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Brown, Shea Austin. "N-Acylethanolamines and Plant Phospholipase D." Thesis, University of North Texas, 1998. https://digital.library.unt.edu/ark:/67531/metadc279270/.
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Recently, three distinct isoforms of phospholipase D (PLD) were identified in Arabidopsis thaliana. PLD α represents the well-known form found in plants, while PLD β and γ have been only recently discovered (Pappan et al., 1997b; Qin et al., 1997). These isoforms differ in substrate selectivity and cofactors required for activity. Here, I report that PLD β and γ isoforms were active toward N-acylphosphatidylethanolamine (NAPE), but PLD α was not. The ability of PLD β and γ to hydrolyze NAPE marks a key difference from PLD α. N-acylethanolamines (NAE), the hydrolytic products of NAPE by PLD β and γ, inhibited PLD α from castor bean and cabbage. Inhibition of PLD α by NAE was dose-dependent and inversely proportional to acyl chain length and degree of unsaturation. Enzyme kinetic analysis suggested non-competitive inhibition of PLD α by NAE 14:0. In addition, a 1.2-kb tobacco (Nicotiana tabacum L.) cDNA fragment was isolated that possessed a 74% amino acid identity to Arabidopsis PLD β indicating that this isoform is expressed in tobacco cells. Collectively, these results provide evidence for NAE producing PLD activities and suggest a possible regulatory role for NAE with respect to PLD α.
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Hodson, Jane E. "Tobacco Phospholipase D β1: Molecular Cloning and Biochemical Characterization." Thesis, University of North Texas, 2002. https://digital.library.unt.edu/ark:/67531/metadc3341/.
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Transgenic tobacco plants were developed containing a partial PLD clone in antisense orientation. The PLD isoform targeted by the insertion was identified. A PLD clone was isolated from a cDNA library using the partial PLD as a probe: Nt10B1 shares 92% identity with PLDβ1 from tomato but lacks the C2 domain. PCR analysis confirmed insertion of the antisense fragment into the plants: three introns distinguished the endogenous gene from the transgene. PLD activity was assayed in leaf homogenates in PLDβ/g conditions. When phosphatidylcholine was utilized as a substrate, no significant difference in transphosphatidylation activity was observed. However, there was a reduction in NAPE hydrolysis in extracts of two transgenic plants. In one of these, a reduction in elicitor- induced PAL expression was also observed.
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Mebarek, Azzam Saida. "La Phospholipase D, une voie de signalisation lipidique impliquée dans de multiples fonctions cellulaires : morphologie, prolifération, différenciation." Lyon, INSA, 2006. http://theses.insa-lyon.fr/publication/2006ISAL0003/these.pdf.
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La phospholipase D (PLD), qui hydrolyse la phosphatidylcholine des membranes cellulaires et donne naissance à un médiateur phospholipidique, l'acide phosphatidique, joue un rôle central dans diverses fonctions cellulaires. Nous avons tout d'abord mis en évidence l'implication des céramides dans la différenciation des myoblastes de rat L6. Ces cellules sont capables de se différencier in vitro en myotubes polynucléés exprimant divers marqueurs du tissu musculaire, en présence de vasopressine. Cette hormone induit une régulation biphasique des taux cellulaires de céramides, avec une décroissance dans les premières heures, suivie d'une augmentation jusqu'au 8e jour, attribuable à l'activation de la voie de biosynthèse de novo. Les céramides ainsi formés ont un effet régulateur négatif sur la différenciation, au moins en partie à cause du contrôle négatif qu'ils exercent sur l'expression de l'isoforme PLD1, dont nous avions démontré le caractère indispensable à la myogénèse. Nous avons de plus observé que les céramides inhibent le réorganisation PLD1- dépendante du cytosquelette d'actine, une des premières étapes du processus myogénique. La régulation de la PLD par les céramides semble constituer une cible pharmacologique intéressante pour des actions visant à favoriser la régénération musculaire dans diverses situations pathologiques. Par ailleurs, nous avons montré que la PLD intervient dans le contrôle de la perméabilité paracellulaire d'une monocouche de cellules endothéliales HUV-EC-C aux macromolécules, grâce à ses effets sur le cytosquelette d'actine. Les lipoprotéines de faible densité (LDL), natives ou oxydées, mises en contact avec la monocouche, sont capables de stimuler l'activité PLD des cellules et la perméabilité aux macromolécules. Elles provoquent en parallèle un remodelage PLD-dépendant du cytosquelette avec la formation de fibres de stress, et un changement de morphologie cellulaire. Les LDL stimulent donc leur propre passage transendothélial, via leur capacité à réguler la PLD. Un défaut de régulation de la PLD pourrait contribuer à une accumulation subendothéliale anormale des LDL, et, étant donné le rôle proathérogène de ces molécules, à l'accélération du processus athéroscléreux. Des travaux antérieurs avaient établi la présence de l'isoforme PLD1 au niveau des radeaux lipidiques membranaires de lymphocytes périphériques humains, et suggéré un lien entre la délocalisation de l'enzyme, son activation, et une inhibition de la réponse aux mitogènes. Nous avons confirmé que la perturbation des radeaux, par des agents agissant spécifiquement sur les lipides de ce compartiment, est associée à une activation de la PLD et à une inhibition de la prolifération lymphocytaire. Par des expériences de surexpression, nous avons montré que l'isoforme PLD1, mas pas l'isoforme PLD2, est spécifiquement responsable d'un contrôle négatif de l'activation lymphocytaire. La mise en évidence de la régulation de PLD1 par inclusion / exclusion du compartiment radeaux lipidiques, et la démonstration de son implication dans le contrôle de la réponse lymphocytaire, devraient permettre de mieux comprendre les mécanismes moléculaires impliqués dans diverses pathologies de l'immunitéPhospholipase D (PLD) hydrolyses phosphatidylcholine of cell membranes in response to a variety of agonists, to generate phosphatidic acid, a second messenger implicated in cell functions such as cytoskeletal reorganization. In L6 skeletal myoblasts induced to differentiate, a short-lived lowering of ceramide levels was followed by a long-lasting elevation due to de novo synthesis. Ceramide mediates a negative control of myogenic differentiation, at least in part through down-regulation of PLD1 isoform expression and inhibition of PLD1-dependent formation of stress fibers. Moreover, we show that PLD is involved in paracellular permeability of endothelial cells through actin cytoskeleton reorganization, and morphological changes. In addition, we show that disruption of membrane lipid rafts by agents specifically active on the lipids of this compartment, induces an activation of PLD and generates anti-proliferative signals in lymphocytes
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Rahier-Corticchiato, Renaud. "Caractérisation biochimique des phospholipases D et de leurs domaines fonctionnels : nouvelle méthode de mesure de l’activité phospholipase D." Thesis, Lyon, 2016. http://www.theses.fr/2016LYSE1292/document.
