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Molecular Biology of the Cell Mar 2024Phospholipase D1 (PLD1) activity is essential for the stimulated exocytosis of secretory vesicles where it acts as a lipid-modifying enzyme to produces phosphatidic acid...
Phospholipase D1 (PLD1) activity is essential for the stimulated exocytosis of secretory vesicles where it acts as a lipid-modifying enzyme to produces phosphatidic acid (PA). PLD1 localizes to the plasma membrane and secretory vesicles, and PLD1 inhibition or knockdowns reduce the rate of fusion. However, temporal data resolving when and where PLD1 and PA are required during exocytosis is lacking. In this work, PLD1 and production of PA are measured during the trafficking, docking, and fusion of secretory vesicles in PC12 cells. Using fluorescently tagged PLD1 and a PA-binding protein, cells were imaged using TIRF microscopy to monitor the presence of PLD1 and the formation of PA throughout the stages of exocytosis. Single docking and fusion events were imaged to measure the recruitment of PLD1 and the formation of PA. PLD1 is present on mobile, docking, and fusing vesicles and also colocalizes with Syx1a clusters. Treatment of cells with PLD inhibitors significantly reduces fusion, but not PLD1 localization to secretory vesicles. Inhibitors also alter the formation of PA; when PLD1 is active, PA slowly accumulates on docked vesicles. During fusion, PA is reduced in cells treated with PLD1 inhibitors, indicating that PLD1 produces PA during exocytosis.
Topics: Rats; Animals; Phosphatidic Acids; Biological Transport; Cell Membrane; Secretory Vesicles; Phospholipase D; Exocytosis
PubMed: 38117597
DOI: 10.1091/mbc.E23-05-0189 -
Plant Physiology Oct 2005
Review
Topics: Calcium; Cell Death; Cytoskeleton; GTP-Binding Proteins; Phosphatidic Acids; Phospholipase D; Plant Cells; Plant Development; Plant Proteins; Plants; Reactive Oxygen Species
PubMed: 16219918
DOI: 10.1104/pp.105.068809 -
International Journal of Molecular... Feb 2022Salt stress is a major adverse abiotic factor seriously affecting fruit tree growth and development. It ultimately lowers fruit quality and reduces yield....
Salt stress is a major adverse abiotic factor seriously affecting fruit tree growth and development. It ultimately lowers fruit quality and reduces yield. Phosphatidylcholine (PC) is an important cell membrane component that is critical for cell structure and membrane stability maintenance. In this study, we found that the addition of external PC sources significantly increased the tolerance of one-year-old peach trees, (L.) Batsch., to salt stress and attenuated their damage. The effect of exogenous application of 200 mg/L PC exerted the most significant positive effect. Its use caused seedling leaf stomatal opening, contributing to normal gas exchange. Moreover, beneficial effects were exerted also to the root system, which grew normally under salt stress. Meanwhile, phospholipase D activity in the cell was promoted. The production of phosphatidic acid (PA) was enhanced by increased decomposition of phospholipids; PA serves as a secondary messenger involved in plant biological process regulation and the reduction in the reactive oxygen species- and peroxide-induced damage caused by salt stress. The possible mechanism of action is via promoted plant osmotic regulation and tolerance to salt stress, reducing salt stress-induced injury to plants.
Topics: Cell Membrane; Gene Expression Regulation, Plant; Homeostasis; Phosphatidic Acids; Phosphatidylcholines; Salt Stress; Seedlings; Stress, Physiological
PubMed: 35269728
DOI: 10.3390/ijms23052585 -
The Journal of Biological Chemistry Jul 1999In the present study, we report that phosphatidic acid (PA) functions as a novel, potent, and selective inhibitor of protein phosphatase 1 (PP1). The catalytic subunit...
