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ABSTRACT: Lipid phosphate phosphatases (LPPs, E.C. 3.1.3.4) catalyse the dephosphorylation of diacylglycerol pyrophosphate (DGPP) and phosphatidic acid (PA), which are secondary messengers in abscisic acid (ABA) signalling. In this study, we investigated the effect of ABA on the expression of AtLPP genes as they encode putative ABA-signalling partners. We observed that AtLPP2 expression was down-regulated by ABA and we performed experiments on Atlpp2-2, an AtLPP2 knockout mutant, to determine whether AtLPP2 was involved in ABA signalling. We observed that Atlpp2-2 plantlets contained about twice as much PA as the wild-type Col-0 and exhibited higher PA kinase (PAK) activity than Col-0 plants. In addition, we showed that ABA stimulated diacylglycerol kinase (DGK) activity independently of AtLPP2 activity but that the ABA-stimulation of PAK activity recorded in Col-0 was dependent on AtLPP2. In order to evaluate the involvement of AtLPP2 activity in guard cell function, we measured the ABA sensitivity of Atlpp2-2 stomata. The inhibition of stomatal opening was less sensitive to ABA in Atlpp2-2 than in Col-0. Watered and water-stressed plants of the two genotypes accumulated ABA to the same extent, thus leading us to consider Atlpp2-2 an ABA-signalling mutant. Taken together our observations show that AtLPP2 is a part of ABA signalling and participate to the regulation of stomatal movements.
Plant Physiology and Biochemistry 03/2011; 49(3):357-62. · 2.84 Impact Factor
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ABSTRACT: Diacylglycerol pyrophosphate (DGPP) is a phosphorylated form of phosphatidic acid (PA) found in plants and yeast but not in
mammals. DGPP is a minor lipid that accumulates transiently under various abiotic stresses and during biotic interactions.
DGPP formation may be a way of attenuating PA content, but DGPP itself might also be a signaling lipid. DGPP is the product
of the phosphorylation of PA catalyzed by PA kinase (PAK). Unfortunately, studies describing the role of PAK are limited as
gene encoding PAK has not been identified yet. DGPP is dephosphorylated by lipid phosphate phosphatase (LPP) activity to produce
PA. LPPs with DGPP phosphatase activity are found in a wide variety of organisms including bacteria, yeast, plants, and mammals.
In Arabidopsis, four genes encoding LPPs have been identified, and a role for DGPP in abscisic acid signaling is becoming
apparent.
03/2009: pages 263-276;
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ABSTRACT: Protein tyrosine (Tyr) phosphorylation plays a central role in many signaling pathways leading to cell growth and differentiation in animals. Tyr phosphorylated proteins have been detected in higher plants, and the roles of protein Tyr phosphatases and protein Tyr kinases in some physiological responses have been shown. We investigated the involvement of Tyr phosphorylation events in abscisic acid (ABA) signaling using a pharmacological approach. Phenylarsine oxide, a specific inhibitor of protein Tyr phosphatase activity, abolished the ABA-dependent accumulation of RAB18 (responsive to ABA 18) transcripts. Protein Tyr kinase inhibitors like genistein, tyrphostin A23, and erbstatin blocked the RAB18 expression induced by ABA in Arabidopsis (Arabidopsis thaliana). Stomatal closure induced by ABA was also inhibited by phenylarsine oxide and genistein. We studied the changes in the Tyr phosphorylation levels of proteins in Arabidopsis seeds after ABA treatment. Proteins were separated by two-dimensional gel electrophoresis, and those phosphorylated on Tyr residues were detected using an anti-phosphotyrosine antibody by western blot. Changes were detected in the Tyr phosphorylation levels of 19 proteins after ABA treatment. Genistein inhibited the ABA-dependent Tyr phosphorylation of proteins. The 19 proteins were analyzed by matrix-assisted laser-desorption ionization time-of-flight/time-of-flight mass spectrometry. Among the proteins identified were storage proteins like cruciferins, enzymes involved in the mobilization of lipid reserves like aconitase, enolase, aldolase, and a lipoprotein, and enzymes necessary for seedling development like the large subunit of Rubisco. Additionally, the identification of three putative signaling proteins, a peptidyl-prolyl isomerase, an RNA-binding protein, and a small ubiquitin-like modifier-conjugating enzyme, enlightens how Tyr phosphorylation might regulate ABA transduction pathways in plants.
