of CpG oligodeoxynucleotide (CpG-ODN), the TLR9 ligand, on polymicrobial sepsis–induced cardiac dysfunction. Methods.?Male C57BL/6 mice were treated with CpG-ODN, control CpG-ODN (control-ODN), or inhibitory CpG-ODN (iCpG-ODN) 1 hour prior to cecal ligation and puncture (CLP)–induced sepsis. Mice that underwent sham surgery served as sham controls. Cardiac function was examined by echocardiography before and 6 hours after CLP. Results.?Cardiac function was significantly decreased 6 hours after CLP. CpG-ODN prevented CLP-induced cardiac dysfunction, as evidenced by maintenance of the ejection fraction and fractional shortening. Control-ODN or iCpG-ODN did not alter CLP-induced cardiac dysfunction. CpG-ODN significantly attenuated CLP-induced myocardial apoptosis and increased myocardial Akt and extracellular-signal-related kinase (ERK) phosphorylation levels following CLP. In vitro experiments demonstrated that CpG-ODN promotes an association between TLR9 and Ras, resulting in Akt and ERK phosphorylation. Inhibition of phosphoinositide 3-kinase (PI3K) by Ly294002 or inhibition of ERK by U0126 in vivo abolished CpG-ODN attenuation of CLP-induced cardiac dysfunction. Conclusions.?CpG-ODN prevents CLP-induced cardiac dysfunction, in part through activation of PI3K/Akt and ERK signaling. Modulation of TLR9 could be an effective approach for treatment of cardiovascular dysfunction in patients with sepsis or septic shock. PMID:23359590

  15. A Potent Inhibitor of Phosphoinositide 3-Kinase (PI3K) and Mitogen Activated Protein (MAP) Kinase Signalling, Quercetin (3, 3', 4', 5, 7-Pentahydroxyflavone) Promotes Cell Death in Ultraviolet (UV)-B-Irradiated B16F10 Melanoma Cells.

    PubMed

    Rafiq, Rather A; Quadri, Afnan; Nazir, Lone A; Peerzada, Kaiser; Ganai, Bashir A; Tasduq, Sheikh A

    2015-01-01

    Ultraviolet (UV) radiation-induced skin damage contributes strongly to the formation of melanoma, a highly lethal form of skin cancer. Quercetin (Qu), the most widely consumed dietary bioflavonoid and well known inhibitor of phosphoinositide 3-kinase (PI3K) and mitogen activated protein (MAP) kinase signalling, has been reported to be chemopreventive in several forms of non-melanoma skin cancers. Here, we report that the treatment of ultraviolet (UV)-B-irradiated B16F10 melanoma cells with quercetin resulted in a dose dependent reduction in cell viability and increased apoptosis. The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (??M). Promotion of UVB-induced cell death by quercetin was further revealed by cleavage of chromosomal DNA, caspase activation, poly (ADP) ribose polymerase (PARP) cleavage, and an increase in sub-G1 cells. Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-?B nuclear translocation. Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF). Overall, these results suggest the possibility of using quercetin in combination with UVB as a possible treatment option for melanoma in future. PMID:26148186

  16. Mechanisms of palmitate-induced cell death in human osteoblasts

    PubMed Central

    Gunaratnam, Krishanthi; Vidal, Christopher; Boadle, Ross; Thekkedam, Chris; Duque, Gustavo

    2013-01-01

    Summary Lipotoxicity is an overload of lipids in non-adipose tissues that affects function and induces cell death. Lipotoxicity has been demonstrated in bone cells in vitro using osteoblasts and adipocytes in coculture. In this condition, lipotoxicity was induced by high levels of saturated fatty acids (mostly palmitate) secreted by cultured adipocytes acting in a paracrine manner. In the present study, we aimed to identify the underlying mechanisms of lipotoxicity in human osteoblasts. Palmitate induced autophagy in cultured osteoblasts, which was preceded by the activation of autophagosomes that surround palmitate droplets. Palmitate also induced apoptosis though the activation of the Fas/Jun kinase (JNK) apoptotic pathway. In addition, osteoblasts could be protected from lipotoxicity by inhibiting autophagy with the phosphoinositide kinase inhibitor 3-methyladenine or by inhibiting apoptosis with the JNK inhibitor SP600125. In summary, we have identified two major molecular mechanisms of lipotoxicity in osteoblasts and in doing so we have identified a new potential therapeutic approach to prevent osteoblast dysfunction and death, which are common features of age-related bone loss and osteoporosis. PMID:24285710

  17. AMEAerospace & Mechanical

    E-print Network

    Wang, Hai

    AMEAerospace & Mechanical Engineering #12;Aerospace and Mechanical Engineers design complex mechanical, thermal, fluidic, acousti- cal, optical, and electronic systems, with char- acteristic sizes space. Aerospace and Mechanical Engineering (AME) students conduct basic and applied research within

  18. Investigation of molecular mechanisms and regulatory pathways of pro-angiogenic nanorods

    NASA Astrophysics Data System (ADS)

    Nethi, Susheel Kumar; Veeriah, Vimal; Barui, Ayan Kumar; Rajendran, Saranya; Mattapally, Saidulu; Misra, Sanjay; Chatterjee, Suvro; Patra, Chitta Ranjan

    2015-05-01

    Angiogenesis, a process involving the growth of new blood vessels from the pre-existing vasculature, plays a crucial role in various pathophysiological conditions. We have previously demonstrated that europium hydroxide [EuIII(OH)3] nanorods (EHNs) exhibit pro-angiogenic properties through the generation of reactive oxygen species (ROS) and mitogen activated protein kinase (MAPK) activation. Considering the enormous implication of angiogenesis in cardiovascular diseases (CVDs) and cancer, it is essential to understand in-depth molecular mechanisms and signaling pathways in order to develop the most efficient and effective alternative treatment strategy for CVDs. However, the exact underlying mechanism and cascade signaling pathways behind the pro-angiogenic properties exhibited by EHNs still remain unclear. Herein, we report for the first time that the hydrogen peroxide (H2O2), a redox signaling molecule, generated by these EHNs activates the endothelial nitric oxide synthase (eNOS) that promotes the nitric oxide (NO) production in a PI3K (phosphoinositide 3-kinase)/Akt dependent manner, eventually triggering angiogenesis. We intensely believe that the investigation and understanding of the in-depth molecular mechanism and signaling pathways of EHNs induced angiogenesis will help us in developing an effective alternative treatment strategy for cardiovascular related and ischemic diseases where angiogenesis plays an important role.Angiogenesis, a process involving the growth of new blood vessels from the pre-existing vasculature, plays a crucial role in various pathophysiological conditions. We have previously demonstrated that europium hydroxide [EuIII(OH)3] nanorods (EHNs) exhibit pro-angiogenic properties through the generation of reactive oxygen species (ROS) and mitogen activated protein kinase (MAPK) activation. Considering the enormous implication of angiogenesis in cardiovascular diseases (CVDs) and cancer, it is essential to understand in-depth molecular mechanisms and signaling pathways in order to develop the most efficient and effective alternative treatment strategy for CVDs. However, the exact underlying mechanism and cascade signaling pathways behind the pro-angiogenic properties exhibited by EHNs still remain unclear. Herein, we report for the first time that the hydrogen peroxide (H2O2), a redox signaling molecule, generated by these EHNs activates the endothelial nitric oxide synthase (eNOS) that promotes the nitric oxide (NO) production in a PI3K (phosphoinositide 3-kinase)/Akt dependent manner, eventually triggering angiogenesis. We intensely believe that the investigation and understanding of the in-depth molecular mechanism and signaling pathways of EHNs induced angiogenesis will help us in developing an effective alternative treatment strategy for cardiovascular related and ischemic diseases where angiogenesis plays an important role. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr01327e

  19. Puzzling Mechanisms

    ERIC Educational Resources Information Center

    van Deventer, M. Oskar

    2009-01-01

    The basis of a good mechanical puzzle is often a puzzling mechanism. This article will introduce some new puzzling mechanisms, like two knots that engage like gears, a chain whose links can be interchanged, and flat gears that do not come apart. It illustrates how puzzling mechanisms can be transformed into real mechanical puzzles, e.g., by…

  20. Bohmian Mechanics

    E-print Network

    Detlef Duerr; Sheldon Goldstein; Roderich Tumulka; Nino Zanghi

    2009-03-15

    Bohmian mechanics is a theory about point particles moving along trajectories. It has the property that in a world governed by Bohmian mechanics, observers see the same statistics for experimental results as predicted by quantum mechanics. Bohmian mechanics thus provides an explanation of quantum mechanics. Moreover, the Bohmian trajectories are defined in a non-conspiratorial way by a few simple laws.

  1. Plasma membrane nanoporation as a possible mechanism behind infrared excitation of cells

    NASA Astrophysics Data System (ADS)

    Beier, Hope T.; Tolstykh, Gleb P.; Musick, Joshua D.; Thomas, Robert J.; Ibey, Bennett L.

    2014-12-01

    Objective. Short infrared (IR) laser pulses have been used to stimulate action potentials in neurons both in vivo and in vitro. However, the mechanism(s) underlying this phenomenon has remained elusive. In vitro studies have found that pulsed IR exposure generates a nearly instant change in capacitance in the plasma membrane, characterized by inward rectification, a common feature in pore-forming exposures, such as electrical pulses and acoustic shock waves. Based on this similarity, we hypothesize that the mechanism of IR stimulation is the formation of short-lived nanopores in the plasma membrane. These transient, small-diameter pores allow the influx of extracellular ions that lead to action potential generation, possibly through activation of secondary messenger pathways or depolarization of the cell membrane resulting in activation of voltage-gated ion channels. Approach. A variety of fluorescent markers are used to observe the cell response to IR stimulation to monitor for effects indicative of nanoporation in other modalities. Main results. We observe rapid, transient rises in intracellular Ca2+, influx of YO-PRO-1 and propidium iodide into the cell signifying membrane permeabilization, cellular blebbing and swelling, and activation of the intracellular phosphoinositides lipid signaling pathway. Significance. This conclusion better explains the experimental observations and limitations of IR-induced neurological stimulation and represents a distinct theoretical shift in the understanding of the mechanism of IR-induced stimulation.

  2. Calcitriol mediates the activity of SGLT1 through an extranuclear initiated mechanism that involves intracellular signaling pathways.

    PubMed

    Castaneda-Sceppa, Carmen; Castaneda, Francisco

    2010-06-01

    The present study explored whether calcitriol plays a role in the regulation of sodium-dependent glucose transporter protein 1 (SGLT1) activity. For this purpose, alpha-methyl glucoside (AMG) uptake in stable transfected Chinese hamster ovary (CHO-G6D3) cells expressing rabbit SGLT1 (rbSGLT1) was used. The involvement of second messengers, intracellular signaling pathways, and pro-inflammatory cytokines were examined using specific inhibitors before incubation with calcitriol for 15 min. The present study demonstrated the involvement of second messengers produced by phospholipase A(2), phospholipase C, calmodulin, diacylglycerol kinase, and phosphoinositide 3 kinase on calcitriol-regulated AMG uptake. Pretreatment with inhibitors of the mitogen-activated protein kinase (MAPK) signaling pathway increased calcitriol-induced AMG uptake. In contrast, inhibition of the phosphoinositide 3-kinase PI3K/Akt/mTOR signaling pathway decreased the effect of calcitriol on AMG uptake. These findings suggest that calcitriol regulates rbSGLT1 activity through a rapid, extranuclear initiated mechanism of action stimulated by MAPK and inhibited by PI3K/Akt/mTOR. Another important finding was the effect of pro-inflammatory cytokines on calcitriol-induced AMG uptake. Interleukin-6 increased while tumor necrosis factor-alpha decreased calcitriol-induced AMG uptake. In conclusion, the present study demonstrates the involvement of calcitriol in the regulation of rbSGLT1 activity. This is due to the activation of intracellular signaling pathways triggered by second messenger molecules and cytokines after a short time (15 min) exposure to calcitriol. PMID:20428992

  3. Bohmian mechanics contradicts quantum mechanics

    E-print Network

    Neumaier, Arnold

    Bohmian mechanics contradicts quantum mechanics Arnold Neumaier Institut fur Mathematik, Universit://solon.cma.univie.ac.at/#24;neum/ Abstract. It is shown that, for a harmonic oscillator in the ground state, Bohmian mechanics and quantum mechanics predict values of opposite sign for certain time correlations. The discrepancy can

  4. Mesenchymal Stem Cells Alleviate Bacteria-Induced Liver Injury in Mice by Inducing Regulatory Dendritic Cells

    PubMed Central

    Zhang, Yi; Cai, Wei; Huang, Qingrong; Gu, Yuting; Shi, Yufang; Huang, Jiefang; Zhao, Fang; Liu, Qiang; Wei, Xunbin; Jin, Min; Wu, Changping; Xie, Qing; Zhang, Yi; Wan, Bing; Zhang, Yanyun

    2014-01-01

    Fulminant hepatic failure (FHF) is a clinical syndrome characterized by sudden and severe impairment of liver function. Mesenchymal stem cells (MSCs) have been proposed as a promising therapeutic approach for FHF. In this study we used Propionibacterium acnes (P. acnes)-primed, lipopolysaccharide (LPS)-induced liver injury in mice as an animal model of human FHF. We demonstrated that administration of MSCs significantly ameliorated liver injury and improved the survival rates of mice subjected to P. acnes plus LPS-induced FHF. Allogeneic MSCs showed similar treatment efficacy as autologous MSCs did in FHF. Treatment efficacy of MSCs could be attributed to decreased infiltration and activation of CD4+ T cells in the liver, inhibition of T helper 1 cells, and induction of regulatory T cells (Tregs). Moreover, decreased DNA copies of P. acnes were detected in the liver of MSC-treated mice. Intriguingly, a distinct liver population of CD11c+MHCIIhiCD80loCD86lo regulatory dendritic cells (DCs) was induced by MSCs. Moreover, these DCs induced Treg differentiation through transforming growth factor-? production. Further mechanistic studies demonstrated that MSC-derived prostaglandin E2 and one of its receptors, EP4, played essential roles in the differentiation of CD11c+B220? DC precursors into regulatory DCs in a phosphoinositide 3-kinase-dependent manner. Conclusion: MSCs induce regulatory DCs from CD11c+B220? DC precursors. This study elucidates an immunoregulatory mechanism of MSCs and lays a foundation for application of MSCs in FHF therapy. (Hepatology 2014;59:671–682) PMID:23929707

  5. Investigation of molecular mechanisms and regulatory pathways of pro-angiogenic nanorods.

    PubMed

    Nethi, Susheel Kumar; Veeriah, Vimal; Barui, Ayan Kumar; Rajendran, Saranya; Mattapally, Saidulu; Misra, Sanjay; Chatterjee, Suvro; Patra, Chitta Ranjan

    2015-05-21

    Angiogenesis, a process involving the growth of new blood vessels from the pre-existing vasculature, plays a crucial role in various pathophysiological conditions. We have previously demonstrated that europium hydroxide [Eu(III)(OH)3] nanorods (EHNs) exhibit pro-angiogenic properties through the generation of reactive oxygen species (ROS) and mitogen activated protein kinase (MAPK) activation. Considering the enormous implication of angiogenesis in cardiovascular diseases (CVDs) and cancer, it is essential to understand in-depth molecular mechanisms and signaling pathways in order to develop the most efficient and effective alternative treatment strategy for CVDs. However, the exact underlying mechanism and cascade signaling pathways behind the pro-angiogenic properties exhibited by EHNs still remain unclear. Herein, we report for the first time that the hydrogen peroxide (H2O2), a redox signaling molecule, generated by these EHNs activates the endothelial nitric oxide synthase (eNOS) that promotes the nitric oxide (NO) production in a PI3K (phosphoinositide 3-kinase)/Akt dependent manner, eventually triggering angiogenesis. We intensely believe that the investigation and understanding of the in-depth molecular mechanism and signaling pathways of EHNs induced angiogenesis will help us in developing an effective alternative treatment strategy for cardiovascular related and ischemic diseases where angiogenesis plays an important role. PMID:25963768

  6. Inhibition of mechanical stress-induced hypertrophic scar inflammation by emodin

    PubMed Central

    LIU, CHENG

    2015-01-01

    At least 50% of hypertrophic scarring (HS) is characterized by inflammation, for which there is currently no effective treatment available. Emodin is a major component of the widely used Chinese herb, rhubarb, which has been used to treat inflammation in several types of disease. However, few studies have investigated the efficacy of emodin in the treatment of HS. In the present study, a mouse model with mechanical stress-induced HS was used to investigate the effects of emodin (20, 40, 80, or 120 mg/ml) on HS, and to determine the potential underlying mechanisms. Treatment with emodin significantly attenuated HS inflammation, as determined by histopathological assessment of the scar elevation index, collagen structure and inflammation (P<0.05). Furthermore, treatment with emodin (40 mg/ml) markedly inhibited phosphoinositide 3-kinase (PI3K)/Akt activity (P<0.01) and this attenuation was associated with reduced expression levels of tumor necrosis factor-?, interleukin-6 and monocyte chemoattractant protein-1 (P<0.05) in the HS tissue. The results of the present study indicated that administration of emodin had therapeutic effects on the progression of HS and the underlying mechanism of this may be due to inhibition of the PI3K/Akt signaling pathway. PMID:25634255

  7. Fractofusion Mechanism

    Microsoft Academic Search

    Tatsuoki Takeda; Tomonori Takizuka

    1989-01-01

    With respect to cold fusion, a fractofusion mechanism is investigated. Deuterons are accelerated by the electric field generated between crack surfaces in a crystal. By assuming the possible magnitude of the potential difference, we calculate the fusion rate and energy multiplication factor. The results are consistent with those of the cold fusion experiments, and the fractofusion mechanism can explain the

  8. Cytoskeletal Mechanics

    NASA Astrophysics Data System (ADS)

    Mofrad, Mohammad R. K.; Kamm, Roger D.

    2006-10-01

    1. Introduction and the biological basis for cell mechanics Mohammad R. K. Mofrad and Roger Kamm; 2. Experimental measurements of intracellular mechanics Paul Janmey and Christoph Schmidt; 3. The cytoskeleton as a soft glassy material Jeffrey Fredberg and Ben Fabry; 4. Continuum elastic or viscoelastic models for the cell Mohammad R. K. Mofrad, Helene Karcher and Roger Kamm; 5. Multiphasic models of cell mechanics Farshid Guuilak, Mansoor A. Haider, Lori A. Setton, Tod A. Laursen and Frank P. T. Baaijens; 6. Models of cytoskeletal mechanics based on tensegrity Dimitrije Stamenovic; 7. Cells, gels and mechanics Gerald H. Pollack; 8. Polymer-based models of cytoskeletal networks F. C. MacKintosh; 9. Cell dynamics and the actin cytoskeleton James L. McGrath and C. Forbes Dewey, Jr; 10. Active cellular motion: continuum theories and models Marc Herant and Micah Dembo; 11. Summary Mohammad R. K. Mofrad and Roger Kamm.

  9. Cytoskeletal Mechanics

    NASA Astrophysics Data System (ADS)

    Mofrad, Mohammad R. K.; Kamm, Roger D.