In the present study, we report that phosphatidic acid (PA) functions as a novel, potent, and selective inhibitor of protein phosphatase 1 (PP1). The catalytic subunit of PP1alpha was inhibited by PA dose-dependently in a noncompetitive manner with a K(i) value of 80 nM. The inhibition by PA was specific to PP1 as PA failed to inhibit protein phosphatase 2A (PP2A) or PP2B. Furthermore, PA was the most effective and potent inhibitor of PP1 compared with other phospholipids. Because we recently showed that ceramides activated PP1, we next examined the effects of PA on ceramide stimulation of PP1. PA inhibited both basal and ceramide-stimulated PP1 activities, and ceramide showed potent and stereoselective activation of PP1 in the presence of PA. Next, the effects of PA on ceramide-induced responses were examined. Molt-4 cells took up PA dose- and time-dependently such that by 1 and 3 h, uptake of PA was 0.37 and 0. 65% of total PA added, respectively. PA at 30 microM and calyculin A at 10 nM (an inhibitor of PP1 and PP2A at low concentrations), but not okadaic acid at 10 nM (a PP2A inhibitor at low concentrations) prevented poly(ADP-ribose) polymerase proteolysis induced by C(6)-ceramide. Moreover, the combination of PA with okadaic acid prevented retinoblastoma gene product dephosphorylation induced by C(6)-ceramide. These data suggest that PA functions as a specific regulator of PP1 and may reverse or counteract those effects of ceramide that are mediated by PP1, such as apoptosis and retinoblastoma gene product dephosphorylation.
Topics: Animals; Ceramides; Dose-Response Relationship, Drug; Enzyme Inhibitors; Humans; Jurkat Cells; Phosphatidic Acids; Phosphoprotein Phosphatases; Protein Phosphatase 1; Protein Phosphatase 2; Rabbits; Retinoblastoma Protein
PubMed: 10409693
DOI: 10.1074/jbc.274.30.21335 -
The Journal of Biological Chemistry Apr 2013Phosphatidic acid (PA) is a class of lipid messengers involved in a variety of physiological processes. To understand how PA mediates cell functions in plants, we used a...
Phosphatidic acid (PA) is a class of lipid messengers involved in a variety of physiological processes. To understand how PA mediates cell functions in plants, we used a PA affinity membrane assay to isolate PA-binding proteins from Camelina sativa followed by mass spectrometric sequencing. A cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) was identified to bind to PA, and detailed analysis was carried out subsequently using GAPC1 and GAPC1 from Arabidopsis. The PA and GAPC binding was abolished by the cation zinc whereas oxidation of GAPCs promoted the PA binding. PA had little impact on the GAPC catalytic activity in vitro, but the PA treatment of Arabidopsis seedlings induced proteolytic cleavage of GAPC2 and inhibited Arabidopsis seedling growth. The extent of PA inhibition was greater in GAPC-overexpressing than wild-type seedlings, but the greater PA inhibition was abolished by application of zinc to the seedling. The PA treatment also reduced the expression of genes involved in PA synthesis and utilization, and the PA-reduced gene expression was partially recovered by zinc treatment. These data suggest that PA binds to oxidized GAPDH and promotes its cleavage and that the PA and GAPC interaction may provide a signaling link coordinating carbohydrate and lipid metabolism.
Topics: Arabidopsis; Arabidopsis Proteins; Carbohydrate Metabolism; Cytoplasm; Glyceraldehyde-3-Phosphate Dehydrogenases; Lipid Metabolism; Phosphatidic Acids; Proteolysis; Seedlings; Zinc
PubMed: 23504314
DOI: 10.1074/jbc.M112.427229 -
The Journal of Biological Chemistry Nov 1980Horse neutrophils incorporate exogenous [14C]arachidonate into phosphatidic acid very rapidly. This acylation of phosphatidate with arachidonate is followed quickly and...