Plant physiology 10/2008; 148(3):1668-80. · 6.53 Impact Factor
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ABSTRACT: In Arabidopsis thaliana suspension cells, ABA was previously shown to promote the activation of anion channels and the reduction of proton pumping that both contribute to the plasma membrane depolarization. These two ABA responses were shown to induce two successive [Ca(2+)](cyt) spikes. As reactive oxygen species (ROS) have emerged as components of ABA signaling pathways especially by promoting [Ca(2+)](cyt) variations, we studied whether ROS were involved in the regulation of anion channels and proton pumps activities. Here we demonstrated that ABA induced ROS production which triggered the second of the two [Ca(2+)](cyt) increases observed in response to ABA. Blocking ROS generation using diphenyleneiodonium (DPI) impaired the proton pumping reduction, the anion channel activation and the RD29A gene expression in response to ABA. Furthermore, H(2)O(2) was shown to activate anion channels and to inhibit plasma membrane proton pumping, as did ABA. However, ROS partially mimicked ABA's effects since H(2)O(2) treatment elicited anion channel activation but not the subsequent expression of the RD29A gene as did ABA. This suggests that expression of the RD29A gene in response to ABA results from the activation of multiple concomitant signaling pathways: blocking of one of them would impair gene expression whereas stimulating only one would not. We conclude that ROS are a central messenger of ABA in the signaling pathways leading to the plasma membrane depolarization induced by ABA.
Plant and Cell Physiology 09/2008; 49(10):1495-507. · 4.70 Impact Factor
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ABSTRACT: The plant hormone abscisic acid (ABA) controls numerous physiological traits: dormancy and germination of seeds, senescence and resistance to abiotic stresses. In order to get more insight into the role of protein tyrosine phosphatase (PTP) in ABA signalling, we obtained eight homozygous T-DNA insertion lines in Arabidopsis thaliana PTP genes. One mutant, named phs1-3, exhibited a strong ABA-induced inhibition of germination as only 26% of its seeds germinated after 3 days instead of 92% for the Columbia (Col-0) line. Genetic and molecular analyses of phs1-3 showed that it bears a unique T-DNA insertion in the promoter of the gene and that the mutation is recessive. PHS1 expression in the mutant is about half that of the Col-0 line. The upregulation of two ABA-induced genes (At5g06760, RAB18) and the downregulation of two ABA-repressed genes (AtCLC-A, ACL) are enhanced in the phs1-3 mutant compared with the wild-type. The 'in planta' aperture of phs1-3 stomata is reduced and the inhibition of the light-induced opening of stomata by ABA is stronger in phs1-3 leaves than in Col-0 leaves. Finally, PHS1 expression is upregulated in the presence of ABA in both phs1-3 and Col-0 but more intensively in the mutant. Thus, phs1-3 is hypersensitive to ABA. Taken together, these results show that PHS1, which encodes a dual-specificity PTP, is a negative regulator of ABA signalling.
The Plant Journal 10/2006; 47(5):711-9. · 6.16 Impact Factor
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ABSTRACT: Diacylglycerol pyrophosphate (DGPP) was recently shown to be a possible intermediate in abscisic acid (ABA) signaling. In this study, reverse transcription-PCR of ABA up-regulated genes was used to evaluate the ability of DGPP to trigger gene expression in Arabidopsis (Arabidopsis thaliana) suspension cells. At5g06760, LTI30, RD29A, and RAB18 were stimulated by ABA and also specifically expressed in DGPP-treated cells. Use of the Ca2+ channel blockers fluspirilene and pimozide and the Ca2+ chelator EGTA showed that Ca2+ was required for ABA induction of DGPP formation. In addition, Ca2+ participated in DGPP induction of gene expression via stimulation of anion currents. Hence, a sequence of Ca2+, DGPP, and anion currents, constituting a core of early ABA-signaling events necessary for gene expression, is proposed.
Plant physiology 09/2006; 141(4):1555-62. · 6.53 Impact Factor
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ABSTRACT: The sink demand was increased on a source maize leaf (Zea mays L. cv. F7F2) by darkening all the leaves except the fourth, which was maintained under the prevailing irradiance conditions. The parameters of carbon metabolism were measured precisely during the first hours, and then daily during one week. The ambient photosynthetic activity and the maximum photosynthetic capacity were not altered by the treatment but the soluble carbohydrate and starch contents diminished, while ADP-glucose pyrophosphorylase (EC 2.7.7.27) activity increased. The carbon export rate, evaluated by the rate of disappearance of radioactivity after a 1-min 14CO2 pulse, was faster than in control leaves. A compartmental analysis of the time course of 14C export further indicated that the sucrose pool providing the export flux was largely increased by the dark treatment. The darkened leaf 5, taken as an example of the darkened sources, was completely depleted of its carbohydrate content after one day in the dark and remained devoid of carbohydrates during the following week.