    2011-08-01

    1. Introduction and the biological basis for cell mechanics Mohammad R. K. Mofrad and Roger Kamm; 2. Experimental measurements of intracellular mechanics Paul Janmey and Christoph Schmidt; 3. The cytoskeleton as a soft glassy material Jeffrey Fredberg and Ben Fabry; 4. Continuum elastic or viscoelastic models for the cell Mohammad R. K. Mofrad, Helene Karcher and Roger Kamm; 5. Multiphasic models of cell mechanics Farshid Guuilak, Mansoor A. Haider, Lori A. Setton, Tod A. Laursen and Frank P. T. Baaijens; 6. Models of cytoskeletal mechanics based on tensegrity Dimitrije Stamenovic; 7. Cells, gels and mechanics Gerald H. Pollack; 8. Polymer-based models of cytoskeletal networks F. C. MacKintosh; 9. Cell dynamics and the actin cytoskeleton James L. McGrath and C. Forbes Dewey, Jr; 10. Active cellular motion: continuum theories and models Marc Herant and Micah Dembo; 11. Summary Mohammad R. K. Mofrad and Roger Kamm.

  10. Tissue Mechanics

    NSDL National Science Digital Library

    Integrated Teaching and Learning Program,

    Students reflect on their experiences making silly putty (the previous hands-on activity in the unit), especially why changing the borax concentration alters the mechanical properties of silly putty and how this pertains to tissue mechanics. Students learn why engineers must understand tissue mechanics in order to design devices that will be implanted or used inside bodies, to study pathologies of tissues and how this alters tissue function, and to design prosthetics. Finally, students learn about collagen, elastin and proteoglycans and their roles in giving body tissues their unique functions. This prepares them for the culminating design-build-test activity of the unit.

  11. & Mechanical Engineering

    E-print Network

    Zhou, Chongwu

    control, emerging fuel cell technologies, computational uid mechanics, combustion, heat transfer-style competition racecars, and custom wave-tanks for testing surfboards. AME RESEARCH Our faculty, Aerotek, Boeing, Honeywell, Jet Propul- sion Laboratory, Lockheed Martin, NASA Facilities, Scaled

  12. Fault mechanics

    SciTech Connect

    Segall, P. (USAF, Geophysics Laboratory, Hanscom AFB, MA (United States))

    1991-01-01

    Recent observational, experimental, and theoretical modeling studies of fault mechanics are discussed in a critical review of U.S. research from the period 1987-1990. Topics examined include interseismic strain accumulation, coseismic deformation, postseismic deformation, and the earthquake cycle; long-term deformation; fault friction and the instability mechanism; pore pressure and normal stress effects; instability models; strain measurements prior to earthquakes; stochastic modeling of earthquakes; and deep-focus earthquakes. Maps, graphs, and a comprehensive bibliography are provided. 220 refs.

  13. Statistical Mechanics

    Microsoft Academic Search

    V. Jaksi ´; C.-A. Pillet

    1937-01-01

    We review and further develop a mathematical framework for non-equilibrium quantum statistical mechanics recently proposed in (JP4, JP5, JP6, Ru3, Ru4, Ru5, Ru6). In the alge- braic formalism of quantum statistical mechanics we introduce notions of non-equilibrium steady states, entropy production and heat fluxes, and study their properties. Our basic paradigm is a model of a small (finite) quantum system

  14. Mural propagation of descending vasa recta responses to mechanical stimulation

    PubMed Central

    Zhang, Zhong; Payne, Kristie; Cao, Chunhua

    2013-01-01

    To investigate the responses of descending vasa recta (DVR) to deformation of the abluminal surface, we devised an automated method that controls duration and frequency of stimulation by utilizing a stream of buffer from a micropipette. During stimulation at one end of the vessel, fluorescent responses from fluo4 or bis[1,3-dibutylbarbituric acid-(5)] trimethineoxonol [DiBAC4(3)], indicating cytoplasmic calcium ([Ca2+]CYT) or membrane potential, respectively, were recorded from distant cells. Alternately, membrane potential was recorded from DVR pericytes by nystatin whole cell patch-clamp. Mechanical stimulation elicited reversible [Ca2+]CYT responses that increased with frequency. Individual pericyte responses along the vessel were initiated within a fraction of a second of one another. Those responses were inhibited by gap junction blockade with 18 ?-glycyrrhetinic acid (100 ?M) or phosphoinositide 3 kinase inhibition with 2-morpholin-4-yl-8-phenylchromen-4-one (50 ?M). [Ca2+]CYT responses were blocked by removal of extracellular Ca2+ or L-type voltage-gated channel blockade with nifedipine (10 ?M). At concentrations selective for the T-type channel blockade, mibefradil (100 nM) was ineffective. During mechanostimulation, pericytes rapidly depolarized, as documented with either DiBAC4(3) fluorescence or patch-clamp recording. Single stimuli yielded depolarizations of 22.5 ± 2.2 mV while repetitive stimuli at 0.1 Hz depolarized pericytes by 44.2 ± 4.0 mV. We conclude that DVR are mechanosensitive and that rapid transmission of signals along the vessel axis requires participation of gap junctions, L-type Ca2+ channels, and pericyte depolarization. PMID:23698119

  15. Computational mechanics

    SciTech Connect

    Goudreau, G.L.

    1993-03-01

    The Computational Mechanics thrust area sponsors research into the underlying solid, structural and fluid mechanics and heat transfer necessary for the development of state-of-the-art general purpose computational software. The scale of computational capability spans office workstations, departmental computer servers, and Cray-class supercomputers. The DYNA, NIKE, and TOPAZ codes have achieved world fame through our broad collaborators program, in addition to their strong support of on-going Lawrence Livermore National Laboratory (LLNL) programs. Several technology transfer initiatives have been based on these established codes, teaming LLNL analysts and researchers with counterparts in industry, extending code capability to specific industrial interests of casting, metalforming, and automobile crash dynamics. The next-generation solid/structural mechanics code, ParaDyn, is targeted toward massively parallel computers, which will extend performance from gigaflop to teraflop power. Our work for FY-92 is described in the following eight articles: (1) Solution Strategies: New Approaches for Strongly Nonlinear Quasistatic Problems Using DYNA3D; (2) Enhanced Enforcement of Mechanical Contact: The Method of Augmented Lagrangians; (3) ParaDyn: New Generation Solid/Structural Mechanics Codes for Massively Parallel Processors; (4) Composite Damage Modeling; (5) HYDRA: A Parallel/Vector Flow Solver for Three-Dimensional, Transient, Incompressible Viscous How; (6) Development and Testing of the TRIM3D Radiation Heat Transfer Code; (7) A Methodology for Calculating the Seismic Response of Critical Structures; and (8) Reinforced Concrete Damage Modeling.

  16. Computational mechanics

    SciTech Connect

    Raboin, P J

    1998-01-01

    The Computational Mechanics thrust area is a vital and growing facet of the Mechanical Engineering Department at Lawrence Livermore National Laboratory (LLNL). This work supports the development of computational analysis tools in the areas of structural mechanics and heat transfer. Over 75 analysts depend on thrust area-supported software running on a variety of computing platforms to meet the demands of LLNL programs. Interactions with the Department of Defense (DOD) High Performance Computing and Modernization Program and the Defense Special Weapons Agency are of special importance as they support our ParaDyn project in its development of new parallel capabilities for DYNA3D. Working with DOD customers has been invaluable to driving this technology in directions mutually beneficial to the Department of Energy. Other projects associated with the Computational Mechanics thrust area include work with the Partnership for a New Generation Vehicle (PNGV) for ''Springback Predictability'' and with the Federal Aviation Administration (FAA) for the ''Development of Methodologies for Evaluating Containment and Mitigation of Uncontained Engine Debris.'' In this report for FY-97, there are five articles detailing three code development activities and two projects that synthesized new code capabilities with new analytic research in damage/failure and biomechanics. The article this year are: (1) Energy- and Momentum-Conserving Rigid-Body Contact for NIKE3D and DYNA3D; (2) Computational Modeling of Prosthetics: A New Approach to Implant Design; (3) Characterization of Laser-Induced Mechanical Failure Damage of Optical Components; (4) Parallel Algorithm Research for Solid Mechanics Applications Using Finite Element Analysis; and (5) An Accurate One-Step Elasto-Plasticity Algorithm for Shell Elements in DYNA3D.

  17. Mechanisms of Hypoxic Up-Regulation of Versican Gene Expression in Macrophages

    PubMed Central

    Sotoodehnejadnematalahi, Fattah; Staples, Karl J.; Chrysanthou, Elvina; Pearson, Helen; Ziegler-Heitbrock, Loems; Burke, Bernard

    2015-01-01

    Hypoxia is a hallmark of many pathological tissues. Macrophages accumulate in hypoxic sites and up-regulate a range of hypoxia-inducible genes. The matrix proteoglycan versican has been identified as one such gene, but the mechanisms responsible for hypoxic induction are not fully characterised. Here we investigate the up-regulation of versican by hypoxia in primary human monocyte-derived macrophages (HMDM), and, intriguingly, show that versican mRNA is up-regulated much more highly (>600 fold) by long term hypoxia (5 days) than by 1 day of hypoxia (48 fold). We report that versican mRNA decay rates are not affected by hypoxia, demonstrating that hypoxic induction of versican mRNA is mediated by increased transcription. Deletion analysis of the promoter identified two regions required for high level promoter activity of luciferase reporter constructs in human macrophages. The hypoxia-inducible transcription factor HIF-1 has previously been implicated as a key potential regulator of versican expression in hypoxia, however our data suggest that HIF-1 up-regulation is unlikely to be principally responsible for the high levels of induction observed in HMDM. Treatment of HMDM with two distinct specific inhibitors of Phosphoinositide 3-kinase (PI3K), LY290042 and wortmannin, significantly reduced induction of versican mRNA by hypoxia and provides evidence of a role for PI3K in hypoxic up-regulation of versican expression. PMID:26057378

  18. Why increased nuchal translucency is associated with congenital heart disease: a systematic review on genetic mechanisms.

    PubMed

    Burger, Nicole B; Bekker, Mireille N; de Groot, Christianne J M; Christoffels, Vincent M; Haak, Monique C

    2015-06-01

    This overview provides insight into the underlying genetic mechanism of the high incidence of cardiac defects in fetuses with increased nuchal translucency (NT). Nuchal edema, the morphological equivalent of increased NT, is likely to result from abnormal lymphatic development and is strongly related to cardiac defects. The underlying genetic pathways are, however, unknown. This study aims to present a systematic overview of genes involved in both cardiac and lymphatic development in mouse embryos. A search of PubMed and the Mammalian Phenotype Browser was performed. Fifteen candidate genes involved in both cardiac and lymphatic development were identified: Adrenomedullin; Chicken ovalbumin upstream promoter-transcription factor 2 (COUP-TFII); Cyp51; Ephrin-B2; Forkhead box protein C2 (Foxc2); Nuclear factor of activated T cells, cytoplasmic 1 (Nfatc1); Neurofibromatosis type 1 (Nf1); Phosphoinositide 3-kinase encoding isoform p110? (Pik3ca); Podoplanin; Prospero-related homeobox 1 (Prox1); T-box 1 (Tbx1); Tyrosine kinase with immunoglobulin-like and endothelial growth factor-like domains 1 (Tie1); vascular endothelial growth factor (Vegf)-A; Vegf receptor-3 (Vegfr-3); and Vascular endothelial zinc finger 1 (Vezf1). Mutations in all but one gene (Pik3ca) resulted in both a cardiac defect and nuchal edema. Candidate genes - mainly encoding for endothelium - are involved in both cardiac and lymphatic development. Alterations in candidate genes are associated with the strong relation between increased NT and cardiac defects. © 2015 John Wiley & Sons, Ltd. PMID:25728762

  19. ErbB4-neuregulin signaling modulates synapse development and dendritic arborization through distinct mechanisms.

    PubMed

    Krivosheya, Daria; Tapia, Lucia; Levinson, Joshua N; Huang, Kun; Kang, Yunhee; Hines, Rochelle; Ting, Annie K; Craig, Ann Marie; Mei, Lin; Bamji, Shernaz X; El-Husseini, Alaa

    2008-11-21

    Perturbations in neuregulin-1 (NRG1)/ErbB4 function have been associated with schizophrenia. Affected patients exhibit altered levels of these proteins and display hypofunction of glutamatergic synapses as well as altered neuronal circuitry. However, the role of NRG1/ErbB4 in regulating synapse maturation and neuronal process formation has not been extensively examined. Here we demonstrate that ErbB4 is expressed in inhibitory interneurons at both excitatory and inhibitory postsynaptic sites. Overexpression of ErbB4 postsynaptically enhances size but not number of presynaptic inputs. Conversely, knockdown of ErbB4 using shRNA decreases the size of presynaptic inputs, demonstrating a specific role for endogenous ErbB4 in synapse maturation. Using ErbB4 mutant constructs, we demonstrate that ErbB4-mediated synapse maturation requires its extracellular domain, whereas its tyrosine kinase activity is dispensable for this process. We also demonstrate that depletion of ErbB4 decreases the number of primary neurites and that stimulation of ErbB4 using a soluble form of NRG1 results in exuberant dendritic arborization through activation of the tyrosine kinase domain of ErbB4 and the phosphoinositide 3-kinase pathway. These findings demonstrate that NRG1/ErbB4 signaling differentially regulates synapse maturation and dendritic morphology via two distinct mechanisms involving trans-synaptic signaling and tyrosine kinase activity, respectively. PMID:18819924

  20. Quantum Mechanics

    NSDL National Science Digital Library

    De Raedt, Hans

    This website contains a number of descriptions of quantum mechanical phenomena, using 3D animations to illustrate the physics. The goal is to introduce basic concepts and phenomena using simulations rather than complex mathematics. The time-dependence of quantum systems is a focus of this material.

  1. Automotive Mechanics.

    ERIC Educational Resources Information Center

    Brown, Desmond

    This curriculum guide provides materials for a competency-based course in automotive mechanics at the secondary level. The curriculum design uses the curriculum infused model for the teaching of basic skills as part of vocational education and demonstrates the relationship of vocationally related skills to communication, mathematics, and science…

  2. Cratering mechanics

    NASA Technical Reports Server (NTRS)

    Ivanov, B. A.

    1986-01-01

    Main concepts and theoretical models which are used for studying the mechanics of cratering are discussed. Numerical two-dimensional calculations are made of explosions near a surface and high-speed impact. Models are given for the motion of a medium during cratering. Data from laboratory modeling are given. The effect of gravitational force and scales of cratering phenomena is analyzed.

  3. Mechanical allodynia.

    PubMed

    Lolignier, Stéphane; Eijkelkamp, Niels; Wood, John N

    2015-01-01

    Mechanical allodynia (other pain) is a painful sensation caused by innocuous stimuli like light touch. Unlike inflammatory hyperalgesia that has a protective role, allodynia has no obvious biological utility. Allodynia is associated with nerve damage in conditions such as diabetes, and is likely to become an increasing clinical problem. Unfortunately, the mechanistic basis of this enhanced sensitivity is incompletely understood. In this review, we describe evidence for the involvement of candidate mechanosensitive channels such as Piezo2 and their role in allodynia, as well as the peripheral and central nervous system mechanisms that have also been implicated in this form of pain. Specific treatments that block allodynia could be very useful if the cell and molecular basis of the condition could be determined. There are many potential mechanisms underlying this condition ranging from alterations in mechanotransduction and sensory neuron excitability to the actions of inflammatory mediators and wiring changes in the CNS. As with other pain conditions, it is likely that the range of redundant mechanisms that cause allodynia will make therapeutic intervention problematic. PMID:24846747

  4. Orbital mechanics

    Microsoft Academic Search

    Vladimir A. Chobotov

    1991-01-01

    The present work on the 'applied', or engineering-related aspects of orbital mechanics gives attention to the geographic and azimuth-elevation coordinate systems, as well as their transformations; the orbital parameters of a satellite; the universal approach to a body's position and velocity as a function of time, and geodetic and geocentric altitudes; and such issues in orbital maneuvering as the general

  5. Social Mechanisms

    Microsoft Academic Search

    Peter Hedström; Richard Swedberg

    1996-01-01

    In this article it is argued that the search for 'social mechanisms' is of crucial importance for the development of sociological theory. With this concept - which is occasionally used in the sociological literature but has received little systematic attention - attention is called to an intermediary level of analysis in-between pure description and story- telling, on the one hand,

  6. Structural Basis and Mechanism of Autoregulation in 3-Phosphoionsitide-Dependent Grp1 Family Arf GTPase Exchange Factors

    SciTech Connect

    DiNitto,J.; Delprato, A.; Lee, M.; Cronin, T.; Huang, S.; Guilherme, A.; Czech, M.; Lambright, D.

    2007-01-01

    Arf GTPases regulate membrane trafficking and actin dynamics. Grp1, ARNO, and Cytohesin-1 comprise a family of phosphoinositide-dependent Arf GTPase exchange factors with a Sec7-pleckstrin homology (PH) domain tandem. Here, we report that the exchange activity of the Sec7 domain is potently autoinhibited by conserved elements proximal to the PH domain. The crystal structure of the Grp1 Sec7-PH tandem reveals a pseudosubstrate mechanism of autoinhibition in which the linker region between domains and a C-terminal amphipathic helix physically block the docking sites for the switch regions of Arf GTPases. Mutations within either element result in partial or complete activation. Critical determinants of autoinhibition also contribute to insulin-stimulated plasma membrane recruitment. Autoinhibition can be largely reversed by binding of active Arf6 to Grp1 and by phosphorylation of tandem PKC sites in Cytohesin-1. These observations suggest that Grp1 family GEFs are autoregulated by mechanisms that depend on plasma membrane recruitment for activation.

  7. PI(4,5)P2-Mediated Cell Signaling: Emerging Principles and PTEN as a Paradigm for Regulatory Mechanism

    PubMed Central

    Gericke, Arne; Leslie, Nicholas R.; Lösche, Mathias; Ross, Alonzo H.

    2013-01-01

    PI(4,5)P2 (phosphatidylinositol 4,5-bisphosphate) is a relatively common anionic lipid that regulates cellular functions by multiple mechanisms. Hydrolysis of PI(4,5)P2 by phospholipase C yields inositol trisphosphate and diacylglycerol. Phosphorylation by phosphoinositide 3-kinase yields PI(3,4,5)P3, which is a potent signal for survival and proliferation. Also, PI(4,5)P2 can bind directly to integral and peripheral membrane proteins. As an example of regulation by PI(4,5)P2, we discuss phosphatase and tensin homologue deleted on chromosome 10 (PTEN) in detail. PTEN is an important tumor suppressor and hydrolyzes PI(3,4,5)P3. PI(4,5)P2 enhances PTEN association with the plasma membrane and activates its phosphatase activity. This is a critical regulatory mechanism, but a detailed description of this process from a structural point of view is lacking. The disordered lipid bilayer environment hinders structural determinations of membrane-bound PTEN. A new method to analyze membrane-bound protein measures neutron reflectivity for proteins bound to tethered phospholipid membranes. These methods allow determination of the orientation and shape of membrane-bound proteins. In combination with molecular dynamics simulations, these studies will provide crucial structural information that can serve as a foundation for our understanding of PTEN regulation in normal and pathological processes. PMID:23775692

  8. Mechanical capacitor

    NASA Technical Reports Server (NTRS)

    Kirk, J. A.; Studer, P. A.; Evans, H. E.

    1976-01-01

    A new energy storage system (the mechanical capacitor), using a spokeless magnetically levitated composite ring rotor, is described and design formulas for sizing the components are presented. This new system is configured around a permanent magnet (flux biased) suspension which has active servo control in the radial direction and passive control in the axial direction. The storage ring is used as a moving rotor and electronic commutation of the stationary armature coils is proposed. There is no mechanical contact with the rotating spokeless ring; therefore, long life and near zero rundown losses are projected. A 7-kW h system is sized to demonstrate feasibility. A literature review of flywheel energy storage systems is also presented and general formulas are developed for comparing rotor geometries.