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La phospholipase D (PLD) hydrolyse les phospholipides membranaires en libérant leur tête polaire afin de générer l'acide phosphatidique (PA), impliqué dans la signalisation cellulaire. Pour comprendre les propriétés biochimiques des PLDs, les travaux présentés ont été réalisés autour de deux axes. Le premier axe concerne l'expression recombinante et la purification de la PLDa d'Arabidopsis thaliana (AtPLDa) dans la levure Pichia pastoris. La détermination de la séquence N-terminale a révélé que l'AtPLDa est amputée de ses 35 premiers résidus, suggérant ainsi la participation d'un mécanisme de maturation. Cependant, la région N-terminale des PLDs de plantes est homologue au domaine C2, impliqué dans leur interaction Ca2+-dépendante avec la membrane. Afin d'évaluer l'impact d'un tel clivage, les domaines C2 de l'AtPLDa mais également de l'AtPLDß, à titre de comparaison, ont été étudiés sous leur forme entière ou mature. Ainsi, la caractérisation de leur affinité pour les phospholipides, associée à leur modélisation tridimensionnelle, ont permis de démontrer que les différences de régulation par le Ca2+, observées entre les formes entières et mature, provenait de la présence d'une hélice a amphipathique, retirée lors du processus de maturation. Le second axe concerne le développement d'une nouvelle méthode de mesure des activités PLD via le dosage de manière direct, spécifique et continu du PA grâce à la propriété d'amplification de fluorescence par chélation de la 8-hydroxyquinoléine, en présence de Ca2+. Ainsi, ce test apparait adapté pour le suivi de l'inhibition des PLDs et pour l'étude de leur spécificité de substrat, en utilisant des phospholipides naturels avec différentes tête polaires, et à l'échelle d'une microplaquePhospholipase D (PLD) hydrolyses membrane phospholipids, leading to the formation of free polar headgroup and phosphatidic acid releasing, involved in cell signaling. To understand the biochemical properties of PLDs, this work has been made around two axes. The one first concerns the recombinant expression and purification of the PLDa of Arabidopsis thaliana (AtPLDa) in the yeast Pichia pastoris. The N-terminal sequence of the recombinant AtPLDa has been determined and found to lack its first 35 amino acids, suggesting the involvement of a maturing mechanism. However, plant PLDs exhibit a C2-lipid binding domain at their N-terminal region, which is involved in their Ca2+-dependent membrane targeting. Thus, to assess the impact of such a cleavage, whole and mature-like C2 domains of AtPLDa, as well as of AtPLDß, for the sake of comparison were studied. Thus, the characterization of their affinity for phospholipids, combined with their three-dimensional modeling have demonstrated that the differences observed in their regulation by Ca2+, observed between whole and mature-like forms, originated from the presence of a N-terminus amphipathic a helix, removed during the maturation process. The second axis concerns the development of a novel PLD assay that measure PA in a direct, specific and continuous manner, using the chelation enhanced fluorescence property of 8-hydroxyquinoline in the presence of Ca2+. Thus, this assay appears suitable for monitoring both the inhibition of PLDs as well as their substrate specificity, using natural phospholipids with different polar headgroups, and at a microplate scale
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Li, Liang. "Regulation of phospholipase D in submandibular glands." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape4/PQDD_0018/NQ53062.pdf.
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Lee, Jung Hoon. "Suppression of phospholipase D[Alpha] in soybean." Diss., Manhattan, Kan. : Kansas State University, 2008. http://hdl.handle.net/2097/828.
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McKinnon, Murray. "Studies on mammalian phosphatidylcholine specific phospholipase D." Thesis, Open University, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.315440.
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Skafi, Najwa. "Role of Phospholipase D in Vascular Calcification." Thesis, Lyon, 2017. http://www.theses.fr/2017LYSE1339/document.
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La calcification vasculaire est l’accumulation de cristaux de calcium dans les vaisseaux sanguins à travers un processus pathologique qui ressemble à la formation de l’os ou du cartilage. Elle apparaît notamment chez les patients diabétiques ou atteints d’une insuffisance rénale chronique. La conséquence principale de la calcification vasculaire est la perte de l’élasticité qui est indispensable pour la fonction des larges artères, elle est de plus associée à la mortalité des patients hémodialysés. Les traitements contre la calcification vasculaire sont généralement limités à ceux qui corrigent les facteurs causatifs des problèmes de santé mais aucune intervention efficace, spécifique et ciblée n’est disponible. Par conséquence, une compréhension profonde des mécanismes moléculaires impliqués dans la calcification vasculaire est nécessaire dans le but de trouver de nouvelles cibles thérapeutiques. La phospholipase D catalyse l’hydrolyse des phospholipides en acide phosphatidique et une tête polaire, elle est aussi impliquée dans différentes fonctions cellulaires et maladies. Il a été démontré qu’elle peut être activée par des facteurs impliqués dans l’ostéogenèse et par d’autres impliqués dans la calcification vasculaire. Ainsi, nous avons étudié le rôle de la phospholipase D dans la calcification vasculaire dans 3 modèles différents. Le premier est un modèle in-vitro de cellules musculaires lisses murines (lignée cellulaire MOVAS), elles sont cultivées en présence d’acide ascorbique et de β-glycérophosphate. Le deuxième est un modèle ex-vivo d’explants d’aortes cultivés en présence de fortes concentrations de phosphate et le troisième est un modèle in-vivo d’insuffisance rénale chronique produite chez des rats. Dans ce dernier modèle, la calcification vasculaire est induite par un régime riche en phosphore et en calcium et par des injections de vitamine D active. La calcification dans ces trois modèles a été suivie par l’analyse de la minéralisation en dosant les dépôts de calcium, de l’activité phosphatase alcaline, et de l’expression de différents marqueurs ostéo-chondrocytaires. Une augmentation de l’expression génique de Pld1 a été observée dans les trois modèles, en particulier au cours des premières étapes de la calcification, et a été accompagnée d'une activité accrue de la phospholipase D dans les modèles in vitro et ex-vivo. L’inhibition de l’activité phospholipase D dans ces deux modèles ou de la phospholipase D1 dans le modèle MOVAS a bloqué complètement la calcification. Par contre, l’inhibition spécifique de la phospholipase D2 n’a pas montré des effets significatifs. Deux voies par lesquelles la phospholipase D peut être activée ont été testées, la voie de la protéine kinase C et la voie de la sphingosine-1-phosphate. Ces deux voies métaboliques se sont révélées être impliquées dans le processus de calcification mais pas forcément dans l’activation de la phospholipase D au cours de ce processus. Des résultats préliminaires ont montré que la phospholipase D pourrait agir après activation de la sphingosine kinase 2 dont l’activité s’est avérée nécessaire pour la calcification dans le modèle MOVAS. Des études supplémentaires sont nécessaires pour comprendre par quels mécanismes la phospholipase D est activée et comment elle agit. La phospholipase D pourrait être une nouvelle cible thérapeutique pour le traitement de la calcification vasculaire vu que son inhibition ne semble pas avoir des effets secondaires chez les patientsVascular calcification is the accumulation of calcium phosphate crystals in blood vessels via a pathological process that resembles physiological bone or cartilage formation. Calcification in the medial layer is mainly seen in diabetic and chronic kidney disease patients. Its main consequence is the loss of elasticity which is indispensable for the function of large arteries. Accordingly, vascular medial calcification was significantly associated with mortality in hemodialysis patients. Vascular calcification treatments are limited to those that correct its causative health problems, but no efficient, specific and targeted interventions are available. Therefore, a deep understanding of its molecular mechanisms is needed to find novel therapeutic targets. Phospholipase D catalyses the hydrolysis of phospholipids into phosphatidic acid and a head group. It is implicated in different cellular functions and diseases. It was found to be activated by factors involved in osteogenesis and others involved in vascular calcification. Thus, we investigated its role in vascular calcification in 3 models: an in-vitro model of murine smooth muscle cell line MOVAS cultured with ascorbic acid and β-glycerophosphate, an ex-vivo model of rat aortas cultured in high phosphate medium, and an in-vivo model of adenine-induced kidney disease in rats in which vascular calcification is induced by further administration of high phosphorus/calcium diet and active vitamin D injections. Calcification was detected in these models using different approaches including alkaline phosphatase activity, calcium dosage, and/or evaluation of osteo-chondrocytic markers expression. Pld1 expression was seen upregulated in all the three models, especially during early stages of calcification, and was accompanied with increased phospholipase D activity in the in-vitro and ex-vivo model. The inhibition of total phospholipase D activity in these two models, or that of phospholipase D1 in case of MOVAS model, abolished calcification. Phospholipase D2-specific inhibition did not induce significant effects. Two pathways by which phospholipase D can be activated were tested, protein kinase C and sphingosine 1-phosphate pathways, but they were found to be involved in calcification but not necessary for phospholipase D activation during this process. Alternatively, the preliminary results showed that PLD may be acting by activation of sphingosine kinase 2 whose activity was found necessary for calcification in the MOVAS model. Further investigations are needed to understand the mechanisms by which phospholipase D is activated and by which it is acting. Phospholipase D could be a novel target for vascular calcification especially that its inhibition in patients did not induce adverse health effects
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Arhab, Yani. "Caractérisation structurale et fonctionnelle des phospholipases D." Thesis, Lyon, 2018. http://www.theses.fr/2018LYSE1225/document.