Horse neutrophils incorporate exogenous [14C]arachidonate into phosphatidic acid very rapidly. This acylation of phosphatidate with arachidonate is followed quickly and spontaneously by its deacylation. This transient formation of arachidonyl-phosphatidate, which reflects a rapidly turning over pool of arachidonate-associated lipid, is not observed with stearic acid or other phospholipids or triglycerides. Phosphatidylcholine, phosphatidylethanolamine, phosphatidylinosinositol, and triglycerides are slowly but increasingly labeled with time. Ionophore A23187 (10 microM) stimulates the extent of labeling of phosphatidate while decreasing the labeling of all the other phospholipids and triglycerides. Phosphatidate is not transiently labeled with [14C]stearate of (32P)orthophosphate, either in the presence or absence of ionophore A23187. When cells are prelabeled for 2 h with very high quantities of (32P)orthophosphate a very substantial fraction (i.e. 20 to 30%) of the phospholipid radioactivity is associated with phosphatidic acid. However, on addition of exogenous arachidonate, there is no increase in [32P]phosphatidate in these prelabeled cells. Thus, the entire phosphatidate molecule does not appear to be turned over during the process described above. Inhibitors of cyclooxygenase and lipoxygenase activities such as BW755C, nordihydroguaiaretic acid, and low concentrations of indomethacin do not affect the labeling of phospholipids. However, eicosatetraynoic acid, an analog of arachidonate, and high concentration (0.1 mM) of indomethacin can block [14C]arachidonate incorporation into lipids. The rapid turnover of the 2-acyl position in phosphatidate might be related to a specific process of fatty acid mobilization within neutrophils.
Topics: Acylation; Animals; Arachidonic Acids; Calcimycin; Horses; Indomethacin; Kinetics; Neutrophils; Phosphatidic Acids; Phospholipids
PubMed: 6776123
DOI: No ID Found -
FEBS Letters Jun 2020Serotonin transporter (SERT) is involved in serotonergic system regulation and in the pathophysiology/therapeutics of serotonin-/SERT-related diseases such as...
Serotonin transporter (SERT) is involved in serotonergic system regulation and in the pathophysiology/therapeutics of serotonin-/SERT-related diseases such as obsessive-compulsive disorder, depression, autism, and schizophrenia. We recently revealed that diacylglycerol (DG) kinase (DGK) δ induces ubiquitination/degradation of SERT in a DGK activity-dependent manner through Praja-1 E3 ubiquitin-protein ligase. However, it is still unclear how Praja-1 activity is regulated by DGKδ. Here, we reveal that 1-stearoyl-2-docosahexaenoyl (18:0/22:6)-phosphatidic acid (PA) and 18:0/22:6-DG are simultaneously decreased and accumulated, respectively, in the DGKδ-knockout mouse brain, indicating that DGKδ selectively phosphorylates 18:0/22:6-DG to generate 18:0/22:6-PA. Moreover, we find that 18:0/22:6-PA selectively binds to Praja-1 and enhances its activity. These results strongly suggest that 18:0/22:6-PA generated by DGKδ activates Praja-1 to degrade SERT in the brain.
Topics: Animals; Brain; COS Cells; Chlorocebus aethiops; Diacylglycerol Kinase; Enzyme Activation; Male; Mice; Phosphatidic Acids; Serotonin Plasma Membrane Transport Proteins; Substrate Specificity; Ubiquitin-Protein Ligases
PubMed: 32134507
DOI: 10.1002/1873-3468.13765 -
Plant, Cell & Environment Apr 2010The activity of phospholipase D (PLD) in plants increases under different hyperosmotic stresses, such as dehydration, drought, and salinity. Recent results begin to shed...
The activity of phospholipase D (PLD) in plants increases under different hyperosmotic stresses, such as dehydration, drought, and salinity. Recent results begin to shed light onto the involvement of PLD in response to water deficits and salinity. Different PLDs have unique and overlapping functions in these responses. PLDalpha1 promotes stomatal closure and reduces water loss. PLDalpha1 and PLDdelta are involved in seedling tolerance to salt stress. PLDalpha3 and PLDepsilon enhance plant growth and hyperosmotic tolerance. The different PLDs regulate the production of phosphatidic acid (PA) that is a key class of lipid mediators in plant response to environmental stresses. Further studies on the upstream regulators that activate different PLDs and the downstream effectors of PLDs and PA have the potential to unveil the linkage between the stimulus perception at the cell membrane to intracellular responses to drought and salinity stresses.