  9. Venting mechanism

    SciTech Connect

    Kielar, S.J.; Rasmer, C.L.; Stimpson, F.F.

    1990-11-20

    This patent describes a venting mechanism for a drive axle assembly or the like. It comprises: a housing with an interior and an exterior, a gear assembly arranged in the housing, the housing being filled with a lubricant up to a predetermined level, and a flow of the lubricant being formed in the housing above the predetermined level when the gear assembly is operated, the venting mechanism comprising: a tube member mounted on the housing above the predetermined level and extending through a wall of the housing to allow for passage of air between the interior and the exterior of the housing, and a deflector vane pivotally attached at an open end of the tube member in the interior of the housing to prevent passage of lubricant out of the housing through the open end of the tube member.

  10. Mechanical clutch

    NASA Technical Reports Server (NTRS)

    Withey, Michael M. (Inventor); Lucas-Dean, Rob G. (Inventor)

    1995-01-01

    The present invention is directed to a mechanical clutch which limits transmission of torque to a desired, predetermined maximum torque from a first clutch plate to a second clutch plate. More specifically, the mechanical clutch includes at least one stepper member, preferably three or more evenly spaced stepper members, which transmit the torque from a first clutch plate to a second clutch plate providing a desired maximum torque is not exceeded. However, if the desired maximum torque is exceeded, the stepper member will rotate and move between the clutch plates so that the torque to the second clutch plate does not exceed the desired maximum torque. The desired maximum torque is set by the axial force compressing the stepper member between the clutch plates and when the applied torque to the first clutch plate exceeds the desired torque, the stepper member will rotate between the clutch plates rather than transmit that torque to the second clutch plate.

  11. Materials & Mechanics

    NSDL National Science Digital Library

    This page presents activities related to materials and mechanics. The lesson plans provided use ideas about building structure to strengthen understanding of larger science topics, the main one being force. Video clips and activities are provided on the topics of bending & torque, tension & compression and shear. Each activity includes a detailed description which will assist teachers in setting up these useful, practical in class lessons.

  12. Seabed mechanics

    SciTech Connect

    Denness, B.

    1985-01-01

    This volume is edited proceedings of the 1983 Seabed Mechanics Symposium, designed for geophysicists, geotechnical engineers, hydraulics engineers, sedimentologists and geologists who are engaged in or interested in activities on the seabed. The book draws on the collective experience of engineers and specialists from a variety of internationally recognized study centers. The topics range from studies of seabed sediments under load, sediment transportation, and hydrodynamics to seabed instrumentation, sampling techniques, and practical site investigations for offshore structures.

  13. Impact Mechanics

    NASA Astrophysics Data System (ADS)

    Stronge, W. J.

    2004-03-01

    Impact mechanics is concerned with the reaction forces that develop during a collision and the dynamic response of structures to these reaction forces. The subject has a wide range of engineering applications, from designing sports equipment to improving the crashworthiness of automobiles. This book develops several different methodologies for analysing collisions between structures. These range from rigid body theory for structures that are stiff and compact, to vibration and wave analyses for flexible structures. The emphasis is on low-speed impact where damage is local to the small region of contact between the colliding bodies. The analytical methods presented give results that are more robust or less sensitive to initial conditions than have been achieved hitherto. As a text, Impact Mechanics builds upon foundation courses in dynamics and strength of materials. It includes numerous industrially relevant examples and end-of-chapter homework problems drawn from industry and sports. Practising engineers will also find the methods presented in this book useful in calculating the response of a mechanical system to impact.

  14. The mechanism of mechanical alloying

    Microsoft Academic Search

    J. S. Benjamin; T. E. Volin

    1974-01-01

    The mechanical alloying process is a new method for producing composite metal powders with controlled microstructures. It\\u000a is unique in that it is an entirely solid state process, permitting dispersion of insoluble phases such as refractory oxides\\u000a and addition of reactive alloying elements such as aluminum and titanium. Interdispersion of the ingredients occurs by repeated\\u000a cold welding and fracture of

  15. Mechanics Mania

    NSDL National Science Digital Library

    Integrated Teaching and Learning Program,

    Through 10 lessons and numerous activities, students explore the natural universal rules engineers and physicists use to understand how things move and stay still. Together, these rules are called "mechanics." The study of mechanics is a way to improve our understanding of everyday movements, such as how gravity pulls things together, how objects balance, spin and twirl, and how things fly and fall. While studying Newton's three laws of motion, students gain hands-on experience with the concepts of forces, changes in motion, and action and reaction. Through hands-on activities, students model the behavior of parachutes and helicopters, closely examine falling objects, build and use a spring scale, examine collisions between skateboards, make model rockets with balloons and string, collect data from cotton ball catapults, study friction with small hovercrafts made from old CDs and balloons, experiment with center of mass by balancing objects on coat hangers and strings, compete to design clay beams with the best strength-to-weight ratio, experiment with weight distribution on homemade spinning tops, experiment with string length, weight and angle of release of pendulums made from fishing weights and string, and use marshmallows and spaghetti to construct their own structures to see which can hold the most weight. For each lesson, associated literacy activities provide additional student engagement. See the Unit Overview section for a list of topics by lesson and descriptions of the associated literacy activities.

  16. Mechanical & Aerospace Engineering

    E-print Network

    Mechanical & Aerospace Engineering An experimental methodology is presented for mechanism verification of physics-based prognosis of mechanical damage, such as fatigue. The proposed experimental methodology includes multi-resolution in-situ mechanical testing, advanced imaging analysis, and mechanism

  17. Department of Mechanical Engineering-

    E-print Network

    Endres. William J.

    Department of Mechanical Engineering- Engineering Mechanics Presidential Council of Alumnae Click #12;The Department of Mechanical Engineering ­ Engineering Mechanics honors its outstanding women Bachelor's of Science Degree in Mechanical Engineering in 1978. She earned her Master's of Science Degree

  18. Applied Mechanics

    NSDL National Science Digital Library

    This resource, created by National Aerospace Technical Education Center (SpaceTEC), is centered on applied mechanics. The introduction states that it should â??provide a basis for demonstrating a basic knowledge which will allow you to: interpret a basic drawing, produce a layout/template and fabricate a project.â?ť Topics in this PowerPoint presentation include: machine shop safety, hand tools, drills, measurement, hardware, materials, basic calculators, blueprint reading and the interpretation of technical drawings. The resource features a quiz, bibliography, a design handbook and a virtual machine shop. Overall, this is thorough presentation of this material. It totals nearly one-hundred twenty slides in length. Instructors could use this either as a presentation or simply to enhance existing curriculum.

  19. quantum mechanics

    PubMed Central

    Bender, Carl M; DeKieviet, Maarten; Klevansky, S. P.

    2013-01-01

    -symmetric quantum mechanics (PTQM) has become a hot area of research and investigation. Since its beginnings in 1998, there have been over 1000 published papers and more than 15 international conferences entirely devoted to this research topic. Originally, PTQM was studied at a highly mathematical level and the techniques of complex variables, asymptotics, differential equations and perturbation theory were used to understand the subtleties associated with the analytic continuation of eigenvalue problems. However, as experiments on -symmetric physical systems have been performed, a simple and beautiful physical picture has emerged, and a -symmetric system can be understood as one that has a balanced loss and gain. Furthermore, the phase transition can now be understood intuitively without resorting to sophisticated mathe- matics. Research on PTQM is following two different paths: at a fundamental level, physicists are attempting to understand the underlying mathematical structure of these theories with the long-range objective of applying the techniques of PTQM to understanding some of the outstanding problems in physics today, such as the nature of the Higgs particle, the properties of dark matter, the matter–antimatter asymmetry in the universe, neutrino oscillations and the cosmological constant; at an applied level, new kinds of -synthetic materials are being developed, and the phase transition is being observed in many physical contexts, such as lasers, optical wave guides, microwave cavities, superconducting wires and electronic circuits. The purpose of this Theme Issue is to acquaint the reader with the latest developments in PTQM. The articles in this volume are written in the style of mini-reviews and address diverse areas of the emerging and exciting new area of -symmetric quantum mechanics. PMID:23509390

  20. Applied quantum mechanics 1 Applied Quantum Mechanics

    E-print Network

    Levi, Anthony F. J.

    Applied quantum mechanics 1 Applied Quantum Mechanics Chapter 5 problems LAST NAME FIRST NAME #12 with the effective electron mass at the band edges. #12;Applied quantum mechanics 3 (c) Write a computer program

  1. Applied quantum mechanics 1 Applied Quantum Mechanics

    E-print Network

    Levi, Anthony F. J.

    Applied quantum mechanics 1 Applied Quantum Mechanics Chapter 8 problems LAST NAME FIRST NAME #12;Applied quantum mechanics 3 (b) If the electron is in a semiconductor and has an effective mass m * 0.07 m

  2. Applied quantum mechanics 1 Applied Quantum Mechanics

    E-print Network

    Levi, Anthony F. J.

    Applied quantum mechanics 1 Applied Quantum Mechanics Chapter 1 problems LAST NAME FIRST NAME #12 happens to the beat frequency if the airplane moves in an arc? #12;Applied quantum mechanics 3 Problem 1

  3. Applied quantum mechanics 1 Applied Quantum Mechanics

    E-print Network

    Levi, Anthony F. J.

    Applied quantum mechanics 1 Applied Quantum Mechanics Chapter 10 problems LAST NAME FIRST NAME #12 ­( ) L/( )= L/ #12;Applied quantum mechanics 3 (d) Use the results of (b) an (c) to draw the electron

  4. Fusogenic mechanisms.

    PubMed

    Lucy, J A

    1984-01-01

    Recent work is described on the fusion of human erythrocytes induced by chlorpromazine, which is accompanied by the endogenous proteolysis of integral and skeletal membrane proteins. This fusion reaction is of interest because it occurs at a high incidence in the presence of EGTA. It thus contrasts with the emphasis that has been placed on the role of Ca2+ in the fusion of lipid bilayers as a model for its behaviour in biomembrane fusion reactions. A survey of fusion in cells induced by chemicals and by Sendai virus, and of fusion in liposomes, in fact reveals numerous reported instances of fusion occurring in the absence of Ca2+. The finding that endogenous proteolysis of erythrocyte membrane proteins accompanies cell fusion induced by chlorpromazine leads to the suggestion that the products of proteolysis may in some cases be fusogenic, and that their formation may provide a general mechanism for the fusion of lipid bilayers in biomembrane fusion reactions, aspects of which are imitated by the fusogenic viruses. PMID:6423351

  5. Applied quantum mechanics 1 Applied Quantum Mechanics

    E-print Network

    Levi, Anthony F. J.

    Applied quantum mechanics 1 Applied Quantum Mechanics Chapter 6 problems LAST NAME FIRST NAME #12 --- and that for a Poisson distribution of such photons #12; 1 2 n ---------------- Applied quantum mechanics 3 (c) Apply conditions is the quantum mechanical result m t 2 2 d d x xd d V x ­= the same Newton's second law in which

  6. Quantum Mechanics II (Undergraduate)

    E-print Network

    Nickrent, Daniel L.

    Quantum Mechanics II (Undergraduate) Applications of Quantum Mechanics Spring, 2014 Physics 440 TEXTBOOK: Introduction to Quantum Mechanics (Second Edition), by David J. Griffiths, and QUNET's wikibook to apply quantum mechanics to some fundamental and important problems such as: better understanding

  7. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar James Bird Department of Mechanical Engineering Boston ­ are discussed. James Bird is an Assistant Professor in the Mechanical Engineering Department at Boston completed post-doctoral research at MIT. His research interests include experimental fluid mechanics

  8. Single Cell Mechanics BIOMATERIALS

    E-print Network

    Single Cell Mechanics BIOMATERIALS Our goal is to develop fundamental tools to measure the response of live cells to mechanical stimulation. The mechanisms by which cells convert mechanical forces evaluate the underlying mechanisms of cell mechanics. Objective Impact and Customers · Cancer, heart

  9. Zanthoxylum schinifolium leaf ethanol extract inhibits adipocyte differentiation through inactivation of the extracellular signal regulated kinase and phosphoinositide 3-kinase/Akt signaling pathways in 3T3-L1 pre-adipocytes.

    PubMed

    Choi, Eun-Ok; Park, Cheol; Shin, Soon Shik; Cho, Eun-Ju; Kim, Byung Woo; Hwang, Jin Ah; Hwang, Hye-Jin; Choi, Yung Hyun

    2015-07-01

    Zanthoxylum schinifolium is widely used as a food flavoring in east Asia. Although this plant has also been used in traditional oriental medicine for the treatment of the common cold, toothache, stomach ache, diarrhea and jaundice, its anti?obesity activity remains to be elucidated. The present study investigated the effects of ethanol extract from the leaves of Z. schinifolium (EEZS) on adipocyte differentiation, and its underlying mechanism, in 3T3?L1 pre?adipocytes. The results demonstrated that EEZS effectively suppressed intracellular lipid accumulation at non?toxic concentrations, and was associated with the downregulation of several adipocyte?specific transcription factors, including peroxisome proliferation?activity receptor ? (PPAR?), CCAAT/enhancer binding protein (C/EBP)? and C/EBP?, in a concentration?dependent manner. Furthermore, it was observed that EEZS markedly inactivated the extracellular signal?regulated protein kinase (ERK) and phosphatidylinositide 3?kinase (PI3K)/Akt pathways, which act upstream of PPAR? and C/EBPs in adipogenesis. These results suggested that EEZS inhibited lipid accumulation by downregulating the major transcription factors involved in the pathway of adipogenesis, including PPAR?, C/EBP? and C/EBP?, via regulation of the ERK and PI3K/Akt signaling pathways in 3T3?L1 adipocyte differentiation. This indicated the potential use of EEZS as an anti?obesity agent. PMID:25760758

  10. Applied quantum mechanics 1 Applied Quantum Mechanics

    E-print Network

    Levi, Anthony F. J.

    Applied quantum mechanics 1 Applied Quantum Mechanics Chapter 6 problems LAST NAME FIRST NAME #12 of the system. (b) Find . (c) Find and show that . Under what conditions is the quantum mechanical result( ) td d A t( ) t A td d A /= A B i 2 --- A^ B^,[ ] A^ B^ Et 2 --- n n 1 2 --- #12;Applied quantum

  11. Molecular mechanism of apoptosis induction in skin cancer cells by the centipedegrass extract

    PubMed Central

    2013-01-01

    Background Centipedegrass extract (CGE) is mainly composed of maysin and its derivatives, which are recognized internationally as natural compounds. Compared to other flavonoids, maysin has a unique structure in that mannose is bound to the flavonoid backbone. CGE exhibits some biological properties in that it can function as an anti-oxidant, anti-inflammatory, anti-adipogenic, and insecticidal. Whether CGE has other biological functions, such as anti-cancer activity, is unknown. Methods B16F1 (mouse) and SKMEL-5 (human) cells were treated with CGE, and their subsequent survival was determined using MTT assay. We performed a cell cycle analysis using propidium iodide (PI), and detected apoptosis using double staining with annexin V-FITC/PI. In addition, we examined mitochondrial membrane potentials using flow cytometry, as well as signaling mechanisms with an immunoblotting analysis. Results CGE inhibited skin cancer cell growth by arresting the cell cycle in the G2/M phase, and increased both early and late apoptotic cell populations without affecting normal cells. Furthermore, we observed mitochondrial transmembrane depolarization, increased cytochrome-c release, caspase-3 and caspase-7 activation, and increased poly ADP-ribose polymerase degradation. CGE also downregulated activation of p-AKT, p-glycogen synthase kinase-3? (GSK-3?), and p-BAD in a time-dependent manner. LY294002 inhibition of phosphoinositide 3-kinase (PI3K) significantly sensitized skin cancer cells, which led to an increase in CGE-induced apoptosis. Conclusions CGE controlled skin cancer cell growth by inhibiting the PI3K/AKT/GSK-3? signaling pathway and activating the effector caspases. This study is the first to demonstrate anti-cancer properties for CGE, and that CGE may be an effective therapeutic agent for treating skin cancer. PMID:24325618

  12. Mechanical systems: A compilation

    NASA Technical Reports Server (NTRS)

    1975-01-01

    A compilation of several mechanized systems is presented. The articles are contained in three sections: robotics, industrial mechanical systems, including several on linear and rotary systems and lastly mechanical control systems, such as brakes and clutches.

  13. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar Katia Bertoldi Harvard University Soft materials in response to diverse stimuli. While the mechanical attributes - such as energy absorption, stiffness and switchable functionalities. Katia Bertoldi is an Assistant Professor of Applied Mechanics at Harvard

  14. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar Sankha Bhowmick Department of Mechanical Engineering comes from nature, where anhydrobiotic organisms survive extreme drought by moving to metabolic stasis of desiccation protocols. Prof. Bhowmick is an associate professor in Mechanical Engineering at University

  15. MECHANICAL ENGINEERING UNDERGRADUATE MAJOR

    E-print Network

    HANDBOOK FOR MECHANICAL ENGINEERING UNDERGRADUATE MAJOR Old Dominion University Department of Mechanical Engineering Batten College of Engineering and Technology Norfolk, Virginia 23529-0247 #12;TABLE OF CONTENTS MECHANICAL ENGINEERING HANDBOOK

  16. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar Robert J. Hannemann The Gordon Institute and the Department of Mechanical Engineering Tufts University Retooling Our Energy Ecosystem: challenges and Chair of the Tufts Department of Mechanical Engineering. His technical and academic interests

  17. MECHANICAL ENGINEERING Ross Schlueter

    E-print Network

    MECHANICAL ENGINEERING Ross Schlueter Engineering Deputy For Mechanical Engineering Russ Wells Mechanical Engineering Department Deputy ELECTRONICS, SOFTWARE & INSTRUMENTATION ENGINEERING Henrik von Der Lippe Deputy ENGINEERING OPERATIONS John Freeman Deputy Project Management & Controls, Finances

  18. Applied quantum mechanics 1 Applied Quantum Mechanics

    E-print Network

    Levi, Anthony F. J.

    Applied quantum mechanics 1 Applied Quantum Mechanics Chapter 5 problems LAST NAME FIRST NAME #12 + --------------------------------------------- k = t 10/= t 1­= Ek 2t kxL( ) 2t 2kxL( )cos+cos= t 10/= t 1­= t 0.2­= #12;Applied quantum mechanics 3 (c) Write a computer program to plot the electron density of states for a square lat- tice

  19. Quantum Mechanics + Open Systems

    E-print Network

    Steinhoff, Heinz-Jürgen

    Quantum Mechanics + Open Systems = Thermodynamics ? Jochen Gemmer T¨ubingen, 09.02.2006 #12., World Scientific) #12;Fundamental Law or Emergent Description? Quantum Mechanics i t = (- 2 2m + V or Emergent Description? Quantum Mechanics i t = (- 2 2m + V ) "Heisenberg Cut" Classical Mechanics: m d2

  20. Mechanical Engineer Company Description

    E-print Network

    Kostic, Milivoje M.

