Cloning, expression and functional characterization of the C2 domain from tomato phospholipase Dα

Tiwari, Krishnaraj; Paliyath, Gopinadhan

doi:10.1016/j.plaphy.2010.09.015

Cited by 32 publications

(16 citation statements)

References 51 publications

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“…Interestingly, C2 domains from other proteins have been shown to bind PA, e.g. tomato PLDα (phospholipase Dα) [42], and mammalian PKCε (protein kinase Cε), which has been shown to specifically respond to PA accumulation in vivo [43]. Whereas HsPTEN is known to bind PS [44], the binding of PA has not been reported for animal PTEN proteins.…”

Section: Discussionmentioning

confidence: 99%

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

Pribat

Sormani

Rousseau‐Gueutin

et al. 2011

Biochemical Journal

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

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Section: Discussionmentioning

confidence: 99%

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

Pribat

Sormani

Rousseau‐Gueutin

et al. 2011

Biochemical Journal

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“…Thus, it has been hypothesized that this dissimilar affinity for Ca 2+ came from the lack of one acidic residue within the CBR1 of the C2 domain of the AtPLD α , due to the substitution of D by Q residue, which could explain the millimolar concentration range of Ca 2+ required for optimal activity of AtPLD α in vitro [35]. However, a recent study on the C2 domain from tomato PLD α tends to disclaim these properties, demonstrating the existence of two high-affinity binding sites for Ca 2+ -binding sites (Kd = 59.73 μ M) [36]. …”

Section: Introductionmentioning

confidence: 99%

Functional Characterization of the N-Terminal C2 Domain fromArabidopsis thalianaPhospholipase Dαand Dβ

Rahier

Noiriel

Abousalham

2016

BioMed Research International

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Most of plant phospholipases D (PLD) exhibit a C2-lipid binding domain of around 130 amino acid residues at their N-terminal region, involved in their Ca2+-dependent membrane binding. In this study, we expressed and partially purified catalytically active PLDα from Arabidopsis thaliana (AtPLDα) in the yeast Pichia pastoris. The N-terminal amino acid sequence of the recombinant AtPLDα was found to be NVEETIGV and thus to lack the first 35 amino acid belonging to the C2 domain, as found in other recombinant or plant purified PLDs. To investigate the impact of such a cleavage on the functionality of C2 domains, we expressed, in E. coli, purified, and refolded the mature-like form of the C2 domain of the AtPLDα along with its equivalent C2 domain of the AtPLDβ, for the sake of comparison. Using Förster Resonance Energy Transfer and dot-blot assays, both C2 domains were shown to bind phosphatidylglycerol in a Ca2+-independent manner while phosphatidic acid and phosphatidylserine binding were found to be enhanced in the presence of Ca2+. Amino acid sequence alignment and molecular modeling of both C2 domains with known C2 domain structures revealed the presence of a novel Ca2+-binding site within the C2 domain of AtPLDα.

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“…Binding sites of PtdIns(4,5)P 2 in domain C2 are also found in PLDα of cabbage [44], poppy [45], and tomatoes [46]. The C2 domain in PLDα of tomatoes binds with high affinity phosphatidic acid but not phosphatidylcholine, but association with phosphoinositides is markedly weakened on increase in the level of their phosphorylation [46]. Domain PX.…”

Section: Specific Features Of Primary Structure Of Pldmentioning

confidence: 97%

“…At low level of Ca 2+ negatively charged PtdInsP and PtdIns(4,5)P 2 molecules attract positively charged amino acid residues R and K in the C2 domain and thus favor the binding with substrates of PLDα of Arabidopsis and catalysis by PLDβ and PLDγ [2,43]. Binding sites of PtdIns(4,5)P 2 in domain C2 are also found in PLDα of cabbage [44], poppy [45], and tomatoes [46]. The C2 domain in PLDα of tomatoes binds with high affinity phosphatidic acid but not phosphatidylcholine, but association with phosphoinositides is markedly weakened on increase in the level of their phosphorylation [46].…”

Section: Specific Features Of Primary Structure Of Pldmentioning

confidence: 99%

“…The presence of Nterminal conservative domain C2 consisting of 130 a.a. and binding Ca 2+ and phospholipids is a unique feature of the majority of plant PLDs [2]. The binding of Ca 2+ with the domain C2 provides for the PLD translocation to substrate on the membrane and induces changes in the enzyme conformation [46]. A direct binding of Ca 2+ in three-looped structures of the C2 domain [11] has been shown for PLDα and PLDβ of Arabidopsis [2].…”

Section: Specific Features Of Primary Structure Of Pldmentioning

confidence: 99%

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Molecular structure of phospholipase D and regulatory mechanisms of its activity in plant and animal cells

Kolesnikov

Nokhrina

Kretynin

et al. 2012

Biochemistry Moscow

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Abbreviations: Gα, α-subunit of heterotrimeric G-protein; PKC, protein kinase C; PLD, phospholipase D; PtdIns(4,5)P 2 , phosphatidylinositol-4,5-bisphosphate. * To whom correspondence should be addressed.Abstract⎯Phospholipase D (PLD) catalyzes hydrolysis of phospholipids with production of phosphatidic acids, which often act as secondary messengers on transmission of intracellular signals. This review summarizes data of various leading laboratories on specific features of organization and regulation of PLD activity in plant and animal cells. The main structural domains of PLD (C2, PX, PH), the active site, and other functionally important parts of the enzyme are considered. Regulatory mechanisms of PLD activity are characterized in detail. Studies associated with molecular design, analysis, and synthesis of new nontoxic substances capable of inhibiting different PLD isoenzymes in vivo are shown to be promising for biotechnology and medicine.

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Cloning, expression and functional characterization of the C2 domain from tomato phospholipase Dα

Cited by 32 publications

References 51 publications

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

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

Functional Characterization of the N-Terminal C2 Domain fromArabidopsis thalianaPhospholipase Dαand Dβ

Molecular structure of phospholipase D and regulatory mechanisms of its activity in plant and animal cells

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