Abscisic Acid Stimulation of Phospholipase D in the Barley Aleurone Is G-Protein-Mediated and Localized to the Plasma Membrane

Phosphatidylethanol is formed by ‘transphosphatidylation’ of phospholipids with ethanol catalysed by phospholipase D and can be accumulated in the plasma membrane of mammalian cells after treatment of animals with ethanol. In the present work we show that phosphatidylalcohols, such as phosphatidylethanol and phosphatidylbutanol, produced a twofold stimulation of the Ca2+-ATPase activity of human erythrocytes. This stimulation occurs with the purified, solubilized enzyme as well as with ghost preparations, where the enzyme is in its natural lipidic environment and is different to that obtained with other acidic phospholipids such as phosphatidylserine. Addition of either phosphatidylserine, phosphatidylethanol or phosphatidylbutanol to the purified Ca2+-ATPase, or to ghosts preparations, increased the affinity of the enzyme for Ca2+ and the maximal velocity of the reaction as compared with controls in the absence of acidic phospholipids. However, in contrast with what occurs with phosphatidylserine, simultaneous addition of phosphatidylalcohols and calmodulin increased the affinity of the enzyme for Ca2+ to a greater extent than each added separately. When ethanol was added to either the purified erythrocyte Ca2+-ATPase or to erythrocyte-ghost preparations in the presence of acidic phospholipids, an additive effect was observed. There was an increase in the affinity for Ca2+ and in the maximal velocity of the reaction, well above the values obtained with ethanol or with the acidic phospholipids tested separately. These findings could have pharmacological importance. It is conceivable that the decrease in the intracellular Ca2+ concentration that has been reported in erythrocytes as a result of ethanol intoxication could be due to the stimulation of the Ca2+-ATPase by the accumulated phosphatidylethanol, to a direct effect of ethanol on the enzyme or to an additive combination of both.

Download Full-text

Stimulation of calcium influx by platelet activating factor in Dictyostelium

Journal of Cell Science ◽

10.1242/jcs.108.4.1597 ◽

1995 ◽

Vol 108 (4) ◽

pp. 1597-1603

Author(s):

R. Schaloske ◽

C. Sordano ◽

S. Bozzaro ◽

D. Malchow

Keyword(s):

Plasma Membrane ◽

G Protein ◽

Dictyostelium Discoideum ◽

Calcium Influx ◽

Platelet Activating Factor ◽

Inactive Compound ◽

Time Period ◽

Dependent Pathway ◽

Stimulation Of

Platelet activating factor (PAF) induces Ca2+ influx in Dictyostelium discoideum. In this investigation we used this activity to analyze the mechanism of PAF action. We found that PAF activity was confined to the period of spike-shaped oscillations and suggest that the role of PAF is to augment cAMP relay. PAF seems to act only a few times during this time period of two hours, since Ca2+ entry adapted to a subsequent stimulus for about 30 minutes. PAF showed a reduced response in the G protein beta- strain LW14 and was unable to induce Ca2+ influx in the G alpha 2- strains HC85 and JM1. The latter expresses the cAMP receptors cAR1 constitutively, and exhibits cAMP-induced Ca2+ influx, albeit at a reduced level. In order to decide whether the inability of PAF to elicit a Ca2+ response in JM1 cells was due to the lack of differentiation and/or the lack of G alpha 2, we inhibited the IP3-dependent pathway with compound U73122 and found that Ca2+ entry was blocked, whereas a closely related inactive compound, U73343, did not alter the response. In agreement with this, NBD-Cl, an inhibitor of Ca2+ uptake into the IP3-sensitive store in Dictyostelium, also abolished PAF activity. The latter was not inhibited by the plasma membrane antagonists BN-52021 or WEB 2170. Therefore PAF seems to operate intracellularly via the IP3-signalling pathway at or upstream of the IP3-sensitive store.

Download Full-text

Phospholipase D is activated by G protein and not by calcium ions in vascular smooth muscle

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1996.270.3.h1031 ◽

1996 ◽

Vol 270 (3) ◽

pp. H1031-H1037

Author(s):

E. F. LaBelle ◽

R. M. Fulbright ◽

R. J. Barsotti ◽

H. Gu ◽

E. Polyak

Keyword(s):