    Mechanical Engineer Company Description Control Solutions Inc. is a small, dynamic, and rapidly. Position Description The Mechanical Engineer is responsible for all aspects associated with the mechanicalE Mechanism, when applicable. · Perform static, dynamic, vibration, thermal, and other engineering analysis

  1. Mechanical Engineering Undergraduate

    E-print Network

    Ghosh, Somnath

    Mechanical Engineering Department Undergraduate Advising Manual for Bachelor of Science Degrees in Mechanical Engineering and Engineering Mechanics 2011-2012 - Updated April 15, 2012 #12;Johns Hopkins University ­ Department of Mechanical Engineering 2011-2012 Undergraduate Student Advising Manual Page 2

  2. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar Yoed Rabin Department of Mechanical Engineering Carnegie at Carnegie Mellon University. Yoed Rabin is a Professor of Mechanical Engineering at Carnegie Mellon University. He received his DSc in Mechanical Engineering in 1994 from the Technion ­ Israel Institute

  3. Mechanical & Biomedical Engineering

    E-print Network

    Barrash, Warren

    Mechanical & Biomedical Engineering Department BACHELOR OF SCIENCE IN MECHANICAL ENGINEERING COURSE 105 Mechanical Engineering Graphics 3 CHEM 111L College Chemistry Lab (DLN) 1 ENGL 102 English Experimental Methods Lab (CID) 2 ME, CE, or ENGR 350 Engineering Mechanics of Materials 3 ME 352 Machine Design

  4. Mechanical Engineering Undergraduate

    E-print Network

    Ghosh, Somnath

    Mechanical Engineering Department Undergraduate Advising Manual for Bachelor of Science Degrees in Mechanical Engineering and Engineering Mechanics 2012-2013 - Updated July 14, 2013 #12;Johns Hopkins University ­ Department of Mechanical Engineering 2012-2013 Undergraduate Student Advising Manual Page 2

  5. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar Virginia Ferguson Department of Mechanical Engineering engineering of this interface. Virginia Ferguson (Ph.D. Mechanical Engineering, 2001) is Assistant Professor of Mechanical Engineering at the Univer- sity of Colorado, Boulder since 2006. She held a joint postdoctoral

  6. INTRODUCTION TO THE MECHANICS

    E-print Network

    Kaminski, Edouard

    INTRODUCTION TO THE MECHANICS OF A CONTINUOUS MEDIUM Lawrence E. Malvern Professor of Mechanics princi- ples common to all branches of solid and fluid mechanics, designed to appeal to the intuition science. The book arose from the need to provide a general preparation in contin- uum mechanics

  7. Research article Expression and evolution of the phosphoinositide-specific

    E-print Network

    Brendel, Volker

    MurashigeeSkoog salts, dehydration, and the plant hormone abscisic acid. Results of this and previous studies, inositol 1,4,5-trisphosphate; DAG, 1,2-diacylglycerol; MS, MurashigeeSkoog; ABA, abscisic acid; GCR1, G

  8. JournalofCellScience ERK and phosphoinositide 3-kinase temporally

    E-print Network

    Amaya, Enrique

    to draw the epithelium and connective tissue forward together in a `purse-string-like' fashion (Martin recovery of tissue integrity and homeostasis, which is vital for survival. Tissue movement in embryonic in a complex tissue setting, namely embryonic wound healing. Key words: ERK, PI3K, Wound healing, Rho GTPases

  9. Phosphoinositide 3-kinase in disease: timing, location, and scaffolding.

    PubMed

    Wymann, Matthias P; Marone, Romina

    2005-04-01

    When PI3Ks are deregulated by aberrant surface receptors or modulators, accumulation of PtdIns(3,4,5)P3 leads to increased cell growth, proliferation and contact-independent survival. The PI3K/PKB/TOR axis controls protein synthesis and growth, while PtdIns(3,4,5)P3-mediated activation of Rho GTPases directs cell motility. PI3K activity has been linked to the formation of tumors, metastasis, chronic inflammation, allergy and cardiovascular disease. Although increased PtdIns(3,4,5)P3 is a well-established cause of disease, it is seldom known which PI3K isoform is implied. Recent work has demonstrated that PI3Kgamma contributes to the control of cAMP levels in the cardiac system, where the protein acts as a scaffold, but not as a lipid kinase. PMID:15780590

  10. Mechanics rules cell biology

    PubMed Central

    2010-01-01

    Cells in the musculoskeletal system are subjected to various mechanical forces in vivo. Years of research have shown that these mechanical forces, including tension and compression, greatly influence various cellular functions such as gene expression, cell proliferation and differentiation, and secretion of matrix proteins. Cells also use mechanotransduction mechanisms to convert mechanical signals into a cascade of cellular and molecular events. This mini-review provides an overview of cell mechanobiology to highlight the notion that mechanics, mainly in the form of mechanical forces, dictates cell behaviors in terms of both cellular mechanobiological responses and mechanotransduction. PMID:20615211

  11. Internal pipe attachment mechanism

    DOEpatents

    Bast, R.M.; Chesnut, D.A.; Henning, C.D.; Lennon, J.P.; Pastrnak, J.W.; Smith, J.A.

    1994-12-13

    An attachment mechanism is described for repairing or extending fluid carrying pipes, casings, conduits, etc. utilizing one-way motion of spring tempered fingers to provide a mechanical connection between the attachment mechanism and the pipe. The spring tempered fingers flex to permit insertion into a pipe to a desired insertion depth. The mechanical connection is accomplished by reversing the insertion motion and the mechanical leverage in the fingers forces them outwardly against the inner wall of the pipe. A seal is generated by crushing a sealing assembly by the action of setting the mechanical connection. 6 figures.

  12. Mechanical bull thumb

    SciTech Connect

    Ginthner, T.P.; Schabel, S.I.

    1981-11-01

    Two patients suffered intra-articular fractures of the thumb while attempting to ride a mechanical bull. The mechanism of injury in these patients and the riding technique which leads to fracture are discussed.

  13. Mechanical and Aerospace Engineering

    E-print Network

    Mechanical and Aerospace Engineering seminar Plasma Actuators for Aerodynamic Flow Control Abstract Plasma actuators have become very popular in aerodynamic flow control research over the last) or thermal (Joule heating) although there is some unavoidable overlap between these mechanisms. Interest

  14. Time in quantum mechanics 

    E-print Network

    Chapin, Kimberly R.

    1997-01-01

    The role of time in quantum mechanics has been and is still very controversial. The purpose of this paper was to explore the historical interpretation of time in quantum mechanics, to determine the current status of this ...

  15. Compliant mechanism learning toolkit

    E-print Network

    Allard, Nicholas (Nicholas A.)

    2006-01-01

    This thesis concerns a toolkit designed to assist in learning the behavior of complaint mechanisms. In the design of complaint mechanisms, increasingly complicated designs behave in ways that are harder to intuitively ...

  16. Section II--Mechanisms Mechanisms of Succession

    E-print Network

    del Moral, Roger

    of Mount St. Helens was initiated by disturbances whose in- tensity and severity varied profoundly in space of successional mechanisms. This figure has evolved from early studies on Mount St. Helens (del Moral 1993

  17. Programmable Mechanical Metamaterials

    E-print Network

    Bastiaan Florijn; Corentin Coulais; Martin van Hecke

    2014-07-17

    We create mechanical metamaterials whose response to uniaxial compression can be programmed by lateral confinement, allowing monotonic, non-monotonic and hysteretic behavior. These functionalities arise from a broken rotational symmetry which causes highly nonlinear coupling of deformations along the two primary axes of these metamaterials. We introduce a soft mechanism model which captures the programmable mechanics, and outline a general design strategy for confined mechanical metamaterials. Finally, we show how inhomogeneous confinement can be explored to create multi stability and giant hysteresis.

  18. Time in quantum mechanics

    E-print Network

    Chapin, Kimberly R.

    1997-01-01

    to describe the quantum mechanical system The first, matrix mechanics, was presented by Heisenberg [31-33] in 1925. The second, wave mechanics, was presented by Schrodinger [34-37] a year later. In 1926, Schrodmger [38] demonstrated the equivalence... can jump &om one state to another. The result is a discontinuous variation in time (i. e. the tune atom) [42]. Throughout the development of quantum mechanics, this atomistic view of time surfaces again and again, For example, In 1925, J. J...

  19. Hybrid Mechanical Systems

    E-print Network

    Philipp Treutlein; Claudiu Genes; Klemens Hammerer; Martino Poggio; Peter Rabl

    2015-01-06

    We discuss hybrid systems in which a mechanical oscillator is coupled to another (microscopic) quantum system, such as trapped atoms or ions, solid-state spin qubits, or superconducting devices. We summarize and compare different coupling schemes and describe first experimental implementations. Hybrid mechanical systems enable new approaches to quantum control of mechanical objects, precision sensing, and quantum information processing.

  20. Giovanni Gallavotti STATISTICAL MECHANICS

    E-print Network

    Roma "La Sapienza", UniversitĂ  di

    Giovanni Gallavotti STATISTICAL MECHANICS Short Treatise Roma 1999 #12; . #12; Short treatise of Statistical Mechanics Giovanni Gallavotti Dipartimento di Fisica Universitâ??a di Roma La Sapienza 00185 Roma that started with my involvement as Coordinator of the Statistical Mechanics section of the Italian Encyclo

  1. Giovanni Gallavotti STATISTICAL MECHANICS

    E-print Network

    Roma "La Sapienza", UniversitĂ  di

    Giovanni Gallavotti STATISTICAL MECHANICS Short Treatise Roma 1999 #12;. #12;Short treatise of Statistical Mechanics Giovanni Gallavotti Dipartimento di Fisica Universit`a di Roma La Sapienza 00185 Roma that started with my involvement as Coordinator of the Statistical Mechanics section of the Italian Encyclo

  2. Fractional quantum mechanics

    Microsoft Academic Search

    Nikolai Laskin

    2000-01-01

    A path integral approach to quantum physics has been developed. Fractional path integrals over the paths of the Lévy flights are defined. It is shown that if the fractality of the Brownian trajectories leads to standard quantum and statistical mechanics, then the fractality of the Lévy paths leads to fractional quantum mechanics and fractional statistical mechanics. The fractional quantum and

  3. Department of Mechanical Engineering

    E-print Network

    Karonis, Nicholas T.

    Department of Mechanical Engineering Undergraduate Program 2007-2008 Northern Illinois University DeKalb, IL #12;DEPARTMENT OF MECHANICAL ENGINEERING NORTHERN ILLINOIS UNIVERSITY Web: http://www.ceet.niu.edu/depts/me/ Phone: 815-753-9979 The mission of the Department of Mechanical Engineering is to provide an up

  4. Department of Mechanical Engineering

    E-print Network

    Karonis, Nicholas T.

    Department of Mechanical Engineering Undergraduate Program 2006-2007 Northern Illinois University DeKalb, IL #12;DEPARTMENT OF MECHANICAL ENGINEERING NORTHERN ILLINOIS UNIVERSITY Web: http://www.ceet.niu.edu/depts/me/ Phone: 815-753-9979 The mission of the Department of Mechanical Engineering is to provide an up

  5. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar William P. King Professor University of Illinois. William P. King is the College of Engineering Bliss Professor in the Department of Mechanical Science and Engineering and Electrical and Computer Engineering. He received the Ph.D. degree in mechanical engineering

  6. Mechanical and Aerospace Engineering

    E-print Network

    Mechanical and Aerospace Engineering seminar Three Dimensional Traction Force Microscopy is the John E. Goode, Jr. Professor of Aerospace and Professor of Mechanical Engineering and Director.E. (Honors) in Mechanical Engineering from the University of Madras in 1981, Sc.M. in Engineering and Applied

  7. Mechanical engineering Department Seminar

    E-print Network

    Lin, Xi

    Mechanical engineering Department Seminar Shuodao Wang Postdoctoral Fellow University of Illinois, and continuous health monitors. Shuodao Wang received his Bachelor's degree in Engineering Mechanics from Tsinghua University in China (2007) and his Ph.D. degree in Mechanical Engineering from Northwestern

  8. Mechanical engineering Department Seminar

    E-print Network

    Lin, Xi

    Mechanical engineering Department Seminar Maureen Lynch Postdoctoral Fellow Cornell University role in this context. Dr. Lynch received her B.S. in Mechanical Engineering from Clemson University in 2005. She received her doctorate in Mechanical Engineering from Cornell University in 2010 under

  9. Missing Mechanism Information

    ERIC Educational Resources Information Center

    Tryon, Warren W.

    2009-01-01

    The first recommendation Kazdin made for advancing the psychotherapy research knowledge base, improving patient care, and reducing the gulf between research and practice was to study the mechanisms of therapeutic change. He noted, "The study of mechanisms of change has received the least attention even though understanding mechanisms may well be…

  10. Introduction to Quantum Mechanics

    E-print Network

    Eduardo J. S. Villaseńor

    2008-04-23

    The purpose of this contribution is to give a very brief introduction to Quantum Mechanics for an audience of mathematicians. I will follow Segal's approach to Quantum Mechanics paying special attention to algebraic issues. The usual representation of Quantum Mechanics on Hilbert spaces is also discussed.

  11. Basic Engineer Equipment Mechanic.

    ERIC Educational Resources Information Center

    Marine Corps Inst., Washington, DC.

    This student guide, one of a series of correspondence training courses designed to improve the job performance of members of the Marine Corps, deals with the skills needed by basic engineer equipment mechanics. Addressed in the four individual units of the course are the following topics: mechanics and their tools (mechanics, hand tools, and power…

  12. Mechanical seal assembly

    DOEpatents

    Kotlyar, Oleg M. (Salt Lake City, UT)

    2001-01-01

    An improved mechanical seal assembly is provided for sealing rotating shafts with respect to their shaft housings, wherein the rotating shafts are subject to substantial axial vibrations. The mechanical seal assembly generally includes a rotating sealing ring fixed to the shaft, a non-rotating sealing ring adjacent to and in close contact with the rotating sealing ring for forming an annular seal about the shaft, and a mechanical diode element that applies a biasing force to the non-rotating sealing ring by means of hemispherical joint. The alignment of the mechanical diode with respect to the sealing rings is maintained by a series of linear bearings positioned axially along a desired length of the mechanical diode. Alternative embodiments include mechanical or hydraulic amplification components for amplifying axial displacement of the non-rotating sealing ring and transferring it to the mechanical diode.

  13. Fractional Quantum Mechanics

    NASA Astrophysics Data System (ADS)

    Laskin, Nikolai

    2000-06-01

    A new application of a fractal concept to quantum physics has been developed. The fractional path integrals over the paths of the Levy flights are defined. It is shown that if fractality of the Brownian trajectories leads to standard quantum mechanics, then the fractality of the Levy paths leads to fractional quantum mechanics. The fractional quantum mechanics has been developed via the new fractional path integrals approach. A fractional generalization of the Schrodinger equation has been discovered. The new relationship between the energy and the momentum of the non-relativistic fractional quantum-mechanical particle has been found, and the Levy wave packet has been introduced into quantum mechanics. We have derived a free particle quantum-mechanical propagator using Fox's H-function. A fractional generalization of the Heisenberg uncertainty relation has been established. We also discuss the relationships between fractional and the well-known Feynman path integrals approaches to quantum mechanics.

  14. Mechanical Seal Assembly

    SciTech Connect

    Kotlyar, Oleg M.

    1999-06-18

    An improved mechanical seal assembly is provided for sealing rotating shafts with respect to their shaft housings, wherein the rotating shafts are subject to substantial axial vibrations. The mechanical seal assembly generally includes a rotating sealing ring fixed to the shaft, a non-rotating sealing ring adjacent to and in close contact with the rotating sealing ring for forming an annular seal about the shaft, and a mechanical diode element that applies a biasing force to the non-rotating sealing ring by means of hemispherical joint. The alignment of the mechanical diode with respect to the sealing rings is maintained by a series of linear bearings positioned axially along a desired length of the mechanical diode. Alternative embodiments include mechanical or hydraulic amplification components for amplifying axial displacement of the non-rotating sealing ring and transferring it to the mechanical diode.

  15. Mechanical seal assembly

    DOEpatents

    Kotlyar, Oleg M. (Salt Lake City, UT)

    2002-01-01

    An improved mechanical seal assembly is provided for sealing rotating shafts with respect to their shaft housings, wherein the rotating shafts are subject to substantial axial vibrations. The mechanical seal assembly generally includes a rotating sealing ring fixed to the shaft, a non-rotating sealing ring adjacent to and in close contact with the rotating sealing ring for forming an annular seal about the shaft, and a mechanical diode element that applies a biasing force to the non-rotating sealing ring by means of hemispherical joint. The alignment of the mechanical diode with respect to the sealing rings is maintained by a series of linear bearings positioned axially along a desired length of the mechanical diode. Alternative embodiments include mechanical or hydraulic amplification components for amplifying axial displacement of the non-rotating sealing ring and transfering it to the mechanical diode.

  16. GFP's Mechanical Intermediate States

    PubMed Central

    Saeger, John; Hytönen, Vesa P.; Klotzsch, Enrico; Vogel, Viola

    2012-01-01

    Green fluorescent protein (GFP) mutants have become the most widely used fluorescence markers in the life sciences, and although they are becoming increasingly popular as mechanical force or strain probes, there is little direct information on how their fluorescence changes when mechanically stretched. Here we derive high-resolution structural models of the mechanical intermediate states of stretched GFP using steered molecular dynamics (SMD) simulations. These structures were used to produce mutants of EGFP and EYFP that mimic GFP's different mechanical intermediates. A spectroscopic analysis revealed that a population of EGFP molecules with a missing N-terminal ?-helix was significantly dimmed, while the fluorescence lifetime characteristic of the anionic chromophore state remained unaffected. This suggests a mechanism how N-terminal deletions can switch the protonation state of the chromophore, and how the fluorescence of GFP molecules in response to mechanical disturbance might be turned off. PMID:23118864

  17. Fractional Classical Mechanics

    E-print Network

    Nick Laskin

    2013-02-03

    Fractional classical mechanics has been introduced and developed as a classical counterpart of the fractional quantum mechanics. Lagrange, Hamilton and Hamilton-Jacobi frameworks have been implemented for the fractional classical mechanics. The Lagrangian of fractional classical mechanics has been introduced, and equation of motion has been obtained. Fractional oscillator model has been launched and solved in 1D case. A new equation for the period of oscillations of fractional classical oscillator has been found. The interplay between the energy dependency of the period of classical oscillations and the non-equidistant distribution of the energy levels for fractional quantum oscillator has been discussed. We discuss as well, the relationships between new equations of fractional classical mechanics and the well-known fundamental equations of classical mechanics.

  18. Mechanical engineering Mechanical engineering is about solving problems, designing processes,

    E-print Network

    Waikato, University of

    Mechanical engineering Mechanical engineering is about solving problems, designing processes, and making products to improve the quality of human life and shape the economy. Mechanical engineers apply, from power stations to cars, robots and computers. The professional training mechanical engineers

  19. Challenges for Mechanism Design

    Microsoft Academic Search

    Marco Ceccarelli

    \\u000a Mechanism Design has been the core of mechanical engineering and indeed of industrial engineering since the beginning of engineering\\u000a practice and particularly in modern times. This has been motivated by the mechanical nature of tasks and interactions with\\u000a machines that will be still fundamental in human-machine operations, although new and new technologies will be available.\\u000a This will identify new interests

  20. Dynamic quantized fracture mechanics

    Microsoft Academic Search

    N. M. Pugno; R. S. Ruoff

    2006-01-01

    A new quantum action-based theory, dynamic quantized fracture mechanics (DQFM), is presented that modifies continuum-based\\u000a dynamic fracture mechanics (DFM). The crack propagation is assumed as quantized in both space and time. The static limit case\\u000a corresponds to quantized fracture mechanics (QFM), that we have recently developed to predict the strength of nanostructures.\\u000a DQFM predicts the well-known forbidden strength and crack

  1. Space Mechanisms Technology Workshop

    NASA Technical Reports Server (NTRS)

    Oswald, Fred B. (Editor)

    2002-01-01

    The Mechanical Components Branch at NASA Glenn Research Center hosted a workshop on Tuesday, May 14, 2002, to discuss space mechanisms technology. The theme for this workshop was 'Working in the Cold,' a focus on space mechanisms that must operate at low temperatures. We define 'cold' as below -60C (210 K), such as would be found near the equator of Mars. However, we are also concerned with much colder temperatures such as in permanently dark craters of the Moon (about 40 K).