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Les phospholipases D (PLD, EC 3.1.4.4) sont des enzymes ubiquitaires retrouvées aussi bien chez les procaryotes (bactéries) que chez les eucaryotes (plantes, animaux et champignons). Les PLD catalysent l'hydrolyse des glycérophospholipides au niveau distal de la liaison phosphodiester pour former de l'acide phosphatidique, un important messager cellulaire impliqué dans de nombreuses voies telles que la prolifération cellulaire, la formation et le trafic vésiculaire, mais aussi la transcription et la survie cellulaire. Les PLD appartiennent à une superfamille de protéines (superfamille des PLD) qui ont en commun un site catalytique HXKX4D, X étant un acide aminé quelconque, contenant les résidus H (Histidyl), K (Lysyl) et D (Aspartyl). Ce site est nommé séquence consensus "HKD" et est dupliqué dans la plupart des membres de la superfamille des PLD. L'étude des PLD de plante est le moyen le plus sûr d'étudier cette famille d'enzyme car ce sont les seules PLD eucaryotiques purifiées à homogénéité et en grande quantité à ce jour. Ces travaux proposent une caractérisation fonctionnelle des résidus conservés au sein des PLD végétales menant à une caractérisation structurale avec la cristallisation de cette protéine. Dans un second temps l'activité de l'enzyme est modulée avec l'étude du domaine minimum, de la maturation post-traductionnelle de l'enzyme et le recherche d'un nouvel inhibiteur. Enfin, nous proposons le clonage d'une nouvelle PLD et la mise au point d'un système de détection in vivo de l'activité PLDPhospholipases D (PLD, EC 3.1.4.4) are ubiquitary enzymes found in prokaryotes (bacteria) as well as in eukaryotes (plant, animals and fungi). PLD catalyzes the hydrolysis of the distal phosphoester bound of phospholipids thus forming phosphatidic acid, an important cell signaling messenger implicated in numerous pathways such as cell proliferation, vesicular formation and trafficking but also transcription and cell survival. PLDs belong to a superfamily of protein which share a common catalytic site called “HKD†for HXKX4D, X is a random amino acid, containing H (Histidyl), K (lysyl) and D (aspartyl) residues. This consensus sequence is duplicated in most of the PLD superfamily members. The study of plant PLD is the best way to understand this family of proteins as they are the sole eukaryotic PLDs to be purified to homogeneity so far. This work provides a functional characterization of the most conserved residues in plant PLDs leading to a structural characterization with the crystallization of this enzyme. A second part of this work proposes the modulation of the enzyme hydrolysis activity by studying the minimal domain necessary for the activity and post-translational maturation undergone by plant PLDs. Also, we look for a new specific inhibitory molecule. Finally, we propose the cloning of a new plant PLD and the development of a new way to detect in vivo PLD activity
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Heung, Yen Ming Mary. "Molecular selectivity of phospholipase D in granulocyte function." Thesis, University of Southampton, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.241935.
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Zeiller, Caroline. "Phospholipase D, perméabilité endothéliale, et apoptose TNFα dépendante." Lyon, INSA, 2007. http://theses.insa-lyon.fr/publication/2007ISAL0087/these.pdf.
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La phospholipase D (PLD) est une enzyme membranaire qui intervient au cœur du métabolisme lipidique en générant un second messager, l’acide phosphatidique (PA) impliqué dans de nombreuses fonctions cellulaires. Deux isoformes PLD1 et PLD2 contribuent à sa synthèse chez les mammifères. Nous avons montré que la PLD augmente la perméabilité de monocouches de cellules endothéliales (lignée HUV-EC-C) aux macromolécules en permettant un remodelage du cytosquelette d’actine. L’isoforme PLD2 jouerait un rôle prépondérant via sa localisation au sein des cavéoles membranaires. Nous avons étudié le rôle de la PLD dans l’apoptose des cellules ECV304 induite par le TNFα, une cytokine activant à la fois des voies de survie et des voies pro-apoptotiques. Nous avons aussi étudié le rôle de la PLD dans l’apoptose des cellules ECV304 induite par le TNFα une cytokine activant à la fois des voies de survie et des voies pro-apoptotiques. Nous avons montré que la PLD a un rôle protecteur contre la mort cellulaire induite par TNFα en présence de cycloheximide, l’isoforme PLD1 étant plus spécifiquement mise en jeu. Divers mécanismes expliquant cet effet protecteur sont proposésPLD is a membrane-bound enzyme which plays a key role in lipid metabolism by generating phosphatidic acid, an anionic phospholipid involved in many cellular functions. Two isoforms PLD 1 and PLD2 exist in mammals. We have shown that PLD enhances the permeability of endothelial cell monolayers (HUV-EC-C cells) through an actin reorganization which is characterized by synthesis of stress fibers. PLD2 might be more particularly implicated, because of its sub cellular localization to membrane caveolae. We also studied the role of PLD in Tumor Necrosis Factor alpha (TNFα)-induced apoptosis of ECV304 cells. TNFα, a pleiotropic cytokine, activates both apoptotic and pro-survival signals depending on the cell model. We showed that PLD exerts a protective effect against cell death induced by TNFα in the presence of an inhibitor of protein synthesis, cycloheximide. PLD 1 isoform plays a predominant role in this process. Different mechanisms explaining the protective role of PLD are proposed
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Kusner, David John. "Regulation of phospholipase D activity in U937 cells." Case Western Reserve University School of Graduate Studies / OhioLINK, 1994. http://rave.ohiolink.edu/etdc/view?acc_num=case1057945948.
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Fang, Mimi. "The role of phospholipase d in osteoblasts in response to titanium surfaces." Thesis, Atlanta, Ga. : Georgia Institute of Technology, 2008. http://hdl.handle.net/1853/26462.
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Thesis (M.S.)--Biomedical Engineering, Georgia Institute of Technology, 2009.Committee Chair: Boyan, Barbara; Committee Member: Eskin, Suzanne; Committee Member: Lobachev, Kirill; Committee Member: Schwartz, Zvi. Part of the SMARTech Electronic Thesis and Dissertation Collection.
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Raymond, Frank Damian. "Inositol specific phospholipase D activity : a GPI-cleaving enzyme." Thesis, University College London (University of London), 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.321569.
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McCulloch, Derek A. "Activation of phospholipase D by G protein-coupled receptors." Thesis, University of Edinburgh, 1999. http://hdl.handle.net/1842/22467.
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The activation of phospholipase D (PLD) by the G protein-coupled receptor (GPCR) family was investigated with special attention paid to the interaction of receptors with small G proteins such as ADP-ribosylation factor (ARF) and RhoA. Agonist-stimulated activation of PLD by the M3 muscarinic, B2 bradykinin and H1 histamine receptors was sensitive to brefeldin A (BFA), an inhibitor of guanine-nucleotide exchange on ARF. In contrast the thrombin and thromboxane A2 receptors stimulated a PLD response insensitive to BFA. The Rho inhibitor C3 exoenzyme from Clostridium botulinum and a negative functional construct of RhoA markedly reduced PLD activity stimulated by the M3 muscarinic but not the thrombin receptor. The receptors investigated which couple to PLD by a mechanism involving ARF all contain in their seventh transmembrane helix (TM7), the amino acid sequence AsnProXXTyr (where X is any amino acid). In contrast, the receptors such as thrombin which activate PLD in an ARF-independent manner have an AspProXXTyr motif. The importance of this highly conserved motif was further indicated by studies on the mouse gonadotropin-releasing hormone (GnRH) receptor which contains the AspProXXTyr sequence and a mutant form of the GnRH receptor where the aspartate at position 318 is replaced with an asparagine thereby restoring the AsnProXXTyr motif present in the majority of group I GPCRs. The wild-type GnRH receptor stimulated PLD in a BFA-insensitive manner, while the Asn318 mutant GnRH form gained BFA-sensitivity. The co-immunoprecipitation of the agonist-treated Asn318 mutant but not wild-type GnRH receptor using antibodies raised to ARF 1/3 and RhoA indicated that a direct physical association between the receptor and small G proteins occurs and appears to be dependent on the presence of the AsnProXXTyr motif.