Smooth Muscle ◽

Vascular Smooth Muscle ◽

G Protein ◽

Phospholipase D ◽

Pertussis Toxin ◽

Alpha Toxin ◽

Tail Artery ◽

Agonist Stimulation ◽

Rat Tail ◽

Stimulation Of

We assessed the sensitivity of phospholipase D (PLD) activity in vascular smooth muscle to cytosolic Ca2+ by increasing cytosolic Ca2+ levels independently of agonist stimulation. When rat tail artery was preloaded with the Ca2+ indicator fluo 3 pentaacetoxymethyl ester, the addition of high extracellular K+, caffeine, or norepinephrine rapidly enhanced cytosolic Ca2+ levels. Neither increased extracellular K+ nor caffeine addition increased phosphatidylethanol production, indicating that cytosolic Ca2+ elevation alone did not stimulate PLD. In contrast, norepinephrine stimulated phosphatidylethanol production in this tissue. In strips of tail artery permeabilized with alpha-toxin and incubated in solutions containing free Ca2+ concentrations observed during physiological stimulation (pCa 6.4), PLD was not stimulated, whereas incubation with guanosine 5'-O-(3-thiotriphosphate) at pCa 7.0 activated this enzyme. Aluminum fluoride (AlF4-) stimulated PLD, and this activity was insensitive to pertussis toxin after stimulation by either norepinephrine or AlF4-. These results indicate that PLD in vascular smooth muscle is activated by norepinephrine via stimulation of a pertussis toxin-insensitive G protein and not via an increase in intracellular Ca2+ levels.

Download Full-text

Abscisic acid signal transduction in the barley aleurone is mediated by phospholipase D activity

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.95.5.2697 ◽

1998 ◽

Vol 95 (5) ◽

pp. 2697-2702 ◽

Cited By ~ 115

Author(s):

S. Ritchie ◽

S. Gilroy

Keyword(s):

Signal Transduction ◽

Abscisic Acid ◽

Phospholipase D ◽

Barley Aleurone

Download Full-text

ADP-ribosylation-factor-regulated phospholipase D activity localizes to secretory vesicles and mobilizes to the plasma membrane following N-formylmethionyl-leucyl-phenylalanine stimulation of human neutrophils

Biochemical Journal ◽

10.1042/bj3250581 ◽

1997 ◽

Vol 325 (3) ◽

pp. 581-585 ◽

Cited By ~ 62

Author(s):

C. P. MORGAN ◽

H. SENGELOV ◽

J. WHATMORE ◽

N. BORREGAARD ◽

S. COCKCROFT

Keyword(s):

Plasma Membrane ◽

Phospholipase D ◽

Small Gtpases ◽

Human Neutrophils ◽

Secretory Vesicles ◽

Rho Proteins ◽

Adp Ribosylation ◽

Activated Neutrophils ◽

Adp Ribosylation Factor ◽

Stimulation Of

Phospholipase D (PLD) is responsible for the hydrolysis of phosphatidylcholine to produce phosphatidic acid and choline. Human neutrophils contain PLD activity which is regulated by the small GTPases, ADP-ribosylation factor (ARF) and Rho proteins. In this study we have examined the subcellular localization of the ARF-regulated PLD activity in non-activated neutrophils and cells ‘primed‘ with N-formylmethionyl-leucyl-phenylalanine (fMetLeuPhe). We report that PLD activity is localized at the secretory vesicles in control cells and is mobilized to the plasma membrane upon stimulation with fMetLeuPhe. We conclude that the ARF-regulated PLD activity is translocated to the plasma membrane by secretory vesicles upon stimulation of neutrophils with fMetLeuPhe in inflammatory/priming doses. We propose that this relocalization of PLD is important for the subsequent events occurring during neutrophil activation.

Download Full-text

G protein-regulated endocytic trafficking of adenylyl cyclase type 9

10.1101/2020.04.23.057026 ◽

2020 ◽

Author(s):

André M. Lazar ◽

Roshanak Irannejad ◽

Tanya A. Baldwin ◽

Aparna A. Sundaram ◽

J. Silvio Gutkind ◽

...

Keyword(s):

Plasma Membrane ◽

Adenylyl Cyclase ◽

G Protein ◽

Subcellular Location ◽

Endocytic Pathway ◽

Heterotrimeric G Proteins ◽

Camp Production ◽

Stimulation Of

SummaryGPCRs are increasingly recognized to initiate signaling via heterotrimeric G proteins as they move through the endocytic network, but little is known about how relevant G protein effectors are localized. Here we report dynamic trafficking of adenylyl cyclase type 9 (AC9) from the plasma membrane to endosomes, while adenylyl cyclase type 1 (AC1) remains in the plasma membrane, and stimulation of AC9 trafficking by ligand-induced activation of Gs-coupled GPCRs or Gs. AC9 transits a similar dynamin-dependent early endocytic pathway as activated GPCRs but, in contrast to GPCR trafficking which is regulated by β-arrestin but not Gs, AC9 trafficking is regulated by Gs but not β-arrestin. We also show that AC9, but not AC1, contributes to cAMP production from endosomes. These results reveal dynamic and isoform-specific trafficking of adenylyl cyclase in the endocytic network, and a discrete role of a heterotrimeric G protein in controlling subcellular location of a relevant effector.

Download Full-text