  2. ENGINEERING MECHANICS SEMINARSENGINEERING MECHANICS SEMINARS THINK COMPOSITE

    E-print Network

    Ponce, V. Miguel

    and homogenized laminate designs as opposed to more traditional composite designs typically imitating metals@stanford.edu Use of fiber reinforced composites has continuously increased in structural designs for aerospace composite laminates for improved mechanical behaviour. Specifically, the epoxy cross-linking P

  3. Mechanisms of Vertebrate Synaptogenesis

    E-print Network

    Sandini, Giulio

    Mechanisms of Vertebrate Synaptogenesis Clarissa L. Waites,1 Ann Marie Craig,2 and Craig C. Garner1 . . . . . . . . . . . . . . 263 ACTIVITY-DEPENDENT REGULATION OF SYNAPTOGENESIS . . . . . . . . . . . . 264 Synapse Elimination

  4. Positioning Mechanism For Hoisting

    NASA Technical Reports Server (NTRS)

    Marlin, John D., III; Moore, Barry J.; Myers, Robert I.

    1992-01-01

    Mechanism positions large, heavy objects in container for lifting out by hoist, crane, or winch. Handles objects gently and ensures they are lifted cleanly away in vertical direction without bumping container. Developed for lifting offset pieces of solid-propellant core out of rocket motor through its propellant port. Similar specialized mechanisms can be developed to lift other specially shaped, specially contained heavy objects. Track in base of mechanism guides each trunnion and piece to which attached to middle as hydraulic rods extend. When mechanism lifted, tilted pieces swing inward and come to rest on energy-absorbing paddle.

  5. Structural Mechanics & Solid Mechanics A finite element toolbox to MATLAB

    E-print Network

    Ehrhardt, Matthias

    Structural Mechanics & Solid Mechanics Department of Mechanics and Materials CALFEM A finite.3 Copyright © 1999 by Structural Mechanics, LTH, Sweden. Printed by JABE Offset, Lund, Sweden. ISRN LUTVDG/TVSM--99/9001--SE (1-265) ISSN 0281-6679 Department of Mechanics and Materials Structural Mechanics #12;The

  6. Integrated Mechanical & Electrical Engineering (IMEE)

    E-print Network

    Burton, Geoffrey R.

    Integrated Mechanical & Electrical Engineering (IMEE) Department of Electronic & Electrical Engineering and Department of Mechanical Engineering #12;Graduates able to work in both mechanical of Mechanical Engineers (IMechE) n Develop essential engineering skills through extensive project work n Enhance

  7. Statistical Mechanics with focus on

    E-print Network

    Johannesson, Henrik

    Statistical Mechanics with focus on Liquids, Solutions and Colloidal Systems Course contents A. Foundations of statistical mechanics Classical dynamics ­ Hamilton's and Liouville's equations The concept thermodynamics and statistical mechanics. B. Liquid state theory; Equilibrium statistical mechanics for liquids

  8. MECHANICAL ENGINEERING UNDERGRADUATE HONORS THESIS

    E-print Network

    Prinz, Friedrich B.

    MECHANICAL ENGINEERING UNDERGRADUATE HONORS THESIS The Department of Mechanical Engineering offers a program leading to a Bachelor of Science in Mechanical Engineering with Honors. This program provides a unique opportunity for qualified mechanical engineering majors to conduct independent study

  9. MECHANICAL ENGINEERING Program of Study

    E-print Network

    Thomas, Andrew

    MECHANICAL ENGINEERING Program of Study Correspondence The Department of Mechanical Engineering offers graduate programs in the fields of thermal science and engineering mechanics. Current areas of research activity include Biomedical Engineering, Biomimetics, Composite Materials, Computational Mechanics

  10. Mechanical and Aerospace Engineering

    E-print Network

    with the interactions between mechanical stresses and ionic transport in the electrolyte of a solid oxide fuel cell-chemo- mechanical theory, two examples of practical interest will be discussed, namely, solid oxide fuel cells of microelectronic packaging, defects and transport in ionic solids with applications to solid oxide fuel cells

  11. A Simple Escapement Mechanism

    NSDL National Science Digital Library

    2012-06-26

    In this activity, learners build a simple mechanism that regulates the "escape" of energy released by a falling weight by portioning it into discrete amounts. Escapements are found in mechanical clocks, such as those driven by a pendulum or a spring. Learners will build the wrapping form of escapement said to be used in a fifteenth-century German clock.

  12. Understand mechanical seals

    Microsoft Academic Search

    Godse

    1995-01-01

    Most pump failures (about 70%) are attributed to malfunctioning mechanical seals. Rotating machinery consists primarily of centrifugal pumps. The regular revision of the API standard on pumps and the increasing role of environmental regulations bring mechanical seals into sharp focus for centrifugal pumps. A integrated process plant brings a variety of influences into the selection process: end user; process consultant;

  13. Mechanical Systems Signal Processing

    E-print Network

    Ray, Asok

    Mechanical Systems and Signal Processing Mechanical Systems and Signal Processing 21 (2007) 866 and analytical models. This paper attempts to address this inadequacy by taking advantage of advanced signal processing and pattern recognition tools. Since a vast majority of structural components that are prone

  14. Bacteriophage resistance mechanisms

    Microsoft Academic Search

    Simon J. Labrie; Julie E. Samson; Sylvain Moineau

    2010-01-01

    Phages are now acknowledged as the most abundant microorganisms on the planet and are also possibly the most diversified. This diversity is mostly driven by their dynamic adaptation when facing selective pressure such as phage resistance mechanisms, which are widespread in bacterial hosts. When infecting bacterial cells, phages face a range of antiviral mechanisms, and they have evolved multiple tactics

  15. Mechanical Engineering & Thermal Group

    E-print Network

    Mojzsis, Stephen J.

    import and export capability · Analyses involving finite element and finite difference model combinations: ANSYS, Matlab, MathCAD, FEMAP, and NASTRAN. Mechanical analyses performed include: · Static stress design is a critical element of instrument design. LASP mechanism designs are simple, use selectively

  16. Fractals and quantum mechanics

    NASA Astrophysics Data System (ADS)

    Laskin, Nick

    2000-12-01

    A new application of a fractal concept to quantum physics has been developed. The fractional path integrals over the paths of the Lévy flights are defined. It is shown that if fractality of the Brownian trajectories leads to standard quantum mechanics, then the fractality of the Lévy paths leads to fractional quantum mechanics. The fractional quantum mechanics has been developed via the new fractional path integrals approach. A fractional generalization of the Schrödinger equation has been discovered. The new relationship between the energy and the momentum of the nonrelativistic fractional quantum-mechanical particle has been established, and the Lévy wave packet has been introduced into quantum mechanics. The equation for the fractional plane wave function has been found. We have derived a free particle quantum-mechanical kernel using Fox's H-function. A fractional generalization of the Heisenberg uncertainty relation has been found. As physical applications of the fractional quantum mechanics we have studied a free particle in a square infinite potential well, the fractional "Bohr atom" and have developed a new fractional approach to the QCD problem of quarkonium. We also discuss the relationships between fractional and the well-known Feynman path integral approaches to quantum mechanics.

  17. Mechanical seal assembly

    Microsoft Academic Search

    Kotlyar; Oleg M

    2002-01-01

    An improved mechanical seal assembly is provided for sealing rotating shafts with respect to their shaft housings, wherein the rotating shafts are subject to substantial axial vibrations. The mechanical seal assembly generally includes a rotating sealing ring fixed to the shaft, a non-rotating sealing ring adjacent to and in close contact with the rotating sealing ring for forming an annular

  18. Mechanical seal assembly

    Microsoft Academic Search

    Kotlyar; Oleg M

    2001-01-01

    An improved mechanical seal assembly is provided for sealing rotating shafts with respect to their shaft housings, wherein the rotating shafts are subject to substantial axial vibrations. The mechanical seal assembly generally includes a rotating sealing ring fixed to the shaft, a non-rotating sealing ring adjacent to and in close contact with the rotating sealing ring for forming an annular

  19. Mechanical Seal Assembly

    Microsoft Academic Search

    Kotlyar; Oleg M

    1999-01-01

    An improved mechanical seal assembly is provided for sealing rotating shafts with respect to their shaft housings, wherein the rotating shafts are subject to substantial axial vibrations. The mechanical seal assembly generally includes a rotating sealing ring fixed to the shaft, a non-rotating sealing ring adjacent to and in close contact with the rotating sealing ring for forming an annular

  20. Quantum Mechanics Measurements, Mutually

    E-print Network

    Gruner, Daniel S.

    Quantum Mechanics Measurements, Mutually Unbiased Bases and Finite Geometry Or why six is the first) #12;Quantum Mechanics for Dummies Finite dimensional quantum states are represented by trace one,1 -icS1,1[ ] #12;Quantum systems evolve and are measured. The evolution of a quantum system using

  1. Fractional quantum mechanics

    PubMed

    Laskin

    2000-09-01

    A path integral approach to quantum physics has been developed. Fractional path integrals over the paths of the Levy flights are defined. It is shown that if the fractality of the Brownian trajectories leads to standard quantum and statistical mechanics, then the fractality of the Levy paths leads to fractional quantum mechanics and fractional statistical mechanics. The fractional quantum and statistical mechanics have been developed via our fractional path integral approach. A fractional generalization of the Schrodinger equation has been found. A relationship between the energy and the momentum of the nonrelativistic quantum-mechanical particle has been established. The equation for the fractional plane wave function has been obtained. We have derived a free particle quantum-mechanical kernel using Fox's H function. A fractional generalization of the Heisenberg uncertainty relation has been established. Fractional statistical mechanics has been developed via the path integral approach. A fractional generalization of the motion equation for the density matrix has been found. The density matrix of a free particle has been expressed in terms of the Fox's H function. We also discuss the relationships between fractional and the well-known Feynman path integral approaches to quantum and statistical mechanics. PMID:11088808

  2. Fractals and quantum mechanics.

    PubMed

    Laskin, Nick

    2000-12-01

    A new application of a fractal concept to quantum physics has been developed. The fractional path integrals over the paths of the Levy flights are defined. It is shown that if fractality of the Brownian trajectories leads to standard quantum mechanics, then the fractality of the Levy paths leads to fractional quantum mechanics. The fractional quantum mechanics has been developed via the new fractional path integrals approach. A fractional generalization of the Schrodinger equation has been discovered. The new relationship between the energy and the momentum of the nonrelativistic fractional quantum-mechanical particle has been established, and the Levy wave packet has been introduced into quantum mechanics. The equation for the fractional plane wave function has been found. We have derived a free particle quantum-mechanical kernel using Fox's H-function. A fractional generalization of the Heisenberg uncertainty relation has been found. As physical applications of the fractional quantum mechanics we have studied a free particle in a square infinite potential well, the fractional "Bohr atom" and have developed a new fractional approach to the QCD problem of quarkonium. We also discuss the relationships between fractional and the well-known Feynman path integral approaches to quantum mechanics. (c) 2000 American Institute of Physics. PMID:12779428

  3. INJECTION WELL MECHANICAL INTEGRITY

    EPA Science Inventory

    EPA's underground injection control regulations require that all injection wells demonstrate mechanical integrity. The regulations state that an injection well has mechanical integrity if (1) there is no significant leak in the casing, tubing or packer and (2) there is no signifi...

  4. Introduction to Quantum Mechanics

    NSDL National Science Digital Library

    The Concord Consortium

    2011-12-12

    The microscopic world is full of phenomena very different from what we see in everyday life. Some of those phenomena can only be explained using quantum mechanics. This activity introduces basic quantum mechanics concepts about electrons that are essential to understanding modern and future technology, especially nanotechnology. Start by exploring probability distribution, then discover the behavior of electrons with a series of simulations.

  5. Mechanics of Composite Materials

    Microsoft Academic Search

    Robert M. Jones

    1999-01-01

    The mechanical behavior of composites is traditionally evaluated on both microscopic and macroscopic scale to take into account inhomogeneity. Micromechanics attempts to quantify the interactions of fiber and matrix (reinforcement and resin) on a microscopic scale on par with the diameter of a single fiber. Macromechanics treats composites as homogeneous materials, with mechanical properties representative of the laminate as a

  6. Geometrization of Quantum Mechanics

    E-print Network

    J. F. Carinena; J. Clemente-Gallardo; G. Marmo

    2007-03-23

    We show that it is possible to represent various descriptions of Quantum Mechanics in geometrical terms. In particular we start with the space of observables and use the momentum map associated with the unitary group to provide an unified geometrical description for the different pictures of Quantum Mechanics. This construction provides an alternative to the usual GNS construction for pure states.

  7. Membrane Quantum Mechanics

    E-print Network

    Okazaki, Tadashi

    2015-01-01

    We consider the multiple M2-branes wrapped on a compact Riemann surface and study the arising quantum mechanics by taking the limit where the size of the Riemann surface goes to zero. The IR quantum mechanical models resulting from the BLG-model and the ABJM-model compactified on a torus are N = 16 and N = 12 superconformal gauged quantum mechanics. After integrating out the auxiliary gauge fields we find OSp(16|2) and SU(1,1|6) quantum mechanics from the reduced systems. The curved Riemann surface is taken as a holomorphic curve in a Calabi-Yau space to preserve supersymmetry and we present a prescription of the topological twisting. We find the N = 8 superconformal gauged quantum mechanics that may describe the motion of two wrapped M2-branes in a K3 surface.

  8. Mechanical code comparator

    DOEpatents

    Peter, Frank J. (Albuquerque, NM); Dalton, Larry J. (Bernalillo, NM); Plummer, David W. (Albuquerque, NM)

    2002-01-01

    A new class of mechanical code comparators is described which have broad potential for application in safety, surety, and security applications. These devices can be implemented as micro-scale electromechanical systems that isolate a secure or otherwise controlled device until an access code is entered. This access code is converted into a series of mechanical inputs to the mechanical code comparator, which compares the access code to a pre-input combination, entered previously into the mechanical code comparator by an operator at the system security control point. These devices provide extremely high levels of robust security. Being totally mechanical in operation, an access control system properly based on such devices cannot be circumvented by software attack alone.

  9. 07SCHOOL OF MECHANICAL ENGINEERING

    E-print Network

    Dimitrova, Vania

    07SCHOOL OF MECHANICAL ENGINEERING UNDERGRADUATE DEGREES School of Mechanical Engineering FACULTY OF ENGINEERING Undergraduate Degrees 2015 #12;www.engineering.leeds.ac.uk/mechanical UNDERGRADUATE DEGREES SCHOOL OF MECHANICAL ENGINEERING The School of Mechanical Engineering offers both a broad mechanical engineering degree

  10. Loperamide Inhibits Corticotrophic Cell Function by a Naloxone-lnsensitive Mechanism in the Rat in vitro

    Microsoft Academic Search

    Christoph Josef Auernhammer; Ulrich Renner; Otto-Albrecht Müller; Johanna Stalla; Günter Karl Stalla

    1993-01-01

    The effect of the antidiarrheal drug loperamide, a ?-opiate agonist, on ACTH secretion and biosynthesis, cAMP generation and phosphoinositide turnover was studied in rat anterior pituitary cell cultures. The cAMP-dependent protein kinase A pathway was stimulated with both corticotropin-releasing hormone (CRH; 2–5 nM) and the membrane-permeable Bu(2)cAMP (0.5–2.5 mM). The protein kinase C pathway was stimulated with 1 ?M arginine

  11. Naringenin Inhibits Glucose Uptake in MCF7 Breast Cancer Cells: A Mechanism for Impaired Cellular Proliferation

    Microsoft Academic Search

    Anne W. Harmon; Yashomati M. Patel

    2004-01-01

    Certain flavonoids inhibit glucose uptake in cultured cells. In this report, we show that the grapefruit flavanone naringenin inhibited insulin-stimulated glucose uptake in proliferating and growth-arrested MCF-7 breast cancer cells. Our findings indicate that naringenin inhibits the activity of phosphoinositide 3-kinase (PI3K), a key regulator of insulin-induced GLUT4 translocation, as shown by impaired phosphorylation of the downstream signaling molecule Akt.

  12. Overview of Bohmian Mechanics

    E-print Network

    Xavier Oriols; Jordi Mompart

    2013-01-09

    This chapter provides a comprehensive overview of the Bohmian formulation of quantum mechanics. It starts with a historical review of the difficulties found by Louis de Broglie, David Bohm, and John S. Bell to convince the scientific community about the validity and utility of Bohmian mechanics. Then, a formal explanation of Bohmian mechanics for nonrelativistic, single-particle quantum systems is presented. The generalization to many-particle systems, where the exchange interaction and the spin play an important role, is also presented. After that, the measurement process in Bohmian mechanics is discussed. It is emphasized that Bohmian mechanics exactly reproduces the mean value and temporal and spatial correlations obtained from the standard, that is the Copenhagen or orthodox, formulation. The ontological characteristics of Bohmian mechanics provide a description of measurements as another type of interaction without the need for introducing the wave function collapse. Several solved problems are presented at the end of the chapter, giving additional mathematical support to some particular issues. A detailed description of computational algorithms to obtain Bohmian trajectories from the numerical solution of the Schrodinger or the Hamilton-Jacobi equations are presented in an appendix. The motivation of this chapter is twofold: first, as a didactic introduction to Bohmian formalism, which is used in the subsequent chapters, and second, as a self-contained summary for any newcomer interested in using Bohmian mechanics in his or her daily research activity.

  13. Mechanism of Gravity Impulse

    E-print Network

    Ning Wu

    2005-10-01

    It is well-known that energy-momentum is the source of gravitational field. For a long time, it is generally believed that only stars with huge masses can generate strong gravitational field. Based on the unified theory of gravitational interactions and electromagnetic interactions, a new mechanism of the generation of gravitational field is studied. According to this mechanism, in some special conditions, electromagnetic energy can be directly converted into gravitational energy, and strong gravitational field can be generated without massive stars. Gravity impulse found in experiments is generated by this mechanism.

  14. Quantum Mechanics Without Observers

    E-print Network

    W. H. Sulis

    2013-03-03

    The measurement problem and the role of observers have plagued quantum mechanics since its conception. Attempts to resolve these have introduced anthropomorphic or non-realist notions into physics. A shift of perspective based upon process theory and utilizing methods from combinatorial games, interpolation theory and complex systems theory results in a novel realist version of quantum mechanics incorporating quasi-local, nondeterministic hidden variables that are compatible with the no-hidden variable theorems and relativistic invariance, and reproduce the standard results of quantum mechanics to a high degree of accuracy without invoking observers.