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Pisch-Heberle, Sandra. "Untersuchungen zur Stabilisierung von Membranproteinen mit ungewöhnlichen Phospholipiden." [S.l.] : Universität Stuttgart , Fakultät Chemie , Institut für Technische Biochemie, 2000. http://www.bsz-bw.de/cgi-bin/xvms.cgi?SWB8619088.
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Jaafar, Rami. "Régulation des la voie mTOR par la phospholipase D dans le muscle squelettique : implication dans le contrôle de la différenciation myogénique et de la taille des myocytes." Phd thesis, INSA de Lyon, 2011. http://tel.archives-ouvertes.fr/tel-00679827.
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La phospholipase D (PLD) hydrolyse la phosphatidylcholine des membranes cellulaires, libérant le messager acide phosphatidique. La capacité de la PLD à influer sur la voie de signalisation de mTOR, acteur central dans le contrôle du tissu musculaire, nous a incités à étudier son rôle dans ce tissu. Mes travaux de thèse ont pour but d'étudier les mécanismes par lesquels la PLD intervient dans la différenciation myogénique et dans la régulation de la masse musculaire. Dans un premier temps, nous avons montré que le contrôle de la différenciation des myoblastes L6 par la PLD met en jeu l'activation des deux complexes de mTOR (mTORC1 et mTORC2). mTORC2 active la différenciation, probablement via son effecteur PKCalpha, alors que mTORC1 la réprime via son effecteur S6K1, en induisant la phosphorylation de rictor, un composant de mTORC2, et l'inhibition de ce complexe. Nous avons par ailleurs montré que l'extinction de PLD par interférence de I'ARN induit l'atrophie de myotubes L6 en culture, ainsi qu'une baisse de la phosphorylation de S6K1 et 4E-BP1, effecteurs de mTORC1. Inversement, la surexpression de PLD à l'aide de vecteurs adénoviraux induit une hypertrophie des myotubes, associée à une activation de la voie mTORC1, et de Akt, effecteur de mTORC2. De plus, la surexpression de PLD atténue l'atrophie induite par la dexaméthasone. Ces résultats mettent en évidence un rôle hypertrophique et anti-atrophique de la PLD, qui pourrait s'exercer par stimulation de la voie mTOR. Nos résultats suggèrent que la PLD est susceptible de jouer un rôle clé dans le muscle squelettique, en agissant tant au niveau de la régénération du tissu qu'au niveau de la régulation de sa masse.
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Grant, Margaret Rose. "Characterisation of the pleckstrin homology (PH) domain of phospholipase D." Thesis, University of Birmingham, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.397782.
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Knapek, Katie J. "The Role of Phospholipase D (PLD) and Grb2 in Chemotaxis." Wright State University / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=wright1230574811.
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Ganesan, Ramya. "Phospholipase D: Key Player in Macrophage-mediated Inflammation and Resolution." Wright State University / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=wright1516049400177206.
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Yozwiak, Michael Leo 1963. "Effect of Corynebacterium pseudotuberculosis phospholipase D on ovine neutrophil function." Thesis, The University of Arizona, 1990. http://hdl.handle.net/10150/277329.
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Corynebacterium pseudotuberculosis causes caseous lymphadenitis of sheep and goats and produces a phospholipase D (PLD) exotoxin which is putatively important in pathogenesis. Viability and function of ovine polymorphonuclear leukocytes (PMN) treated with crude and purified forms of PLD were determined by various assays. PMN viability by dye exclusion showed the PLD-treatment had a significant effect only after 24 hour incubation. Scanning electron microscopy of PLD-treated ovine erythrocytes revealed membrane alterations, but no such alterations were seen in PLD-treated PMN. Transmission electron microscopy revealed significantly fewer granules in PMN treated with PLD, although other facets of phagocytic function appeared to be normal, including phagosome-lysosome fusion. PLD-treated PMN were significantly reduced in their ability to internalize Corynebacterium pseudotuberculosis and to attach to or phagocytose Staphylococcus epidermidis. Purified PLD activated normal sheep serum, producing chemotactic factors. PLD treatment of PMN significantly reduced the ability of these cells to migrate toward activated sheep serum.
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Tanguy, Emeline. "Implication de l’acide phosphatidique dans le trafic membranaire : rôle et régulation de la phospholipase D au cours de la phagocytose et de l’exocytose régulée." Thesis, Strasbourg, 2019. http://www.theses.fr/2019STRAJ112.
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La mise en évidence du rôle des lipides dans le trafic membranaire fait partie des avancées majeures de ces dernières années. Mes travaux de thèse se sont focalisés sur le plus simple des phospholipides, l’acide phosphatidique (PA). La production de PA par la phospholipase D (PLD) joue un rôle crucial au cours de la phagocytose et l’exocytose régulée, mais la dynamique de sa formation, tout comme les mécanismes d’action des différentes formes de PA, restent à ce jour totalement inconnus. Durant mon doctorat, j’ai contribué à la caractérisation de trois domaines de liaison peptidique au PA, qui nous ont permis de mieux comprendre les mécanismes d’interaction des protéines avec le PA et de générer des sondes moléculaires pour repérer ce lipide au sein des cellules. Ainsi, j’ai pu visualiser la production de PA au cours de la phagocytose et mis en évidence l’implication de la GTPase Arf6 dans la régulation de la synthèse de PA par la PLD. J’ai également pu montrer que la PLD est impliquée dans plusieurs étapes de l’exocytose dans des cellules neuroendocrines. Des expériences de lipidomique et de restauration ont révélé notamment que des formes mono- et polyinsaturées du PA contrôlent des étapes distinctes de l’exocytose que j’ai pu définirThe discovery of the involvement of lipids in membrane trafficking is one of the major recent progress in cell biology. My thesis work focused on phosphatidic acid (PA), the simplest phospholipid. PA synthesis by phospholipase D (PLD) plays a crucial role during phagocytosis and regulated exocytosis, but its precise dynamics, as well as the mode of action of the different PA species, remain unknown. I characterized three PA binding domains allowing a better understanding of the interaction between proteins and PA and leading to the generation of genetic sensors for PA in cells. Thus I could visualize PA synthesis during phagocytosis and identified that the small GTPase Arf6 regulates PLD activity and consequently PA synthesis. My work also reveals that PLD modulates several steps during exocytosis in neuroendocrine cells. Further lipidomics and rescue experiments allowed me to show that mono- and polyunsaturated forms of PA are involved in distinct steps of exocytosis
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Petersen, Gitte. "Characterization and partial purification of N-acylethanolamine phospholipid-hydrolyzing phospholipase D /." Kbh. : Danmarks Farmaceutiske Højskole, 2001. http://www.dfh.dk/phd/defences/gittepedersen.html.
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Parre, Elodie. "La signalisation lipidique et le métabolisme de la proline en réponse à des contraintes hydriques : rôles des phospholipases C et D chez Arabidopsis thaliana." Paris 6, 2008. http://www.theses.fr/2008PA066213.