Physiologia Plantarum 04/2006; 94(2):319 - 327. · 3.11 Impact Factor
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Christine Zalejski,
Zongshen Zhang,
Anne-Laure Quettier,
Régis Maldiney,
Magda Bonnet,
Mathias Brault,
Chantal Demandre,
Emile Miginiac,
Jean-Pierre Rona,
Bruno Sotta, Emmanuelle Jeannette
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ABSTRACT: In plants, the importance of phospholipid signaling in responses to environmental stresses is becoming well documented. The involvement of phospholipids in abscisic acid (ABA) responses is also established. In a previous study, we demonstrated that the stimulation of phospholipase D (PLD) activity and plasma membrane anion currents by ABA were both required for RAB18 expression in Arabidopsis thaliana suspension cells. In this study, we show that the total lipids extracted from ABA-treated cells mimic ABA in activating plasmalemma anion currents and induction of RAB18 expression. Moreover, ABA evokes within 5 min a transient 1.7-fold increase in phosphatidic acid (PA) followed by a sevenfold increase in diacylglycerol pyrophosphate (DGPP) at 20 min. PA activated plasmalemma anion currents but was incapable of triggering RAB18 expression. By contrast, DGPP mimicked ABA on anion currents and was also able to stimulate RAB18 expression. Here we show the role of DGPP as phospholipid second messenger in ABA signaling.
The Plant Journal 05/2005; 42(2):145-52. · 6.16 Impact Factor
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ABSTRACT: Abscisic acid (ABA) plays a key role in the control of stomatal aperture by regulating ion channel activities and water exchanges across the plasma membrane of guard cells. Changes in cytoplasmic calcium content and activation of anion and outward-rectifying K(+) channels are among the earliest cellular responses to ABA in guard cells. In Arabidopsis suspension cells, we have demonstrated that outer plasmalemma perception of ABA triggered similar early events. Furthermore, a Ca(2+) influx and the activation of anion channels are part of the ABA-signaling pathway leading to the specific expression of RAB18. Here, we determine whether phospholipases are involved in ABA-induced RAB18 expression. Phospholipase C is not implicated in this ABA pathway. Using a transphosphatidylation reaction, we show that ABA plasmalemma perception results in a transient stimulation of phospholipase D (PLD) activity, which is necessary for RAB18 expression. Further experiments showed that PLD activation was unlikely to be regulated by heterotrimeric G proteins. We also observed that ABA-dependent stimulation of PLD was necessary for the activation of plasma anion current. However, when ABA activation of plasma anion channels was inhibited, the ABA-dependent activation of PLD was unchanged. Thus, we conclude that in Arabidopsis suspension cells, ABA stimulation of PLD acts upstream from anion channels in the transduction pathway leading to RAB18 expression.
Plant physiology 10/2002; 130(1):265-72. · 6.53 Impact Factor
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ABSTRACT: Based on known amino acid sequences, probes have been generated by PCR and used for the subsequent isolation of cDNAs and genes coding for two thioredoxins (m and h) of Chlamydomonas reinhardtii. Thioredoxin m, a chloroplastic protein, is encoded as a preprotein of 140 amino acids (15 101 Da) containing a transit peptide of 34 amino acids with a very high content of Ala and Arg residues. The sequence for thioredoxin h codes for a 113 amino acid protein with a molecular mass of 11817 Da and no signal sequence. The thioredoxin m gene contains a single intron and seems to be more archaic in structure than the thioredoxin h gene, which is split into 4 exons. The cDNA sequences encoding C. reinhardtii thioredoxins m and h have been integrated into the pET-3d expression vector, which permits efficient production of proteins in Escherichia coli cells. A high expression level of recombinant thioredoxins was obtained (up to 50 mg/l culture). This has allowed us to study the biochemical/biophysical properties of the two recombinant proteins. Interestingly, while the m-type thioredoxin was found to have characteristics very close to the ones of prokaryotic thioredoxins, the h-type thioredoxin was quite different with respect to its kinetic behaviour and, most strikingly, its heat denaturation properties.
Plant Molecular Biology 05/1995; 28(3):487-503. · 4.15 Impact Factor
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ABSTRACT: ADPglucose pyrophosphorylase activity (measured in the direction of glucose-IP synthesis) from adult maize leaves was shown to present diurnal variations in activity which were not related to the starch accumulation rate. Extractable ADPG-PPase activity was maximum at the end of the dark period and for the first 8 hours in the light. Subsequently, ADPG-PPase activity progressively declined to half the initial value, by the end of the day. A short 1 hour-dark period during the day may transiently reverse the inhibition. Experiments with excised leaves placed on sucrose or sorbitol solution further showed that the magnitude of ADPG-PPase inhibition was related to high leaf sucrose and starch contents. The differences in activity were not due to changes in the quantity of ADPG-PPase protein as shown by immunodot and immunoprecipitation techniques nor to the differential response of the mesophyll and bundle sheath isoforms nor to a time-dependent susceptibility of the enzyme to proteolysis. Thus, changes in ADPG-PPase specific activity were inferred. A search for in vivo phosphorylation of the protein was proved negative, whatever the light or dark pre-treatment which modified the ADPG-PPase activity. The difference between the 7:00 (morning) and 21:00 (evening) forms was also observed when activity was measured by formation of ADPglucose (the physiological direction) in presence or absence of PGA, as activator. The two forms retained different activity upon gel filtration.