  15. Advanced Mechanics. Mathematical Introduction

    E-print Network

    G. Giachetta; L. Mangiarotti; G. Sardanashvily

    2010-01-20

    Classical non-relativistic mechanics in a general setting of time-dependent transformations and reference frame changes is formulated in the terms of fibre bundles over the time-axis R. Connections on fibre bundles are the main ingredient in this formulation of mechanics which thus is covariant under reference frame transformations. The basic notions of a non-relativistic reference frame, a relative velocity, a free motion equation, a relative acceleration, an external force are formulated. Newtonian, Lagrangian, Hamiltonian mechanical systems and the relations between them are defined. Lagrangian and Hamiltonian conservation laws are considered.

  16. Is quantum mechanics exact?

    SciTech Connect

    Kapustin, Anton [California Institute of Technology, Pasadena, California 91125 (United States)] [California Institute of Technology, Pasadena, California 91125 (United States)

    2013-06-15

    We formulate physically motivated axioms for a physical theory which for systems with a finite number of degrees of freedom uniquely lead to quantum mechanics as the only nontrivial consistent theory. Complex numbers and the existence of the Planck constant common to all systems arise naturally in this approach. The axioms are divided into two groups covering kinematics and basic measurement theory, respectively. We show that even if the second group of axioms is dropped, there are no deformations of quantum mechanics which preserve the kinematic axioms. Thus, any theory going beyond quantum mechanics must represent a radical departure from the usual a priori assumptions about the laws of nature.

  17. Mechanics and mechanisms of ultrasonic metal welding

    NASA Astrophysics Data System (ADS)

    de Vries, Edgar

    During ultrasonic welding of sheet metal, normal and shear forces act on the parts to be welded and the weld interface. These forces are a result of the ultrasonic vibrations of the tool, pressed onto the parts to be welded. Furthermore they determine the weld quality and the power that is needed to produce the weld. The main goal in this study is to measure and calculate the tangential forces during ultrasonic metal welding that act on the parts and the weld interface and correlate them to weld quality. In this study a mechanics based model was developed which included a model for the temperature generation during welding and its effect on the mechanical material properties. This model was then used to calculate the interface forces during welding. The model results were in good agreement with the experimental results, which included the measured shear force during welding. With the knowledge of the forces that act at the interface it might be possible to control weld quality (strength) and avoid sonotrode welding (sticking of the sonotrode to the parts). Without a solution to these two problems USMW will never be applicable to large scale automated production use, despite its advantages. In the experiments the influence of part dimensions, friction coefficient, normal force and vibration amplitude on weld quality and sonotrode adhesion were examined. The presented model is capable of predicting and explaining unfavorable welding conditions, therefore making it possible to predetermine weld locations on larger parts or what surface preparation of the parts to be welded would lead to an improved welding result. Furthermore shear force at the anvil measured during welding could be correlated to changing welding conditions. This is a new approach of explaining the process of USMW, because it is based on mechanical considerations. The use of a shear force measuring anvil has the potential to be implemented into welding systems and the shear force would provide an additional means of process control.

  18. The mechanism of DNA mechanical unzipping

    NASA Astrophysics Data System (ADS)

    Volkov, S. N.; Solov'yov, A. V.

    2009-09-01

    The process of DNA double helix unzipping is considered to be a nonlinear dynamical process powered by an external force. In the course of unzipping the most probable pathway of the mechanical strands separation is determined. This pathway accounts for the structural organization of the double helix, the manner in which the external force is applied, and the kinetic parameters of the base pair opening in the double-stranded DNA chain. It is assumed in our model that the base pair unzipping includes the stretch of DNA complementary base pairs and the double helix torsion. For the description of the DNA unzipping the two-stage mechanism of the external force action is proposed. The mechanism explains the cooperative nature of the unzipping process under the physiological conditions and the threshold character of external force action. On the first stage the external force transforms the conformational state of the double helix and makes the unzipping possible. On the second stage the unzipping propagates along dsDNA. The boundary between the open and the closed parts of the helix (the so called `fork') is demonstrated to move along the chain as a step-like excitation (kink soliton). It is shown that for stable unzipping propagation along DNA double helix, it is necessary to allow rotation of the DNA chain while keeping the velocity of the unzipping fork propagation small in comparison to the velocity of sound in the DNA. It is demonstrated that fluctuations of the external force at the constant velocity of unzipping and the fluctuations of the DNA unzipping length at the fixed force have the same origin: the heterogeneity of the DNA base pairs.

  19. The orbital mechanics of flight mechanics

    NASA Technical Reports Server (NTRS)

    Dunning, R. S.

    1973-01-01

    A reference handbook on modern dynamic orbit theory is presented. Starting from the most basic inverse-square law, the law of gravity for a sphere is developed, and the motion of point masses under the influence of a sphere is considered. The reentry theory and the orbital theory are discussed along with the relative motion between two bodies in orbit about the same planet. Relative-motion equations, rectangular coordinates, and the mechanics of simple rigid bodies under the influence of a gravity gradient field are also discussed.

  20. Mechanical Engineering Is Mechanical Engineering right for me?

    E-print Network

    Harman, Neal.A.

    Mechanical Engineering Is Mechanical Engineering right for me? If you are interested in the wide range of principles related to mechanical systems then Mechanical Engineering is well suited to you. A Mechanical Engineering degree programme will focus on aspects such as analysis, design, manufacture

  1. Cell Mechanics and Mechanosensitivity 1 Cell Mechanics and

    E-print Network

    Cell Mechanics and Mechanosensitivity 1 Fall 2014 MW 12-2:00 BE535 Cell Mechanics mechanical force and energy. We will closely follow the new text; Cell Mechanics and Mechanobiology" by C biology: fundamentals, Receptors, Experimental methods and Genetics. 4) Review of Solid Mechanics 5

  2. Applied Mechanics Materials

    NSDL National Science Digital Library

    This site from SpaceTEC National Aerospace Technical Education Center presents materials on applied mechanics in the aerospace workplace. Topics include machine shop safety, non-cutting hand tools, cutting hand tools, drilling, measurement, basic calculations, hardware and blueprints.

  3. Department of MECHANICAL ENGINEERING

    E-print Network

    Barthelat, Francois

    : Aerodynamics, Fluids and Thermal Engineering, Mechanics of Materials and Structures, Dynamics and Control .............................................................................................................. 11 Office Space and Desk Allocation ............................................................................................. 13 The Faculty and Their Research Aerodynamics, Fluids and Thermal Engineering

  4. Mechanical Engineering Major Program

    E-print Network

    Chen, Wei

    ) Chemistry (2) Physics (2) EDC (2) Public Speaking (1) ME ­ Advanced Study ­ Design Choose 1 ME ­ Advanced-1 EDC I DSGN 106-2 EDC II GEN_CMN 102 Public Speaking Chem 171 (Honors) Replaces 101/102 Mechanical

  5. Mechanical Compression Heat Pumps 

    E-print Network

    Apaloo, T. L.; Kawamura, K.; Matsuda, J.

    1986-01-01

    to develop, design and test compressors built to meet the needs of the mechanically demanding industrial heat pump applications which often require high compression ratios and temperatures in excess of 200 degrees F. This paper will review the theoretical...

  6. Ultralight, ultrastiff mechanical metamaterials

    E-print Network

    Zheng, Xiaoyu

    The mechanical properties of ordinary materials degrade substantially with reduced density because their structural elements bend under applied load. We report a class of microarchitected materials that maintain a nearly ...

  7. Orbital Mechanics Analysis Program

    NASA Technical Reports Server (NTRS)

    Simon, W. C.; Jankowski, S. C.; Hughes, T. B.

    1985-01-01

    Orbital Mechanics Analysis Program provides engineers with simple tool for analysis or synthesis of any orbital maneuvering function involving vehicle and target. Program useful in such applications as proximity operations and rendezvous maneuvers.

  8. Mechanical & Aerospace Engineering

    E-print Network

    Mechanical & Aerospace Engineering Spall fracture is an important failure mode in metallic, the misorientation across adjacent grains, the presence of an incoherent twin boundary or a terminated twin at a GB

  9. Failure mechanisms in MEMS.

    SciTech Connect

    Walraven, Jeremy Allen

    2003-07-01

    MEMS components by their very nature have different and unique failure mechanisms than their macroscopic counterparts. This paper discusses failure mechanisms observed in various MEMS components and technologies. MEMS devices fabricated using bulk and surface micromachining process technologies are emphasized. MEMS devices offer uniqueness in their application, fabrication, and functionality. Their uniqueness creates various failure mechanisms not typically found in their bulk or IC counterparts. In ICs, electrical precautions are taken to mitigate failure. In MEMS, both electrical and mechanical precautions must be enacted to reduce the risk of failure and increased reliability. Unlike ICs, many MEMS components are designed to interact with their environment, making the fabrication, testing, and packaging processes critical for the success of the device.

  10. MECHANISMS OF PESTICIDE DEGRADATION

    EPA Science Inventory

    This research project was initiated with the overall objective of determining (1) the chemical structures of toxic components of toxaphene, (2) to study anaerobic metabolism to degrade toxaphene and other pesticides, and (3) to understand toxic action mechanism of chlordimeform. ...

  11. Mechanical engineering Department Seminar

    E-print Network

    , and on sabbatical leave at White House Office of Science and Technology Policy where he serves as the assistant in mechanisms and robotics, the 2012 ASME Mecha- nisms and Robotics Award, the 2012 IEEE Robotics and Automation

  12. UNDERGRADUATE STUDENT MANUAL Department of Mechanical Engineering

    E-print Network

    Plotkin, Joshua B.

    1 UNDERGRADUATE STUDENT MANUAL Department of Mechanical Engineering and Applied Mechanics Engineering and Applied Mechanics? ........................................................................3..................................................................................................................................5 Curriculum in Mechanical Engineering and Applied Mechanics

  13. Mechanical Properties of Nanostructures

    Microsoft Academic Search

    Bharat Bhushan

    Knowledge of the mechanical properties of nanostructures is necessary for designing realistic MEMS\\/NEMS devices. Microelectromechanical\\u000a systems (MEMS) refer to microscopic devices that have a characteristic length of less than 1mm but more than 1 µm and combine\\u000a electrical and mechanical components. Nanoelectromechanical systems (NEMS) refer to nanoscopic devices that have a characteristic\\u000a length of less than 1 µm and combine

  14. Ultrasound—biophysics mechanisms

    PubMed Central

    O'Brien, William D.

    2007-01-01

    Ultrasonic biophysics is the study of mechanisms responsible for how ultrasound and biological materials interact. Ultrasound-induced bioeffect or risk studies focus on issues related to the effects of ultrasound on biological materials. On the other hand, when biological materials affect the ultrasonic wave, this can be viewed as the basis for diagnostic ultrasound. Thus, an understanding of the interaction of ultrasound with tissue provides the scientific basis for image production and risk assessment. Relative to the bioeffect or risk studies, that is, the biophysical mechanisms by which ultrasound affects biological materials, ultrasound-induced bioeffects are generally separated into thermal and nonthermal mechanisms. Ultrasonic dosimetry is concerned with the quantitative determination of ultrasonic energy interaction with biological materials. Whenever ultrasonic energy is propagated into an attenuating material such as tissue, the amplitude of the wave decreases with distance. This attenuation is due to either absorption or scattering. Absorption is a mechanism that represents that portion of ultrasonic wave that is converted into heat, and scattering can be thought of as that portion of the wave, which changes direction. Because the medium can absorb energy to produce heat, a temperature rise may occur as long as the rate of heat production is greater than the rate of heat removal. Current interest with thermally mediated ultrasound-induced bioeffects has focused on the thermal isoeffect concept. The non-thermal mechanism that has received the most attention is acoustically generated cavitation wherein ultrasonic energy by cavitation bubbles is concentrated. Acoustic cavitation, in a broad sense, refers to ultrasonically induced bubble activity occurring in a biological material that contains pre-existing gaseous inclusions. Cavitation-related mechanisms include radiation force, microstreaming, shock waves, free radicals, microjets and strain. It is more challenging to deduce the causes of mechanical effects in tissues that do not contain gas bodies. These ultrasonic biophysics mechanisms will be discussed in the context of diagnostic ultrasound exposure risk concerns. PMID:16934858

  15. Quantized fracture mechanics

    Microsoft Academic Search

    Nicola M. Pugno; Rodney S. Ruoff

    2004-01-01

    A new energy-based theory, quantized fracture mechanics (QFM), is presented that modifies continuum-based fracture mechanics; stress- and strain-based QFM analogs are also proposed. The differentials in Griffith's criterion are substituted with finite differences; the implications are remarkable. Fracture of tiny systems with a given geometry and type of loading occurs at ‘quantized’ stresses that are well predicted by QFM: strengths

  16. Quantized fracture mechanics

    Microsoft Academic Search

    Nicola M. Pugno; Rodney S. Ruoff

    2004-01-01

    A new energy-based theory, quantized fracture mechanics (QFM), is presented that modifies continuum-based fracture mechanics; stress- and strain-based QFM analogs are also proposed. The differentials in Griffith's criterion are substituted with finite differences; the implications are remarkable. Fracture of tiny systems with a given geometry and type of loading occurs at 'quantized' stresses that are well predicted by QFM: strengths

  17. Upper Airway Mechanics

    Microsoft Academic Search

    Johan A. Verbraecken; Wilfried A. De Backer

    2009-01-01

    This review discusses the pathophysiological aspects of sleep-disordered breathing, with focus on upper airway mechanics in obstructive and central sleep apnoea, Cheyne-Stokes respiration and obesity hypoventilation syndrome. These disorders constitute the end points of a spectrum with distinct yet interrelated mechanisms that lead to substantial pathology, i.e. increased upper airway collapsibility, control of breathing instability, increased work of breathing, disturbed

  18. Mechanisms of catalyst deactivation

    Microsoft Academic Search

    Calvin H Bartholomew

    2001-01-01

    The literature treating mechanisms of catalyst deactivation is reviewed. Intrinsic mechanisms of catalyst deactivation are many; nevertheless, they can be classified into six distinct types: (i) poisoning, (ii) fouling, (iii) thermal degradation, (iv) vapor compound formation accompanied by transport, (v) vapor-solid and\\/or solid-solid reactions, and (vi) attrition\\/crushing. As (i), (iv), and (v) are chemical in nature and (ii) and (v)

  19. Supersymmetry in quantum mechanics

    Microsoft Academic Search

    Avinash Khare

    1997-01-01

    In the past ten years, the ideas of supersymmetry have been profitably applied to many nonrelativistic quantum mechanical\\u000a problems. In particular, there is now a much deeper understanding of why certain potentials are analytically solvable. In\\u000a this lecture I review the theoretical formulation of supersymmetric quantum mechanics and discuss many of its applications.\\u000a I show that the well-known exactly solvable

  20. Mechanics of Materials

    NSDL National Science Digital Library

    Philpot, Timothy A.

    MecMovies is an extensive collection of examples, theory, and games designed to complement the entire Mechanics of Materials course. The software features impressive graphics and animation that are highly effective in visually communicating course concepts to students. Special emphasis is placed on developing the learner's understanding and proficiency in basic concepts and skills through interactive exercises and games. Classroom implementation of the software has produced improved student performance and more positive student attitudes regarding the Mechanics of Materials course.

  1. Interprocess Communications Mechanisms

    NSDL National Science Digital Library

    Rusling, David

    This website created by David A Rusling explains the various components of Interprocess Communications Mechanisms supported by Linux. Signals, pipes, and sockets are the main features of this website with a section dedicated to each. Each section contains descriptions and explanations, sub-headings, diagrams, and/or charts. This is a great overview to Interprocess Communications Mechanisms and teachers and students alike will find this a worthwhile bookmark in their browsers.

  2. Quantized fracture mechanics

    NASA Astrophysics Data System (ADS)

    Pugno, Nicola M.; Ruoff, Rodney S.

    2004-09-01

    A new energy-based theory, quantized fracture mechanics (QFM), is presented that modifies continuum-based fracture mechanics; stress- and strain-based QFM analogs are also proposed. The differentials in Griffith's criterion are substituted with finite differences; the implications are remarkable. Fracture of tiny systems with a given geometry and type of loading occurs at 'quantized' stresses that are well predicted by QFM: strengths predicted by QFM are compared with experimental results on carbon nanotubes, ?-SiC nanorods, ?-Si3N4 whiskers, and polysilicon thin films; and also with molecular mechanics/dynamics simulation of fracture of carbon nanotubes and graphene with cracks and holes, and statistical mechanics-based simulations on fracture of two-dimensional spring networks. QFM is self-consistent, agreeing to first-order with linear elastic fracture mechanics (LEFM), and to second-order with non-linear fracture mechanics (NLFM). For vanishing crack length QFM predicts a finite ideal strength in agreement with Orowan's prediction. In contrast to LEFM, QFM has no restrictions on treating defect size and shape. The different fracture Modes (opening I, sliding II and tearing III), and the stability of the fracture propagations, are treated in a simple way.

  3. Lung Parenchymal Mechanics

    PubMed Central

    Suki, Béla; Stamenovic, Dimitrije; Hubmayr, Rolf

    2014-01-01

    The lung parenchyma comprises a large number of thin-walled alveoli, forming an enormous surface area, which serves to maintain proper gas exchange. The alveoli are held open by the transpulmonary pressure, or prestress, which is balanced by tissues forces and alveolar surface film forces. Gas exchange efficiency is thus inextricably linked to three fundamental features of the lung: parenchymal architecture, prestress, and the mechanical properties of the parenchyma. The prestress is a key determinant of lung deformability that influences many phenomena including local ventilation, regional blood flow, tissue stiffness, smooth muscle contractility, and alveolar stability. The main pathway for stress transmission is through the extracellular matrix. Thus, the mechanical properties of the matrix play a key role both in lung function and biology. These mechanical properties in turn are determined by the constituents of the tissue, including elastin, collagen, and proteoglycans. In addition, the macroscopic mechanical properties are also influenced by the surface tension and, to some extent, the contractile state of the adherent cells. This article focuses on the biomechanical properties of the main constituents of the parenchyma in the presence of prestress and how these properties define normal function or change in disease. An integrated view of lung mechanics is presented and the utility of parenchymal mechanics at the bedside as well as its possible future role in lung physiology and medicine are discussed. PMID:23733644

  4. Mechanics of the Orbita

    PubMed Central

    Demer, Joseph L.