Full textAbstract:
Parmi les stratégies adaptatives que les plantes ont développées pour faire face aux contraintes hydriques, l’accumulation de proline est une réponse physiologique fréquemment observée chez un grand nombre d’espèces. Alors que le métabolisme de ce soluté compatible a été relativement bien caractérisé, les voies de signalisation impliquées dans sa régulation restent en grande partie inconnues. Par une approche pharmacologique, nous avons mis en évidence l’existence d’un contrôle positif de la biosynthèse de la proline par les phospholipases C – et la synthèse d’ IP3 – en réponse au NaCl et non au mannitol. D’autre part, nos données montrent l’importance du niveau de calcium intracellulaire et non de la signature calcique lors de la réponse prolinique. Au cours de ces dernières années, nous avons montré que la biosynthèse de la proline était régulée négativement par une activité PLD en conditions de croissance favorable. À la lumière de ces résultats, nous avons concentré nos efforts sur les PLD1 et PLD2 qui, parmi les douze isoformes de PLD végétales, sont les seules présentant une activité calcium-indépendante. Par une approche de génétique inverse, nous avons obtenu et caractérisé des mutants d’insertion pour les gènes PLD1 et PLD2. Bien que les niveaux de proline chez ces mutants ne soient pas affectés, ils présentent néanmoins des phénotypes très intéressants au niveau de la capacité de germination et de croissance racinaire en conditions de stress hydrique. Enfin, ce travail nous a permis de montrer que les plantes sont capables de discerner un stress salin d’un stress hyperosmotique par des voies de signalisation spécifiques.
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Fennell, Myles. "Activation of phospholipase D by the LHRH receptor and associated mechanisms." Thesis, University of Edinburgh, 1995. http://hdl.handle.net/1842/28004.
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The αT3-1 gonadotroph-derived cell line was used as a model system to study intracellular signalling systems associated with the LHRH receptor, with particular interest in phospholipase D (PLD) and mechanisms involved in its regulation, such as protein kinase C (PKC) and non-receptor tryosine kinases. Phospholipase D activation in αT3-1 cells proceeded after a delay when stimulated by LHRH, or by the phorbol ester, phorbol 12,13-dibutyrate (PDBu). LHRH-stimulated PLD activity was fully resistant to desensitisation over a 40 min time course. The Ca2+ ionophore, ionomycin was unable to stimulate PLD activity. LHRH-stimulated PLD activity was inhibited by PKC downregulation or by bisindolylmaleimide PKC inhibitors at an approximately 8-fold higher concentration than that seen for the PDBu-stimulated PLD activity. Another PKC inhibitor, H7, inhibited PDBu-stimulated PLD activity in a manner that indicated multiple components to the inhibition and inhibited LHRH-stimulated PLD activity with a low potency. This is consistent with the possibility that one mediator of the LHRH response is a form of PKC which is relatively resistant to certain PKC inhibitors and insensitive to phorbol esters, or that some other unknown kinase is involved. The tyrosine kinase inhibitors, lavendustin A and genistein were able to selectively inhibit the LHRH-stimulated PLD activity. Phospholipase D activation stimulated by Gi and Go are involved. LHRH induced tyrosine phosphorylation of a number of proteins in αT3-1 cells; a similar phosphorylation profile was also produced by stimulation with PDBu.
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Devlin, Marni Allison. "The characterization of TSH-mediated phospholipase D activity in thyroid cells." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape9/PQDD_0017/MQ47020.pdf.
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Flores, Borja Fabian. "Characterisation of the mouse glycosylphosphatidyl inositol phospholipase D (GPI-PLD) gene." Thesis, University College London (University of London), 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.404723.
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Morgan, Clive Paul. "ARF regulated phospholipase D : role in cell signalling and membrane traffic." Thesis, University College London (University of London), 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.288004.
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Fite, Kristen. "Dysregulation of Phospholipase D (PLD) isoforms increases breast cancer cell invasion." Wright State University / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=wright149557402792618.
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Heller, Manfred. "Characterization of glycosylphosphatidylinositol-specific phospholipase D from mammalian liver and serum /." [S.l.] : [s.n.], 1994. http://www.ub.unibe.ch/content/bibliotheken_sammlungen/sondersammlungen/dissen_bestellformular/index_ger.html.
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Xie, Mingsheng. "The regulation and functional significance of phospholipase D in HL-60 cells." Case Western Reserve University School of Graduate Studies / OhioLINK, 1992. http://rave.ohiolink.edu/etdc/view?acc_num=case1056134184.
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Riebeling, Christian. "Mechanisms of regulation of phospholipase D isoforms involvement in differentiation and apoptosis /." [S.l.] : [s.n.], 2001. http://www.diss.fu-berlin.de/2001/100/index.html.
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Ufer, Guido [Verfasser]. "Proteins under the control of phospholipase D in Arabidopsis thaliana / Guido Ufer." Bonn : Universitäts- und Landesbibliothek Bonn, 2016. http://d-nb.info/1132711495/34.
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Uddin, Mohib. "Role of priming in the regulation of phospholipase D in human neutrophils." Thesis, University of Cambridge, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.620569.
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Yamamoto, Izumi. "Studies on Activation and Inhibition of Phospholipase D in Small Unilamellar Vesicles." 京都大学 (Kyoto University), 2000. http://hdl.handle.net/2433/157221.
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本文データは平成22年度国立国会図書館の学位論文(博士)のデジタル化実施により作成された画像ファイルを基にpdf変換したものであるKyoto University (京都大学)
0048
新制・論文博士
博士(薬学)
乙第10300号
論薬博第624号
新制||薬||183(附属図書館)
UT51-2000-C67
(主査)教授 半田 哲郎, 教授 中川 照眞, 教授 多賀 徹
学位規則第4条第2項該当
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McNamara, Peter Joseph. "Attenuation of Corynebacterium pseudotuberculosis by targeted mutagenesis of the phospholipase D gene." Diss., The University of Arizona, 1994. http://hdl.handle.net/10150/186732.
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A toxic phospholipase D (PLD), is thought to be an essential virulence determinant of Corynebacterium pseudotuberculosis, an animal pathogen which causes chronic, suppurative infections. In this study, PLD was purified from C. pseudotuberculosis biovar equi isolate 155 and the gene encoding PLD (pld) was cloned, pld from 155 and C. pseudotuberculosis biovar ovis isolate Whetten 1 were sequenced, Pld⁻ mutants of Whetten 1 were constructed, and one mutant was tested for reduced virulence for goats. PLD was purified by size exclusion chromatography and isoelectric focusing. In the purest fraction, sphingomyelinase, synergistic hemolysis, and staphylococcal β-hemolysin inhibition activities were associated with the presence of 31 and 21 kDa proteins. Only the 31 kDa protein was identified by Western blot analysis with PLD-neutralizing antibodies. Similar 1.6 kb inserts with pld from 155 and Whetten 1, in plasmids pCpE13s and pCpO51s respectively, conferred all three PLD-associated activities on E. coli. DNA sequence analysis revealed that both inserts encode a 307 amino acid protein which, in secreted form, has a predicted a molecular weight of 31.1 kDa and a pI of 8.84. The primary structure of the corynebacterial PLDs were compared to those produced by Corynebacterium ulcerans and Arcanobacterium haemolyticum. The four PLDs were found to shared 64% to 97% identity, and except for small regions of amino acid homology which includes two catalytic domains from glyceraldehyde-3-phosphate dehydrogenases, the sequences were unique when compared to other sequences found in GenBank. Two Pld⁻ strains of Whetten 1 were engineered by allele replacement. In strain W1.23r1, pld was deleted and in strain W1.31r1, pld contains a nonsense mutation which is predicted to truncate PLD after 65 amino acids. The virulence of W1.31r1 was compared to that of Whetten 1 by inoculation of goats. Whetten 1 caused abscessation at the site of infection and spread to the regional lymph node, while W1.31r1 had reduced ability to establish a primary infection and was incapable of dissemination. These results confirm that PLD is a virulence determinant of C. pseudotuberculosis which allows the persistence and spread of the bacteria within the host.
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Abdallah, Dina. "Fonctions de la phospholipase D et des récepteurs de la prostaglandine PGE2 durant la maturation des ostéoblastes, le processus de la minéralisation physiologique et la calcification cardiovasculaire." Thesis, Lyon 1, 2014. http://www.theses.fr/2014LYO10152/document.