Topics: Droughts; Gene Expression Regulation, Plant; Phosphatidic Acids; Phospholipase D; Plant Development; Plant Stomata; Plants; Salinity; Signal Transduction; Stress, Physiological; Water
PubMed: 19968827
DOI: 10.1111/j.1365-3040.2009.02087.x -
The Journal of Biological Chemistry Dec 2003We have identified a novel phospholipase A1, named mPA-PLA1beta, which is specifically expressed in human testis and characterized it biochemically together with...
We have identified a novel phospholipase A1, named mPA-PLA1beta, which is specifically expressed in human testis and characterized it biochemically together with previously identified mPA-PLA1alpha. The sequence of mPAPLA1beta encodes a 460-amino acid protein containing a lipase domain with significant homology to the previously identified phosphatidic acid (PA)-selective PLA1, mPA-PLA1alpha. mPA-PLA1beta contains a short lid and deleted beta9 loop, which are characteristics of PLA1 molecules in the lipase family, and is a member of a subfamily in the lipase family that includes mPA-PLA1alpha and phosphatidylserine-specific PLA1. Both mPA-PLA1beta and mPA-PLA1alpha recombinant proteins exhibited PA-specific PLA1 activity and were vanadate-sensitive. When mPAPLA1beta-expressing cells were treated with bacterial phospholipase D, the cells produced lysophosphatidic acid (LPA). In both mPA-PLA1alpha and beta-expressing cells, most of the PA generated by the phospholipase D (PLD) treatment was converted to LPA, whereas in control cells it was converted to diacylglycerol. When expressed in HeLa cells most mPA-PLA1alpha protein was recovered from the cell supernatant. By contrast, mPA-PLA1beta was recovered almost exclusively from cells. Consistent with this observation, we found that mPA-PLA1beta has higher affinity to heparin than mPA-PLA1alpha. We also found that the membrane-associated mPA-PLA1s were insoluble in solubilization by 1% Triton X-100 and were detected in Triton X-100-insoluble buoyant fractions of sucrose gradients. The present study raises the possibility that production of LPA by mPA-PLA1alpha and -beta occurs on detergent-resistant membrane domains of the cells where they compete with lipid phosphate phosphatase for PA.
Topics: Amino Acid Sequence; Animals; Base Sequence; Blotting, Western; Cell Line; Diglycerides; Fluorescent Antibody Technique; Humans; Mass Spectrometry; Molecular Sequence Data; Phosphatidic Acids; Phospholipases A; Phospholipases A1; Recombinant Proteins; Sequence Homology, Amino Acid; Spodoptera; Substrate Specificity; Vanadates
PubMed: 12963729
DOI: 10.1074/jbc.M213018200 -
The International Journal of... Aug 2012Mitochondria, once viewed as functioning relatively autonomously in the cell, have increasingly been recognized to be involved in numerous signaling networks that impact... (Review)
Review
Mitochondria, once viewed as functioning relatively autonomously in the cell, have increasingly been recognized to be involved in numerous signaling networks that impact on a wide range of cell biological processes. In addition to the many types of proteins that mediate these pathways, the importance of signaling functions regulated via lipids and lipid second messengers generated on the mitochondrial surface is also becoming well appreciated. We focus here on phosphatidic acid, a lipid second messenger produced via several different pathways that can in turn stimulate the formation of multiple other bioactive lipids. Taken together, fascinating roles for phosphatidic acid and the connected lipids in mitochondrial function and interaction with other organelles are being uncovered. These pathways present new opportunities for the development of therapeutic approaches relevant to reproduction, metabolism, and neurodegenerative disease.
Topics: Cell Physiological Phenomena; Humans; Lipid Metabolism; Mitochondria; Neurodegenerative Diseases; Organelles; Phosphatidic Acids; Signal Transduction
PubMed: 22609101
DOI: 10.1016/j.biocel.2012.05.006