    2008-01-01

    The oculomotor periphery was formerly regarded as a simple mechanism executing complex behaviors explicitly specified by innervation. It is now recognized that several fundamental aspects of ocular motility are properties of the extraocular muscles (EOMs) and their associated connective tissue pulleys. The Active Pulley Hypothesis proposes that rectus and inferior oblique EOMs have connective tissue soft pulleys that are actively controlled by the direction action of the EOMs’ orbital layers. Functional imaging and histology have suggested that the rectus pulley array constitutes an inner mechanism, similar to a gimbal, that is rotated torsionally around the orbital axis by an outer mechanism driven by the oblique EOMs. This arrangement may mechanically account for several commutative aspects of ocular motor control, including Listing’s law, yet permits implementation of noncommutative motility as during the vestibulo-ocular reflex. Recent human behavioral studies, as well neurophysiology in monkeys, are consistent with mechanical rather than central neural implementation of Listing’s law. Pathology of the pulley system is associated with predictable patterns of strabismus that are surgically treatable when the pathologic anatomy is characterized by imaging. This mechanical determination may imply limited possibilities for neural adaptation to some ocular motor pathologies, but indicates greater potential for surgical treatments. PMID:17314483

  5. Wear and Tear - Mechanical

    NASA Technical Reports Server (NTRS)

    Swanson, Theodore

    2008-01-01

    The focus of this chapter is on the long term wear and tear, or aging, of the mechanical subsystem of a spacecraft. The mechanical subsystem is herein considered to be the primary support structure (as in a skeleton or exoskeleton) upon which all other spacecraft systems rest, and the associated mechanisms. Mechanisms are devices which have some component that moves at least once, in response to some type of passive or active control system. For the structure, aging may proceed as a gradual degradation of mechanical properties and/or function, possibly leading to complete structural failure over an extended period of time. However, over the 50 years of the Space Age such failures appear to be unusual. In contrast, failures for mechanisms are much more frequent and may have a very serious effect on mission performance. Just as on Earth, all moving devices are subject to normal (and possibly accelerated) degradation from mechanical wear due to loss or breakdown of lubricant, misalignment, temperature cycling effects, improper design/selection of materials, fatigue, and a variety of other effects. In space, such environmental factors as severe temperature swings (possibly 100's of degrees C while going in and out of direct solar exposure), hard vacuum, micrometeoroids, wear from operation in a dusty or contaminated environment, and materials degradation from radiation can be much worse. In addition, there are some ground handling issues such as humidity, long term storage, and ground transport which may be of concern. This chapter addresses the elements of the mechanical subsystem subject to wear, and identifies possible causes. The potential impact of such degradation is addressed, albeit with the recognition that the impact of such wear often depends on when it occurs and on what specific components. Most structural elements of the mechanical system typically are conservatively designed (often to a safety factor of greater than approximately 1.25 on yield for unmanned spacecraft) but do not have backup structure due to the added mass this would impose, and also due to the fact that structural elements can be accurately modeled mathematically and in test. Critical mechanisms or devices may have backups, or alternate work-arounds, since characterization of these systems in a 1g environment is less accurate than structure, and repair in-space is often impossible.

  6. Engineering Mechanics Annual Report 2002

    E-print Network

    Franssen, Michael

    Engineering Mechanics Annual Report 2002 Graduate School Engineering Mechanics c/o Eindhoven.1 1.2 Mission statement 1.2 1.3 Outline of the field of Engineering Mechanics 1.2 1.4 Organization 1 and Engineering 4.1 5. Engineering Mechanics (1) 5.1 6. Aerospace Structures and Computational Mechanics 6.1 7

  7. Engineering Mechanics Annual Report 2001

    E-print Network

    Franssen, Michael

    Engineering Mechanics Annual Report 2001 Graduate School Engineering Mechanics c/o Eindhoven.3 Outline of the field of Engineering Mechanics 1.2 1.4 Organization 1.3 1.5 Participants 1.4 1.6 Research Analysis 6.1 7. Engineering Mechanics (1) 7.1 8. Aerospace Structures and Computational Mechanics 8.1 9

  8. Vanguard Satellite Separation Mechanisms

    NASA Technical Reports Server (NTRS)

    Baumann, Robert C.

    1961-01-01

    Early in the Vanguard program it became apparent that a thoroughly reliable means of separating the satellite packages from the third-stage rocket would be required. A completely self -contained standard mechanism was developed with redundant firing circuits for use on both test vehicles and satellite-launching vehicles. A change in the experimental objectives of the test-vehicle payload units necessitated modification of some of the standard separation mechanisms. A strap, pull-pin, girth-ring separation device was developed which employed the basic actuation of the standard mechanisms. Evidence of residual burning of the third stage made it necessary to delay separation longer than the time designed into the long-delay separation device. The standard separation mechanism was modified and integrated with the satellite command receiver system so that a ground command after third-stage burnout would cause separation. Flight performance of the various separation mechanisms proved their reliability; they performed without failure in all Vanguard launchings.

  9. STANFORD UNIVERSITY MECHANICAL ENGINEERING DEPARTMENT

    E-print Network

    Prinz, Friedrich B.

    STANFORD UNIVERSITY MECHANICAL ENGINEERING DEPARTMENT GRADUATE STUDENT HANDBOOK Academic Year 20092010 Mechanical Engineering Student Services Building 530, Room 125 (650) 7257695 FAX (650) 7234882 Revised 9/09 #12;MECHANICAL ENGINEERING GRADUATE STUDENT HANDBOOK 20092010 TABLE OF CONTENTS Page

  10. Continuous Improvement Plan Mechanical Engineering

    E-print Network

    Fernandez, Eduardo

    Continuous Improvement Plan Mechanical Engineering Undergraduate Curriculum #12;Plan for the Assessment and Continuous Improvement of the Mechanical Engineering Undergraduate Curriculum Department of Ocean and Mechanical Engineering Florida Atlantic University April 4, 2001 (latest modification 3

  11. STANFORD UNIVERSITY MECHANICAL ENGINEERING DEPARTMENT

    E-print Network

    Prinz, Friedrich B.

    STANFORD UNIVERSITY MECHANICAL ENGINEERING DEPARTMENT GRADUATE STUDENT HANDBOOK Academic Year 20062007 Mechanical Engineering Student Services Building 530, Room 125 (650) 7257695 FAX (650) 7234882 Revised 9/06 #12;MECHANICAL ENGINEERING GRADUATE STUDENT HANDBOOK 20062007 TABLE OF CONTENTS (Clickable

  12. The continuum and wave mechanics

    E-print Network

    Collins, Royal Eugene

    1954-01-01

    THE CONTINUUM AND WATS MECHANICS By ROYAL SUGUDD COLLINS Approved as to etyle and content "by: THE CONTINUUM AND WAVE MECHANICS A Dissertation By ROYAL EUGENE COLLINS li i Submitted to the Graduate School of the Agricultural and Mechanical...

  13. Design of compliant microleverage mechanisms

    Microsoft Academic Search

    Xiao-Ping S Su; Henry S Yang

    2001-01-01

    A compliant microleverage mechanism can be used as a mechanical amplifier in micro-electro-mechanical systems (MEMS) to transfer an input to output for achieving mechanical or geometric advantages, such as amplifying force or displacement. Constrained by micro-fabrication technologies, a microleverage mechanism is formed by co-planar flexures, achieving mechanical transformation through elastic deformation. This paper presents the design theory and synthesis of

  14. Mechanisms of hypothermic neuroprotection.

    PubMed

    Drury, Paul P; Gunn, Eleanor R; Bennet, Laura; Gunn, Alistair J

    2014-03-01

    Prolonged, moderate cerebral hypothermia initiated within a few hours after severe hypoxia-ischemia and continued until resolution of the acute phase of delayed cell death can reduce acute brain injury and improve long-term behavioral recovery in term infants and in adults after cardiac arrest. The specific mechanisms of hypothermic neuroprotection remain unclear, in part because hypothermia suppresses a broad range of potential injurious factors. This article examines proposed mechanisms in relation to the known window of opportunity for effective protection with hypothermia. Knowledge of the mechanisms of hypothermia will help guide the rational development of future combination treatments to augment neuroprotection with hypothermia and identify those most likely to benefit. PMID:24524453

  15. [Knee extensor mechanism ruptures].

    PubMed

    Duthon, V B; Fritschy, D

    2011-08-10

    Knee extensor mechanism is composed of the quadriceps and its tendon, patella and patellar tendon. Rupture of either the quadriceps or patellar tendon, or of the patella itself, lead to a disruption of the knee extensor mechanism. Clinical examination reveals an inability to actively extend the knee. Standard radiographs of the knee show a suprapatellar swelling of the soft tissues in case of quadriceps tendon rupture, or a displaced patellar fracture, or a patella alta in case of patellar tendon rupture. Echography and MRI confirm the diagnosis and may reveal associated injuries. In case of knee extensor mechanism rupture, early surgical reconstruction of quadriceps or patellar tendon, or patella osteosynthesis, are mandatory to achieve early functional recovery. PMID:21919393

  16. Epigenetics: Biology's Quantum Mechanics

    PubMed Central

    Jorgensen, Richard A.

    2011-01-01

    The perspective presented here is that modern genetics is at a similar stage of development as were early formulations of quantum mechanics theory in the 1920s and that in 2010 we are at the dawn of a new revolution in genetics that promises to enrich and deepen our understanding of the gene and the genome. The interrelationships and interdependence of two views of the gene – the molecular biological view and the epigenetic view – are explored, and it is argued that the classical molecular biological view is incomplete without incorporation of the epigenetic perspective and that in a sense the molecular biological view has been evolving to include the epigenetic view. Intriguingly, this evolution of the molecular view toward the broader and more inclusive epigenetic view of the gene has an intriguing, if not precise, parallel in the evolution of concepts of atomic physics from Newtonian mechanics to quantum mechanics that are interesting to consider. PMID:22639577

  17. Graphene Statistical Mechanics

    NASA Astrophysics Data System (ADS)

    Bowick, Mark; Kosmrlj, Andrej; Nelson, David; Sknepnek, Rastko

    2015-03-01

    Graphene provides an ideal system to test the statistical mechanics of thermally fluctuating elastic membranes. The high Young's modulus of graphene means that thermal fluctuations over even small length scales significantly stiffen the renormalized bending rigidity. We study the effect of thermal fluctuations on graphene ribbons of width W and length L, pinned at one end, via coarse-grained Molecular Dynamics simulations and compare with analytic predictions of the scaling of width-averaged root-mean-squared height fluctuations as a function of distance along the ribbon. Scaling collapse as a function of W and L also allows us to extract the scaling exponent eta governing the long-wavelength stiffening of the bending rigidity. A full understanding of the geometry-dependent mechanical properties of graphene, including arrays of cuts, may allow the design of a variety of modular elements with desired mechanical properties starting from pure graphene alone. Supported by NSF grant DMR-1435794

  18. MIRO Calibration Switch Mechanism

    NASA Technical Reports Server (NTRS)

    Suchman, Jason; Salinas, Yuki; Kubo, Holly

    2001-01-01

    The Jet Propulsion Laboratory has designed, analyzed, built, and tested a calibration switch mechanism for the MIRO instrument on the ROSETTA spacecraft. MIRO is the Microwave Instrument for the Rosetta Orbiter; this instrument hopes to investigate the origin of the solar system by studying the origin of comets. Specifically, the instrument will be the first to use submillimeter and millimeter wave heterodyne receivers to remotely examine the P-54 Wirtanen comet. In order to calibrate the instrument, it needs to view a hot and cold target. The purpose of the mechanism is to divert the instrument's field of view from the hot target, to the cold target, and then back into space. This cycle is to be repeated every 30 minutes for the duration of the 1.5 year mission. The paper describes the development of the mechanism, as well as analysis and testing techniques.

  19. PROSPECTIVE STUDENT HANDBOOK Mechanical Engineering

    E-print Network

    Wang, Deli

    attitude determination and control, space structures, orbital mechanics, and rocket propulsion, flight mechanics, and stability and control. · Knowledge of topics in astronautical engineering including

  20. Chemical Mechanical Planarization- Chemical

    NSDL National Science Digital Library

    This website includes an animation which illustrates the chemical action of slurry in the chemical-mechanical planarization process. Objective: Explain the mechanical and chemical steps in the CMP process. This simulation is from Module 068 of the Process & Equipment III Cluster of the MATEC Module Library (MML). Find this animation under the section "Process & Equipment III." To view other clusters or for more information about the MML visit http://matec.org/ps/library3/process_I.shtmlKey Phrase: MATEC Animation

  1. Suitlock Docking Mechanism

    NASA Technical Reports Server (NTRS)

    Culbertson, Philip, Jr. (Inventor)

    1997-01-01

    An environmental protective suit used for hazardous clean-up or space applications includes a suitlock docking mechanism that allows for easy egress and ingress of a crew member between a sealed vessel and a possibly contaminated environment. The suitlock docking mechanism comprises a single actuator that controls latches which, in turn, respectfully control rack and pinion assemblies that allow for easy removal and attachment of a life support equipment enclosure shell to the environmental protective suit or to the vehicle from which the operator performs his/her duties.

  2. Dual Quantum Mechanics

    E-print Network

    W. Chagas-Filho

    2009-05-11

    We point out a possible complementation of the basic equations of quantum mechanics in the presence of gravity. This complementation is suggested by the well-known fact that quantum mechanics can be equivalently formulated in the position or in the momentum representation. As a way to support this complementation, starting from the action that describes conformal gravity in the world-line formalism, we show that there are duality transformations that relate the dynamics in the presence of position dependent vector and tensor fields to the dynamics in the presence of momentum dependent vector and tensor fields.

  3. Mechanical Harvesting of Corn.

    E-print Network

    Sorenson, J. W. (Jerome Wallace); Smith, H. P. (Harris Pearson)

    1948-01-01

    - - TEXAS AGRICULTURAL EXPERIMENT STATION R. D. LEWIS, Director ' College Station, Texas BULLETIN 706 OCTOBER 1948 Mechanical Harvesting of Corn H. P. SMITH and J. W. SORENSON, JR. Department of Agricultural Engineering LlBRARY Atricaltr...~ial &$J~he~i~ai CallcgoofTexas Collbge Statign, Toxes THE TEXAS AGRICULTURAL AND MECHANICAL COLLEGE SYSTEM GIBB GILCHRIST, Chancellor P9-1048-6500-L180 [Blank Page in Original Bulletin] Preface 1~x2s farmers annually plant from 4% to 5 million acres...

  4. Reconstructing the Antikythera Mechanism

    NASA Astrophysics Data System (ADS)

    Freeth, Tony

    The Antikythera Mechanism is a geared astronomical calculating machine from ancient Greece. The extraordinary nature of this device has become even more apparent in recent years as a result of research under the aegis of the Antikythera Mechanism Research Project (AMRP) - an international collaboration of scientists, historians, museum staff, engineers, and imaging specialists. Though many questions still remain, we may now be close to reconstructing the complete machine. As a technological artifact, it is unique in the ancient world. Its brilliant design conception means that it is a landmark in the history of science and technology.

  5. Closed loop mechanical ventilation.

    PubMed

    Wysocki, Marc; Jouvet, Philippe; Jaber, Samir

    2014-02-01

    Mechanical ventilation is a sophisticated technique with very narrow therapeutic ranges i.e. highly efficient and able to keep alive the most severe patients, but with considerable side effects and unwanted complications if not properly and timely used. Computerized protocols, closed loop systems, decision support, all terms which need to be defined, may help making mechanical ventilation safer and more efficient. The present paper will provide a short overview on technical and engineering considerations regarding closed loop controlled ventilation as well as tangible clinical evidences supporting the previous statement. PMID:23564277

  6. Soil mechanics experiment

    NASA Technical Reports Server (NTRS)

    Mitchell, J. K.; Bromwell, L. G.; Carrier, W. D., III; Costes, N. C.; Houston, W. N.; Scott, R. F.

    1972-01-01

    The Apollo 15 soil-mechanics experiment has offered greater opportunity for study of the mechanical properties of the lunar soil than previous missions, not only because of the extended lunar-surface stay time and enhanced mobility provided by the lunar roving vehicle (rover), but also because four new data sources were available for the first time. These sources were: (1) the self-recording penetrometer (SRP), (2) new, larger diameter, thin-walled core tubes, (3) the rover, and (4) the Apollo lunar-surface drill (ALSD). These data sources have provided the best bases for quantitative analyses thus far available in the Apollo Program.

  7. PHYSICS 5706 --STATISTICAL MECHANICS 2: NONEQUILIBRIUM STATISTICAL MECHANICS

    E-print Network

    Täuber, Uwe Claus

    PHYSICS 5706 -- STATISTICAL MECHANICS 2: NONEQUILIBRIUM STATISTICAL MECHANICS Syllabus -- Fall 2014 ­ Quantum and Statistical Mechanics. Literature: The lectures will draw from original papers and several. Cowan, Topics in statistical mechanics (Imperial College, 2005) H. Haken, Synergetics (Springer, 3rd ed

  8. MECHANICAL ENGINEERING 121 Department of Mechanical Engineering (MEE)

    E-print Network

    Kostic, Milivoje M.

    MECHANICAL ENGINEERING 121 Department of Mechanical Engineering (MEE) *Available for general education credit. The Department of Mechanical Engineering offers an upper- division curriculum which leads to a B.S. in mechanical engineering. The curriculum is based on a strong foundation of fundamental

  9. 114 MECHANICAL ENGINEERING Department of Mechanical Engineering (MEE)

    E-print Network

    Kostic, Milivoje M.

    114 MECHANICAL ENGINEERING Department of Mechanical Engineering (MEE) *Available for general education credit. The Department of Mechanical Engineering offers an upper- division curriculum which leads to a B.S. in mechanical engineering. The curriculum is based on a strong foundation of fundamental

  10. Program Transformation Mechanics A Classification of Mechanisms for Program Transformation

    E-print Network

    Utrecht, Universiteit

    Program Transformation Mechanics A Classification of Mechanisms for Program Transformation with a Survey of Existing Transformation Systems Jonne van Wijngaarden Eelco Visser UU-CS-2003-048 Institute Transformation Mechanics A Classification of Mechanisms for Program Transformation with a Survey of Existing

  11. Nonlinear Solid Mechanics SOLID MECHANICS AND ITS APPLICATIONS

    E-print Network

    Nonlinear Solid Mechanics #12;SOLID MECHANICS AND ITS APPLICATIONS Volume 160 Series Editors: G and Scope of the Series The fundamental questions arising in mechanics are: Why?, How?, and How much and insight in answering these questions on the subject of mechanics as it relates to solids. The scope

  12. Quantum mechanical light harvesting mechanisms in photosynthesis

    NASA Astrophysics Data System (ADS)

    Scholes, Gregory

    2012-02-01

    More than 10 million billion photons of light strike a leaf each second. Incredibly, almost every red-coloured photon is captured by chlorophyll pigments and initiates steps to plant growth. Last year we reported that marine algae use quantum mechanics in order to optimize photosynthesis [1], a process essential to its survival. These and other insights from the natural world promise to revolutionize our ability to harness the power of the sun. In a recent review [2] we described the principles learned from studies of various natural antenna complexes and suggested how to utilize that knowledge to shape future technologies. We forecast the need to develop ways to direct and regulate excitation energy flow using molecular organizations that facilitate feedback and control--not easy given that the energy is only stored for a billionth of a second. In this presentation I will describe new results that explain the observation and meaning of quantum-coherent energy transfer. [4pt] [1] Elisabetta Collini, Cathy Y. Wong, Krystyna E. Wilk, Paul M. G. Curmi, Paul Brumer, and Gregory D. Scholes, ``Coherently wired light-harvesting in photosynthetic marine algae at ambient temperature'' Nature 463, 644-648 (2010).[0pt] [2] Gregory D. Scholes, Graham R. Fleming, Alexandra Olaya-Castro and Rienk van Grondelle, ``Lessons from nature about solar light harvesting'' Nature Chem. 3, 763-774 (2011).