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Le métabolisme lipidique affecte la maturation et la différenciation des cellules osseuses. L'objectif de ma thèse est d'approfondir deux aspects du métabolisme lipidique mal connus, soit les actions de la phospholipase D (PLD) et celles des récepteurs de prostaglandine PGE2 pendant la différenciation des cellules. Une lignée humaine, les Saos-2 et les ostéoblastes primaires issus de calvaria de souriceaux ont servi de modèles cellulaires de la minéralisation physiologique. La culture d'aorte ex vivo sous des conditions d'hyperphosphatémie a été utilisée pour reproduire la calcification de l'aorte qui est un modèle ex vivo de calcification cardiovasculaire (CCV). Nous avons montré que l'expression et l'activité de la PLD augmentent dans les Saos-2 et les ostéoblastes primaires au bout du 5ème jour de la différenciation tandis qu'elles s'accroissent au bout du 6ème jour de traitement de l'aorte dans un milieu d'hyperphosphatémie. Les inhibiteurs de PLD diminuent l'activité de phosphatase alcaline (TNAP) dans les ostéoblastes et dans l'aorte calcifiée tandis que la surexpression de la PLD1 dans les Saos-2 l'augmente. Dans une deuxième partie de ce travail, nous avons suivi la variation d'expression des récepteurs de PGE2 au cours de la maturation des Saos-2. L'expression du gène EP3 augmente au stade tardif de la minéralisation tandis que celle d'EP4 diminue. Pour conclure, ces résultats indiquent que l'activité de la PLD en affectant l'activité de la TNAP pourrait moduler finement la minéralisation physiologique et la CCV et que la minéralisation s'accompagne d'un changement d'expression des récepteurs de PGE2, dans les Saos-2Lipid metabolism affects the maturation and the differentiation of bone cells. The aim of my PhD thesis is to explore two unknown sides of lipid metabolism which are the actions of phospholipase D (PLD) and those of prostaglandin PGE2 receptors during cell differentiation. Human lineage, Saos-2 cells and primary osteoblasts from calvaria of mice were used as cellular models of physiological mineralization. The ex vivo aorta culture under hyperphosphatemia conditions has been used to reproduce the calcification of the aorta, which is an ex vivo model of cardiovascular calcification (CVC). We showed that the expression and the activity of PLD increased in Saos-2 and primary osteoblasts after the fifth day of differentiation while in the aorta under hyperphosphatemia condition, PLD activity increased at the end of the sixth day. PLD inhibitors decreased the activity of alkaline phosphatase (TNAP) in osteoblasts and in calcified aorta while the overexpression of PLD1 in the Saos-2 increased it. In the second part of this work, we monitored the variation of the expression of PGE2 receptors during the maturation of Saos-2 cells. The EP3 gene expression increased in the late stage of the mineralization while that of EP4 decreased. In conclusion, these results indicated that the PLD activity by affecting the activity of TNAP could modulate the physiological mineralization and CVC. We showed that the mineralization is dependent of the change of the expression of PGE2 receptors in Saos-2 cells
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Hagele, Thomas. "Mercury activates phospholipase D in vascular endothelial cells Implications for environmental cardiovascular disease /." Connect to resource, 2006. http://hdl.handle.net/1811/6486.
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Thesis (Honors)--Ohio State University, 2006.Title from first page of PDF file. Document formatted into pages: contains 47 p.; also includes graphics. Includes bibliographical references (p. 26-30). Available online via Ohio State University's Knowledge Bank.
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Gargett, Caroline Eve, and mikewood@deakin edu au. "Studies of the human lymphocyte P2Z receptor and its activation of phospholipase D." Deakin University, 1997. http://tux.lib.deakin.edu.au./adt-VDU/public/adt-VDU20060727.144101.
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Extracellular adenosine 5′-triphosphate (ATP) is an agonist for the P2Z receptor of human leukaemic lymphocytes and opens a Ca 2+-selective ion channel, which also conducts Ba2+, Sr2+ and the small fluorescent dye, ethidium+. A wide range of receptor agonists, many of which raise cytosolic [Ca2+] activate phospholipase D (PLD). In the present study, it was shown that both ATP and 3′-O-(4-benzoylbenzoyl)-ATP (BzATP) stimulated PLD activity in a concentration-dependent manner, and the inhibitory effects of suramin, oxidised ATP, extracellular Na+ and Mg2+ suggested that the effect of these agonists is mediated by P2Z receptors.
The role of divalent cations in ATP-stimulated PLD activity was investigated. Several agonists (eg ATP, thapsigargin, ionomycin) stimulated a rise in cytosolic [Ca2+] in human lymphocytes, but only ATP and ionomycin stimulated PLD activity. When Ca2+ influx was prevented by EGTA, the majority of ATP-stimulated and all of ionomycin-stimulated PLD activity was inhibited. Preloading cells with the Ca2+ chelator, BAPTA, reduced cytosolic [Ca2+] and, paradoxically, ATP-stimulated PLD activity was potentiated. ATP-stimulated PLD activity was supported by both Ba2+ and Sr2+ when they were substituted for extracellular Ca2+. Furthermore, both ATP-stimulated PLD activity and ATP-stimulated 133Ba2+ influx showed a linear dependence on extracellular [Ba2+]. Thus it was concluded that ATP stimulated PLD activity in direct proportion to the influx of divalent cations through the P2Z ion channel and this PLD activity was insensitive to changes in bulk cytosolic [Ca2+].
The calmodulin (Ca2+/CaM) inhibitor, trifluoperazine (TFP) inhibited ionomycin- and ATP-stimulated PLD activity and ATP-stimulated apoptosis, but had no effect on PLD activity already activated by ATP. However, TFP inhibited ATP-stimulated Ca2+, Ba2+ and ethidium+ fluxes, at concentrations below those which inhibit Ca2+/CaM, suggesting that TFP inhibits the P2Z receptor.
Similarly, the isoquinolinesulphonamide, KN-62, a selective inhibitor of Ca2+/CaM-dependent protein kinase II (CaMKII), also prevented ATP-stimulated apoptosis, but had no effect on pre-activated PLD. In addition, KN-62, and an analogue, KN-04, which has no effect on CaMKII, potently inhibited ATP-stimulated Ba2+ influx (IC50 12.7 ± 1.5 and 17.3 ± 2.7 nM, respectively), ATP-stimulated ethidium+ uptake (IC50 13.1 ± 2.6 and 37.2 ± 8.9 nM, respectively), ATP-stimulated phospholipase D activity (50% inhibition 5.9 ± 1.2 and 9.7 ± 2.8 nM, respectively) and ATP-induced shedding of the surface adhesion molecule, L-selectin (IC50 31.5 ± 4.5 and 78.7 ± 10.8 nM, respectively). They did not inhibit phorbol ester- or ionomycin-stimulated PLD activity or phorbol ester-induced L-selectin shedding. Neither KN-62 nor KN-04 (both 500 nM) have any effect on UTP-stimulated Ca2+ transients in fura-2-loaded human neutrophils, a response which is mediated by the P2Y2 receptor, neither did they inhibit ATP-stimulated contractile responses mediated by the P2X1 receptor of guinea pig urinary bladder. Thus, KN-62 and KN-04 are almost equipotent as P2Z inhibitors with IC50s in the nanomolar, indicating that their actions cannot be due to CaMKII inhibition, but rather that they are potent and direct inhibitors of the P2Z receptor.
Extracellular ATP-induced shedding of L-selectin from lymphocytes into the medium is a Ca2+-independent response. L-selectin is either cleaved by a metalloproteinase or a PLD with specificity for glycosylphosphatidylinositol (GPI). The novel hydroxamic acid-based zinc chelator, Ro-31-9790 blocks ATP-induced L-selectin shedding, but was without effect on ATP-induced Ba2+ influx or ATP-stimulated PLD activity. Furthermore, another zinc chelator, 1,10-phenanthroline, an inhibitor of a GPI-PLD, potentiated rather than inhibited ATP-stimulated PLD activity, suggesting that ATP-induced L-selectin shedding and ATP-stimulated PLD activity are independent of each other.