  13. Mechanical engineering Department Seminar

    E-print Network

    The electric guitar is a fascinating convergence of engineering and music, science and art. ErgonomicallyMechanical engineering Department Seminar Kevin L. Wasson Corning Iinc. Engineering an electric engineering used to create, analyze, and refine the electric guitar. Structural design issues are presented

  14. Mechanical & Aerospace Engineering

    E-print Network

    Mechanical & Aerospace Engineering We present an arbitrary high-order quadrature- free, Runge (Herrmann, 2008). The accuracy, consistency, and the convergence of the resulting method is demonstrated spatial convergence for k- th order orthonormal Legendre polynomial basis functions. We furthermore show

  15. Viral Structure and Mechanics

    Microsoft Academic Search

    Sujit S. Datta

    The prospect of understanding and controlling the structure, formation and properties of nanoscale virus capsids has challenged researchers from disciplines as diverse as biology, chemistry, mechanical engineering and physics for over half a century. In this review, I will describe a number of theoretical approaches to this. The most well-known approach is that of Caspar and Klug, developed in the

  16. ANEUPLOIDY: ETIOLOGY AND MECHANISMS

    EPA Science Inventory

    The 'Symposium on Aneuploidy: Etiology and Mechanisms' was held from March 25-29, 1985. This Symposium developed as a consequence of the concern of the Environmental Protection Agency with the support of the National Institute of Environmental Health Sciences about human exposure...

  17. The Mechanization of Mining.

    ERIC Educational Resources Information Center

    Marovelli, Robert L.; Karhnak, John M.

    1982-01-01

    Mechanization of mining is explained in terms of its effect on the mining of coal, focusing on, among others, types of mining, productivity, machinery, benefits to retired miners, fatality rate in underground coal mines, and output of U.S. mining industry. (Author/JN)

  18. Mechanical Harvesting of Corn. 

    E-print Network

    Sorenson, J. W. (Jerome Wallace); Smith, H. P. (Harris Pearson)

    1948-01-01

    to deflect the ears and prevent them being caught and pinched by the flights on the snapping rollers. A change was made in the wagon elevator to save time and labor and to permit the ears to be dropped into a sack. The long MECHANICAL HARVESTING OF CORN...

  19. Memory Mechanisms in Grasping

    ERIC Educational Resources Information Center

    Hesse, Constanze; Franz, Volker H.

    2009-01-01

    The availability of visual information influences the execution of goal-directed movements. This is very prominent in memory conditions, where a delay is introduced between stimulus presentation and execution of the movement. The corresponding effects could be due to a decay of the visual information or to different processing mechanisms used for…

  20. Mechanical and Aerospace Engineering

    E-print Network

    On superhydrophobic surfaces, condensate drops spontaneously jump upon coalescence, giving rise to self-sustained dropwise condensation without any external driving forces. The self-propelled motion results from: a new self-cleaning mechanism for water-repellent surfaces that works without rain, and a planar phase

  1. Department of Mechanical Engineering

    E-print Network

    Li, Teng

    ). · Undergraduate students have been awarded 3 Goldwater Scholarships, 2 Truman Scholarships, 1 ChurchillDepartment of Mechanical Engineering 2014 Fast Facts Faculty Based on 2013 statistics from National Academy of Engineering Members 42 Professional Society Fellow Memberships 4 PECASE Awardees 16

  2. Mechanical engineering Department Seminar

    E-print Network

    Lin, Xi

    Mechanical engineering Department Seminar Ed Goluch DiPietro Assistant Professor Northeastern for studying biofilm formation and removal, and micro/nano-fluidic devices to trap and isolate exotic bacterial of Chemical Engineering at Northeastern University. Ed received his B.S. in Chemical Engineering from

  3. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar Russ Tedrake Department of Electrical Engineering is the X Consortium Associate Professor of Electrical Engineering and Computer Science and Aeronautics of the NSF CAREER Award, the MIT Jerome Saltzer Award for undergraduate teaching, the DARPA Young Faculty

  4. Electro-Mechanical Curriculum.

    ERIC Educational Resources Information Center

    EASTCONN Regional Educational Services Center, North Windham, CT.

    This electromechanical technician curriculum covers the following general areas: (1) basic soldering; (2) reading diagrams and following schematics; and (3) repairing circuitry and mechanics common to major appliances, vending machines, amusement equipment, and small office machines. The manual includes the following sections: (1) course…

  5. Implementation by Decent Mechanisms

    Microsoft Academic Search

    Jernej Copic; Clara Ponsati

    2002-01-01

    Abstract We address the design of optimal mechanisms for bargaining problems subject to incomplete information on the reservation shares of the agents. Decent rules are those that are Pareto Optimal in the constrained set of rules satisfying strategy proofness, individual rationality and weak eciency,- a requirement on ex post eciency.,We char- acterize decent rules and prove a uniqueness and existence

  6. Photovoltaic array loss mechanisms

    Microsoft Academic Search

    Charles Gonzalez

    1986-01-01

    Loss mechanisms which come into play when solar cell modules are mounted in arrays are identified. Losses can occur either from a reduction in the array electrical performance or with nonoptimal extraction of power from the array. Electrical performance degradation is caused by electrical mismatch, transmission losses from cell surface soiling and steep angle of reflectance, and electrical losses from

  7. Mechanical & Aerospace Engineering

    E-print Network

    Mechanical & Aerospace Engineering Nowadays automobile's technology is going under ever increasing's Advanced Vehicle Technology Competition "EcoCAR" at the Polytechnic School. Dr. Mayyas received his Ph as a senior automotive engineer at the transportation division of the Jordanian Police Department. During his

  8. Industrial Mechanical Maintenance.

    ERIC Educational Resources Information Center

    Hendrix, Laborn J.

    This manual was developed to assist teachers in Oklahoma in preparing students for industrial mechanical maintenance. The materials in this teacher's guide are organized in 14 units of instruction covering the following four areas: receiving and setting equipment; equipment hookup and operation; equipment layout, anchoring, and setup; and…

  9. Mechanical behavior of superalloys

    Microsoft Academic Search

    Floreen

    1986-01-01

    Recent developments affecting the mechanical behavior of superalloys over three ranges of operating temperatures are reviewed. At lower temperatures, activity has been focused on stress corrosion cracking susceptibility in light water reactor and sour gas well environments. The susceptibility to intergranular crack growth is critically dependent upon the grain boundary chemistry, and a method of minimizing the sensitivity of the

  10. Ribosomopathies: mechanisms of disease.

    PubMed

    Nakhoul, Hani; Ke, Jiangwei; Zhou, Xiang; Liao, Wenjuan; Zeng, Shelya X; Lu, Hua

    2014-01-01

    Ribosomopathies are diseases caused by alterations in the structure or function of ribosomal components. Progress in our understanding of the role of the ribosome in translational and transcriptional regulation has clarified the mechanisms of the ribosomopathies and the relationship between ribosomal dysfunction and other diseases, especially cancer. This review aims to discuss these topics with updated information. PMID:25512719

  11. Engine & Vehicle Mechanics Curriculum.

    ERIC Educational Resources Information Center

    Alaska State Dept. of Education, Juneau. Div. of Adult and Vocational Education.

    This competency-based curriculum includes all competencies a student will acquire in an engine and vehicle mechanics educational program. It follows guidelines established for automobile technician training programs leading toward certification and addresses requirements of the National Institute for Automotive Service Excellence (ASE). The…

  12. Diesel Mechanics: Fundamentals.

    ERIC Educational Resources Information Center

    Foutes, William; And Others

    This publication is the first in a series of three texts for a diesel mechanics curriculum. Its purpose is to teach the basic concepts related to employment in a diesel trade. Six sections contain 29 units. Each instructional unit includes some or all of these basic components: unit and specific (performance) objectives, suggested activities for…

  13. Turboprop Propulsion Mechanic.

    ERIC Educational Resources Information Center

    Chanute AFB Technical Training Center, IL.

    This instructional package consists of a plan of instruction, glossary, and student handouts and exercises for use in training Air Force personnel to become turboprop propulsion mechanics. Addressed in the individual lessons of the course are the following: common hand tools, hardware, measuring devices, and safety wiring; aircraft and engine…

  14. Mechanical engineering Department Seminar

    E-print Network

    Lin, Xi

    , University of Washington Acoustic scattering from mine-like objects in an underwater environment: ExperimentsMechanical engineering Department Seminar Aubrey Espańa Professor Applied Physics Laboratory Refreshments served at 10:45 AM Our work concerns the detection and classification of objects in an underwater

  15. Mechanical engineering Department Seminar

    E-print Network

    Career Development Associate Professor in Biomedical Engineering. She is a recipient of the Donald PMechanical engineering Department Seminar Domitilla Del Vecchio Department of Mechanical Engineering MIT A Control Theory Approach to Engineering Biomolecular Networks 11:00 AM Friday, November 2nd

  16. Mechanical engineering Department Seminar

    E-print Network

    Mechanical engineering Department Seminar Yi Zhao Department of Biomedical Engineering Ohio State in the Department of Biomedical Engineering, Ohio State University since 2006. Before then, he received B.S. and M University Electrospun/electrosprayed constructs for microscale cell/tissue engineering 11:00 AM Friday

  17. Mechanical engineering Department Seminar

    E-print Network

    Lin, Xi

    Mechanical engineering Department Seminar Junjie Niu Postdoctoral Associate MIT Engineering Nano+/e-. A low-cost Li-S battery with improved cyclability will also be discussed. Carbon sponge-like matrix and Engineering at Massachusetts Institute of Technology. He received Ph.D. in Materials Science from Zhejiang

  18. Horticultural Mechanics Competencies

    ERIC Educational Resources Information Center

    Shipley, W. Edward

    1974-01-01

    Ornamental horticulture teachers and managers of ornamental horticulture businesses were surveyed to determine which agricultural mechanics knowledges and skills are needed for entry-level employment in nursery, greenhouse, turf, and landscape management, which are common to the four areas, and the appropriate grade level at which they should be…

  19. Mechanisms of Salinity Tolerance

    Microsoft Academic Search

    Rana Munns; Mark Tester

    2008-01-01

    The physiological and molecular mechanisms of tolerance to os- motic and ionic components of salinity stress are reviewed at the cel- lular, organ, and whole-plant level. Plant growth responds to salinity in two phases: a rapid, osmotic phase that inhibits growth of young leaves, and a slower, ionic phase that accelerates senescence of ma- ture leaves. Plant adaptations to salinity

  20. Formic Acid Mechanical,

    E-print Network

    Illinois at Urbana-Champaign, University of

    Formic Acid Fire Ant Starch Mechanical, Industrial Chemical, Petroleum Biological What Do and engineering. Products will range from starch, to polylactic acid, to corn fiber, to motor fuels. Progressive systems. Fire ants make formic acid. U of I researchers are developing fuel cells that use formic acid (1

  1. Why Mechanical Engineering?

    E-print Network

    Li, Teng

    Engineering program to students in the Southern Maryland region in a convenient, student- friendly format provides a quality engineering education to the region's residents and builds technical expertiseWhy Mechanical Engineering? Do you have sharp analytical and problem- solving skills? Are you

  2. Interpretation of quantum mechanics

    Microsoft Academic Search

    Roland Omnčs

    1987-01-01

    New axioms are proposed for the interpretation of quantum mechanics. They rest on a kind of calculus allowing to select meaningful physical statements and giving rules to check a given physical reasoning containing implications. Measurement theory is reformulated. Laboratoire associé au Centre National de la Recherche Scientifique.

  3. Supersymmetry and quantum mechanics

    Microsoft Academic Search

    Fred Cooper; Avinash Khare; Uday Sukhatme

    1995-01-01

    In the past ten years, the ideas of supersymmetry have been profitably applied to many nonrelativistic quantum mechanical problems. In particular, there is now a much deeper understanding of why certain potentials are analytically solvable and an array of powerful new approximation methods for handling potentials which are not exactly solvable. In this report, we review the theoretical formulation of

  4. Ph 125 Quantum Mechanics

    NSDL National Science Digital Library

    Mabuchi, Hideo

    This web site contains resources for a comprehensive quantum mechanics course designed for graduate and advanced undergraduate students at Cal Tech. The course has been revised to include quantum information science, and prepares students for a course in quantum computation. Lecture notes, a syllabus, homework problems with solutions, and exam solutions are available.

  5. Imaging the Antikythera Mechanism

    SciTech Connect

    Malzbender, Tom (Hewlett Packard Laboratories) [Hewlett Packard Laboratories

    2011-01-12

    In 1900, a party of sponge divers chanced on the wreck of a Roman merchant vessel between Crete and mainland Greece. It was found to contain numerous ancient Greek treasures, among them a mysterious lump of clay that split open to reveal 'mathematical gears' as it dried out. This object is now known as the Antikythera Mechanism, one of the most enlightening artifacts in terms of revealing the advanced nature of ancient Greek science and technology. In 2005 we travelled to the National Archeological Museum in Athens to apply our Reflectance Imaging methods to the mechanism in the hopes of revealing ancient writing on the device. We were successful, and along with the results of Microfocus CT imaging, we are able to decipher 3000 characters compared with the original 800 known. This lead to an understanding that the device was a mechanical, astronomical computer from 150 B.C.E. capable of predicting solar and lunar eclipses along with other celestial events. This talk will overview both the imaging methods as well as what they reveal about the Antikythera Mechanism.

  6. Motorcycle Mechanic. Teacher Edition.

    ERIC Educational Resources Information Center

    Baugus, Mickey; Fulkerson, Dan, Ed.

    These teacher's materials are for a 19-unit competency-based course on entry-level motorcycle mechanics at the secondary and postsecondary levels. The 19 units are: (1) introduction to motorcycle repair; (2) general safety; (3) tools and equipment; (4) metric measurements; (5) fasteners; (6) service department operations; (7) motorcycle engines;…

  7. MECHANICAL TEST LAB CAPABILITIES

    E-print Network

    MECHANICAL TEST LAB CAPABILITIES · Static and cyclic testing (ASTM and non-standard) · Impact drop testing · Slow-cycle fatigue testing · High temperature testing to 2500°F · ASTM/ Boeing/ SACMA standard testing · Ability to design and fabricate non-standard test fixtures and perform non-standard tests

  8. Wear-mechanism modelling

    SciTech Connect

    Ashby, M.F. (Cambridge Univ. (United Kingdom). Dept. of Engineering)

    1993-03-01

    Goals of the program are to calculate the surface temperatures in dry sliding, develop a soft wear tester for ceramics, survey the wear mechanisms in brittle solids, and couple the temperature calculations with models to give wear maps for brittle solids. (DLC)

  9. Diesel Mechanics: Fuel Systems.

    ERIC Educational Resources Information Center

    Foutes, William

    This publication is the third in a series of three texts for a diesel mechanics curriculum. Its purpose is to teach the concepts related to fuel injection systems in a diesel trade. The text contains eight units. Each instructional unit includes some or all of these basic components: unit and specific (performance) objectives, suggested activities…

  10. Mechanism of tsunami earthquakes

    Microsoft Academic Search

    Hiroo Kanamori

    1972-01-01

    The mechanism of the Aleutian islands earthquake of 1946 and the Sanriku earthquake of 1896 is studied on the basis of the data on seismic waves from 5 to 100 s and on tsunamis. These earthquakes generated, despite their relatively small earthquake magnitude, two of the largest and most widespread tsunamis in history. The data obtained at different periods are

  11. celestial mechanics populations

    E-print Network

    Fisher, Todd

    celestial mechanics weather populations directions of research Dynamical Systems: Investigations into Chaos Todd Fisher tfisher@math.byu.edu Department of Mathematics Brigham Young University Brigham Young´e's paper there was a mistake found. He was told to buy back every copy of the journal containing his

  12. Mechanical & Aerospace Engineering

    E-print Network

    Shumway, John

    in the areas of pool and flow boiling, microchannel flows, electronics cooling, hydrogen energy, fuel cellsMechanical & Aerospace Engineering Pool boiling heat transfer enhancement is taking a dramatic turn/editor of several books/handbooks including Handbook of Phase Change: Boiling and Condensation published by Taylor

  13. Epigenetic mechanisms in schizophrenia.

    PubMed

    Roth, Tania L; Lubin, Farah D; Sodhi, Monsheel; Kleinman, Joel E

    2009-09-01

    Epidemiological research suggests that both an individual's genes and the environment underlie the pathophysiology of schizophrenia. Molecular mechanisms mediating the interplay between genes and the environment are likely to have a significant role in the onset of the disorder. Recent work indicates that epigenetic mechanisms, or the chemical markings of the DNA and the surrounding histone proteins, remain labile through the lifespan and can be altered by environmental factors. Thus, epigenetic mechanisms are an attractive molecular hypothesis for environmental contributions to schizophrenia. In this review, we first present an overview of schizophrenia and discuss the role of nature versus nurture in its pathology, where 'nature' is considered to be inherited or genetic vulnerability to schizophrenia, and 'nurture' is proposed to exert its effects through epigenetic mechanisms. Second, we define DNA methylation and discuss the evidence for its role in schizophrenia. Third, we define posttranslational histone modifications and discuss their place in schizophrenia. This research is likely to lead to the development of epigenetic therapy, which holds the promise of alleviating cognitive deficits associated with schizophrenia. PMID:19559755

  14. Mechanisms of restenosis.

    PubMed Central

    Casscells, W; Engler, D; Willerson, J T

    1994-01-01

    Restenosis after percutaneous transluminal coronary angioplasty remains a problem, which suggests that we still do not fully understand its mechanisms. We review here the current understanding of the cell biology of restenosis, including clinical correlation (risk factors), randomized clinical trials, human histology, animal models, and in vitro studies. Images PMID:8180514

  15. Sorghum Detoxification Mechanisms

    Microsoft Academic Search

    A. P. Pinto; A. de Varennes; M. L. S. Gonçalves; A. M. Mota

    2006-01-01

    The influence of cadmium (Cd) on internal and external defense mechanisms of Sorghum bicolor (L.) Moench. x Sorghum sudanense was studied by electrochemical methods to infer the type of detoxification processes developed by the plant under environmental stress conditions. Hydroponic experiments with sorghum were conducted in nutrient solutions in the absence and presence of 0.1 mg Cd L. Plant exposure

  16. Mechanical engineering Department Seminar

    E-print Network

    Lin, Xi

    and inspired their utilization in advanced materials. This talk will examine the fundamental mechanics Buckling & Snapping Structures for Advanced Functionality 11:00 AM Friday, 28 February 2014 Room 245, 110 did it provide func- tionality. An increasing focus on soft materials, from rubbers and gels

  17. Residential Mechanical Precooling

    SciTech Connect

    German, A.; Hoeschele, M.

    2014-12-01

    This research conducted by the Alliance for Residential Building Innovation team evaluated mechanical air conditioner pre-cooling strategies in homes throughout the United States. EnergyPlus modeling evaluated two homes with different performance characteristics in seven climates. Results are applicable to new construction homes and most existing homes built in the last 10 years, as well as fairly efficient retrofitted homes.

  18. Mechanical engineering Department Seminar

    E-print Network

    Lin, Xi

    Mechanical engineering Department Seminar Willie Padilla Associate Professor Boston College enabling energy harvesting applications. Willie Padilla received a physics PhD from the University to 2006. In 2006 Professor Padilla joined the Department of Physics at Boston College and is currently an

  19. Diesel Mechanics: Electrical Systems.

    ERIC Educational Resources Information Center

    Foutes, William; And Others

    This publication is the second in a series of three texts for a diesel mechanics curriculum. Its purpose is to teach the concepts related to electricity and circuitry in a diesel trade. The text contains nine units. Each instructional unit includes some or all of these basic components: unit and specific (performance) objectives, suggested…

  20. Quo Vadis Quantum Mechanics?

    Microsoft Academic Search

    W G Unruh

    2006-01-01

    Quantum mechanics is one of the most successful theoretical structures in all of science. Developed between 1925-26 to explain the optical spectrum of atoms, the theory over the succeeding 80 years has been extended, first to quantum field theories, gauge field theories, and now even string theory. It is used every day by thousands of physicists to calculate physical phenomena