Although extracellular ATP is the natural ligand for the lymphocyte P2Z receptor, it is less potent than BzATP in stimulating Ba2+ influx. Concentration-response curves for BzATP- and ATP-stimulated ethidium+ influx gave EC50s 15.4 ± 1.4 µM and 85.6 ± 8.8 µM, respectively. The maximal response to ATP was only 69.8 ± 1.9% of that for BzATP. Hill coefficients were 3.17 ± 0.24 and 2.09 ± 0.45 for BzATP and ATP respectively, suggesting greater positive cooperativity for BzATP than for ATP in opening the P2Z-operated ion channel. A rank order of agonist potency of BzATP > ATP = 2MeSATP > ATPγS was observed for agonist-stimulated ethidium+ influx, while maximal influxes followed a rank order of BzATP > ATP > 2MeSATP > ATPγS. When ATP (300 -1000 µM) was added simultaneously with 30 µM BzATP (EC90), it reduced both ethidium+ and Ba2+ fluxes by 30 - 40% relative to values observed with BzATP alone. KN-62, previously shown to be a specific inhibitor of the lymphocyte P2Z receptor, was a less potent antagonist of BzATP-induced fluxes than ATP, when maximal concentrations of both agonists (50 and 500 µM respectively) were used. However, when BzATP (18 µM) was used at a concentration equiactive with a maximally effective ATP concentration, KN-62 showed the same inhibitory potency for both agonists. The ecto-ATPase antagonist, ARL-67156, inhibited both ATP- and BzATP-stimulated Ba2+ influx, suggesting that the lower efficacy of ATP compared with BzATP was not due to preferential hydrolysis of ATP. Thus, the natural ligand, ATP, is a partial agonist for the P2Z receptor while BzATP is a full agonist. Moreover the competitive studies show that only a single class of P2-receptor (P2Z class) is expressed on human leukaemic lymphocytes.
Both ATP- and BzATP-stimulated PLD activity were significantly inhibited (P < 0.05) when cells were suspended in iso-osmotic choline Cl medium. Choline+ was found to be a permeant for the P2Z ion channel, since ATP induced a large uptake of [14C]choline+ (60 to 150 µmol/ml intracellular water) during a 5 min incubation, which remained in the cells for several hours, and ATP was used to load cells with these levels of choline+. Intracellular choline+ inhibited ATP-, BzATP-, PMA- and ionomycin-stimulated PLD activity. Brief exposure of lymphocytes to ATP increased the subsequent basal rate of ethidium+ uptake, and this was prevented by intracellular choline+. It is proposed that P2Z-mediated Ca2+ influx in lymphocytes activates PLD leading to significantly changes of the phospholipid composition of the plasma membrane, which subsequently produces a permeability lesion, which in turn contributes to cell death.
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Yu, Yuanyuan. "Investigation of potential roles of Phospholipase D in Arabidopsis thaliana seed oil accumulation." Thesis, University of British Columbia, 2009. http://hdl.handle.net/2429/14835.
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Most plants depend on the production of storage compounds in the seed to provide
nutrients during germination. These storage compounds have been exploited by man for a
variety of purposes. For example vegetable oil is an important lipid source for cooking
and the production of lubricants, inks, paints, and biodiesels. Almost all seed oil is stored in the chemical form of triacylglycerol (TAG), and in mature seed, TAG is stored in densely packed oil bodies. Genetic engineering offers great potential to increase the amount and/or manipulate the type of oil produced. There are two routes to this goal: first, the development of existing plant species that already synthesize the desired novel oil into “alternative oilseed’ crops; second, the engineering of existing crops through the
cloning of genes which determine the synthesis of novel oils and their introduction into existing crops such as oilseed rape (Brassica napus) by genetic transformation. Whichever approach is adopted we need to know how storage oil is accumulated in order to optimize yield.
My research is part of a broader effort to explore the possibility of increasing oil content in Brassica species using Arabidopsis thaliana as a model system. My specific project examines whether the phospholipase D zeta (PLDZ) enzyme can be used to control oil production in Arabidopsis thaliana. In plants, PLDZ hydrolyzes membrane phospholipids
to form phosphatidic acid, which is the branch point for the synthesis of diacyiglycerol that is an immediate precursor of storage oil. In addition, previous research had shown that the PLDZ gene was negatively regulated in the leaves by GLABRA 2 (GL2), a transcription factor that was also known to negatively regulate oil content in the seed. In this thesis I show that the pldzlpldz2 double mutant accumulates 11% less oil in the seed
than did wild type, without differences in seed size, plant growth or development. Both
PLDZ genes localize to the embryo and funiculus during the later stages of seed
development. In the g12 mutant background, PLDZ transcript levels in the seeds are lower than in the wild type; besides the pldz]pldz2gl2 triple mutant shows intermediate oil content which is unlike neither pldzlpldz2 double mutant nor g12 mutant phenotype, suggesting that GL2 is not a negative regulator of PLDZ genes during oil production process.
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Whitby, Helen Elizabeth. "The role of glycosylphospatidyl inositol phospholipase D (GDI PLD) in type one hypersensitivity." Thesis, University College London (University of London), 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.251585.
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Planat, Valérie. "Étude des voies de transduction régulant l'activité phospholipase D dans le neutrophile humain." Toulouse 3, 1996. http://www.theses.fr/1996TOU30128.
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En situation inflammatoire ou infectieuse le polymorphonucléaire neutrophile est la première cellule à intervenir dans la défense de l'organisme. Divers messagers extracellulaires sont présents sur le site inflammatoire et susceptibles d'activer le neutrophile, pour induire des réponses cellulaires comme l'adhésion, le chimiotactisme, la bactéricidie ou la production d'ions superoxydes. L'enchainement des réactions cellulaires, depuis la fixation de l'agoniste sur son récepteur membranaire jusqu'à la réponse finale constitue la transduction du signal. Un partenaire récent de la transduction du signal est la phospholipase d (pld). La première partie de ce travail concerne la recherche de nouveaux agonistes de l'activité pld du neutrophile. Cette étude a montré que le peptide chimiotactique fmlp et l'ester de phorbol pma activent une pld, alors que le platelet derived growth factor, l'interleuline 8 et les immunoglobulines g aggregées sont inefficaces. De plus, nous montrons l'absence de récepteurs au pdgf dans le neutrophile. Une deuxième partie a été consacrée à l'étude des modes de régulation des voies de transduction impliquant la pld lors de l'activation du neutrophile. Dans divers modèles cellulaires, il a été rapporté la participation de protéines g trimériques et monomériques, de protéines serine-threonine kinases, de protéines tyrosine kinases et de divers cofacteurs (calcium, phospholipides, acides gras). Il nous a été possible de distinguer dans le neutrophile, d'une part l'activité stimulée par le fmlp, dépendante du calcium, de protéines g et de protéines tyrosine kinases, et d'autre part l'activité pld stimulée par le pma, impliquant la protéine kinase c seule. Les mitogen-activated-protein kinases, activées par phosphorylation par le fmlp et le pma, ne semblent pas impliquées. Des inhibiteurs, capables de bloquer l'activité de la nadph oxydase ont été testés sur l'activité pld: la wortmannine, un inhibiteur de la phosphatidylinositol 3-kinase, et l'oxyde de phenylarsine, un inhibiteur de protéines tyrosines phosphatases. La sensibilite à ces inhibiteurs permet de distinguer trois niveaux d'activité pld pour chacun des deux agonistes, réprésentant en fait une fraction de l'activité globale dans la cellule
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McHugh, John. "Molecular and biochemical characterization of phospholipase D in cotton (Gossypium hirsutum L) seedlings." Thesis, University of North Texas, 2005. https://digital.library.unt.edu/ark:/67531/metadc4732/.
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N-Acylethanolamines (NAEs) are enriched in seed-derived tissues and are believed to be formed from the membrane phospholipid, N-acylphosphatidylethanolamine (NAPE) via the action of phospholipase D (PLD). In an effort to identify a functional NAPE-PLD in cotton seeds and seedlings, we have screened a cotton seedling cDNA (cotyledon mRNA from 48 h dark grown seedlings) library with a 1.2 kb tobacco partial cDNA fragment encoding the middle third of a putative PLDβ/γ (genbank accession, AF195614) isoform. Six plaques were isolated from the Uni-ZAP lambda library, excised as pBluescript SK(-) phagemids and subjected to nucleotide sequence analysis. Alignment of derived sequences with Arabidopsis PLD family members indicated that the cDNAs represent six different PLD gene products -three putative PLD β isoforms and three putative PLD δ isoforms. The PLD β isoforms, designated Ghpldβ1a, GHpldβ1b and a truncated Ghpldβ1b isoform. Both the full-length PLD β proteins contained characteristic HKxxxxD catalytic domains, a PC-binding domain, a PIP2-binding domain and a C2 domain. In addition both cotton PLD β isoforms had a N-terminal "SPQY" rich domain which appeared to be unique to these PLDs. The three PLD δ isoforms, designated Ghpldδ1a, Ghpldδ1b and Ghpldδ1b-2 encode full-length PLDδ proteins, and like the above PLDs, contained the characteristic catalytic and regulatory domains. The expression of Ghpldδ1b showed hydrolytic and transphosphatidylation activity toward radiolabelled phosphatidylcholine (PC) but it appears Ghpldδ1b does not utilize NAPE as a substrate to produce NAEs nor does it seem to be suppressed by NAEs.
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Qin, Chunbo. "Cloning, expression and characterization of multiple forms of phospholipase D in Arabidopsis thaliana /." Search for this dissertation online, 2003. http://wwwlib.umi.com/cr/ksu/main.
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Bühler, Anja [Verfasser]. "Molecular mechanisms regulating phospholipase C-gamma 2 activity / Anja Bühler." Ulm : Universität Ulm. Fakultät für Naturwissenschaften, 2016. http://d-nb.info/1094889202/34.
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Schäffner, Ines. "Identifizierung und rekombinante Herstellung von Phospholipase D-Isoenzymen aus Weisskohl (Brassica oleracea var. capitata)." [S.l. : s.n.], 2001. http://deposit.ddb.de/cgi-bin/dokserv?idn=963174665.
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Yu, Changhua. "Phospholipase D/phosphatidic acid phosphatase signal transduction pathway in post-infarction congestive heart failure." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq23565.pdf.
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Ha, V. L. "The coordinated regulation of phospholipase D by ADP-ribosylation factors and their exchange factors." Thesis, University College London (University of London), 2004. http://discovery.ucl.ac.uk/1446607/.
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Phospholipase D (PLD) is a phospholipid hydrolyzing enzyme, the activation of which has effects on cellular events including cell growth and membrane trafficking in mammalian cells. In a reconstitution assay consisting of permeabilised HL-60 cells (human myeloid leukemic cells), experiments confirmed that members of the ADP- ribosylation factor (ARF) family proteins including ARFl and ARF6 were efficient PLD activators. However, ARFl was a stronger activator of PLD than ARF6, a result that was also found in in vitro PLD assays. Moreover, the myristoylated amino terminal ?-helix of ARF was essential for the activation of PLD. The activation of ARF is in turn regulated by a specific family of small guanine nucleotide exchange factors (GEFs) comprising ARNO, GRP-1 and cytohesin-1. The GEFs catalyze the release of bound GDP and its replacement by GTP, leading to ARF activation. Using the reconstitution assay, the PLD activation mediated by ARF was enhanced by the GEFs. However, the GEFs did not improve the stimulation of PLD by ARF in vitro. Interestingly, these GEFs activated PLD with high potency but none of the three GEFs was more potent than the others in their regulation of PLD in cell- based assays. It seems that this pattern of PLD activation does not reflect differential interactions with major phosphoinositides (PIP2 and PIP3) since these molecules bind equally well to the GEFs. Importantly, PIP2 and PIP3 increased the potency of PLD activation mediated by the coordinated actions of ARF and its exchange factors indicating the involvement of other pathways in the regulation of PLD in vivo. In particular, it emerged that PIP3 is a more potent activator of PLD than PIP2 in the presence and absence of ARF-GEFs.
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Guillemain, Isabelle. "Mode d'action des neurokinines et de divers neurotransmetteurs au niveau de la glande parotide de rat. Regulation de la phospholipase c et de la phospholipase d." Paris 11, 1994. http://www.theses.fr/1994PA112303.
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Dans la premiere partie de notre travail, nous avons caracterise le type de recepteurs des neurokinines impliques dans le mode d'action de la substance p au niveau de la glande parotide de rat. Nous avons etudie les effets d'agonistes selectifs de ces recepteurs des neurokinines sur la production des inositol phosphates et sur les mouvements de calcium. Nous avons montre que seuls les recepteurs nk-1 sont couples a l'activation de la voie de transduction calcium/phospholipides. Dans la seconde partie de notre travail, nous avons demontre pour la premiere fois la presence d'une activite phospholipase d au niveau des acini de glande parotide, en mettant en evidence la reaction de transphosphatidylation catalysee par cet enzyme, a savoir la formation de phosphatidylethanol a partir d'un phospholipide substrat, en presence d'ethanol. Nos resultats indiquent que la phospholipase d peut etre regulee differemment par les agonistes muscariniques et alpha adrenergiques par une voie dependante du calcium et par les esters de phorbol par une voie independante du calcium. L'utilisation de differents inhibiteurs de la proteine kinase c ne nous a pas permis de demontrer l'implication de cet enzyme lors de l'activation de la phospholipase d. Par ailleurs, nous avons observe que l'activation de la phospholipase d ne resulte en aucune production detectable d'acide phosphatidique. Compte tenu de l'ensemble de ces resultats, nous suggerons que l'activation d'une phospholipase d reflete, dans les cellules acineuses de glande parotide, une voie metabolique qui permettrait un echange de base entre des phospholipides et serait impliquee dans le renouvellement du pool de phosphatidylinositol lors d'une stimulation soutenue de la phospholipase c par les agonistes muscariniques et alpha adrenergiques
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Cai, Jingfei. "Probing the Membrane Association Mechanisms for Pulmonary Collectins and Mammalian Phospholipase C." Thesis, Boston College, 2013. http://hdl.handle.net/2345/3872.
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Thesis advisor: Mary F. RobertsThesis advisor: Eranthie Weerapana
Peripheral proteins from mammals often exhibit multi-domain structures and require metal ions such as calcium as co-factors. This dissertation investigates two types of such proteins -- pulmonary collectins (surfactant proteins A and D) and phosphatidylinositol-specific phospholipase C (PLC) delta1 -- and their interactions with model membranes. One approach to work around the complexity brought upon by such multi-domain protein structure is to use a truncated construct or an isolated single domain. For pulmonary collectins, homotrimers consisting of the neck domain and the carbohydrate recognition domain were used in a novel NMR assay for better understanding of their lipid-specific interactions with the membranes. For PLC delta1, we were particularly interested in the role of the EF-hand domain. The isolated EF-hand domain of PLC delta1 was first used to characterize its interactions with membranes and identify key residues responsible for such interactions. These key residues in the N terminal lobe of the EF-hand domain, either cationic or hydrophobic, were then found to affect the hydrolysis activity of the full-length enzyme. A common role for this region of the PLC in facilitating proper membrane association was thus proposed
Thesis (PhD) — Boston College, 2013
Submitted to: Boston College. Graduate School of Arts and Sciences
Discipline: Chemistry