Selectivity of the diacylglycerol kinase inhibitor 3-{2-(4-[bis-(4-fluorophenyl)methylene]-1-piperidinyl)ethyl}-2,3-dihydro-2-thioxo-4(1H)quinazolinone (R59949) among diacylglycerol kinase subtypes

Jiang, Ying; Sakane, Fumio; Kanoh, Hideo; Walsh, James P.

doi:10.1016/s0006-2952(99)00395-0

Cited by 118 publications

(143 citation statements)

References 39 publications

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“…To further confirm that the formation of PA was a result of AtDGK2 activity, the reaction mix was preincubated with the DGK inhibitor R59022, which has been shown to be active in both animal and plant cells (18,37,40,41). In our experiments, the decrease in PA formation was dependent on the concentration of the inhibitor (Fig.…”

Section: Fig 2 Phylogenetic Analysis Of Plant Dgk Isozymessupporting

confidence: 53%

AtDGK2, a Novel Diacylglycerol Kinase from Arabidopsis thaliana, Phosphorylates 1-Stearoyl-2-arachidonoyl-sn-glycerol and 1,2-Dioleoyl-sn-glycerol and Exhibits Cold-inducible Gene Expression

Gómez-Meriño

Brearley

Ornatowska

et al. 2004

Journal of Biological Chemistry

117

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Section: Fig 2 Phylogenetic Analysis Of Plant Dgk Isozymessupporting

confidence: 53%

AtDGK2, a Novel Diacylglycerol Kinase from Arabidopsis thaliana, Phosphorylates 1-Stearoyl-2-arachidonoyl-sn-glycerol and 1,2-Dioleoyl-sn-glycerol and Exhibits Cold-inducible Gene Expression

Gómez-Meriño

Brearley

Ornatowska

et al. 2004

Journal of Biological Chemistry

117

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“…In PAE-KDR cells, by assaying Dgk activity in anti-Dgk-a immunoprecipitates, we provided compelling evidence that Dgk-a isoform is activated by VEGF. In addition, most of the total Dgk activity assayed from PAE-KDR wholecell homogenates is inhibited in vitro by R59949, which inhibits selectively class-I Dgk enzymes, that is, the a-, band g-isoforms, while it does not affect the activity of other Dgk isoforms (Jiang et al 2000), as well as the activity of other lipid and protein kinases (De Chaffoy de Courcelles et al, 1989;Cutrupi et al, 2000). The finding that overexpression of wt Dgk-a reverts the ability of R59949 to impair HGF-induced cell movement and network organization on matrigel, respectively, in PAE and PAE-KDR cells provides further support to the selectivity of R59949 for Dgk-a (Cutrupi et al, 2000; Figure 4c).…”

Section: Discussionmentioning

confidence: 99%

“…In HUVEC cultured in 2% FCS, VEGF-A 165 stimulates DNA synthesis, which was inhibited by R59949 at concentrations equal or higher than 5 mM (Figure 5d). The reduced ability of R59949 to inhibit DNA synthesis in HUVEC may depend on serum sequestration of R59949 (De Chaffoy de Courcelles et al, 1989), although the involvement of other Dgk isoforms, less sensitive to the inhibitor (Jiang et al, 2000), could not be ruled out.…”

Section: Pharmacological Inhibition Of Dgk-a Impairs In Vitro Vegf-a mentioning

confidence: 99%

Activation of diacylglycerol kinase α is required for VEGF-induced angiogenic signaling in vitro

et al. 2004

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“…For this purpose, we used R59949, which inhibits calciumdependent DGK isoforms (type I: ␣, ␤, and ␥) (38). Indeed, 10 M R59949 significantly inhibited DGK␥ activity (ϳ70% inhibition) in vitro (data not shown).…”

Section: D)mentioning

confidence: 99%

Diacylglycerol Kinase γ Serves as an Upstream Suppressor of Rac1 and Lamellipodium Formation

Tsushima

Kai

Yamada

et al. 2004

Journal of Biological Chemistry

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Nine diacylglycerol kinase (DGK) isozymes have been identified. However, our knowledge of their individual functions is still limited. Here, we demonstrate the role of DGK␥ in regulating Rac1-governed cell morphology. We found that the expression of kinase-dead DGK␥, which acts as a dominant-negative mutant, and inhibition of endogenous DGK␥ activity with R59949 induced lamellipodium and membrane ruffle formation in NIH3T3 fibroblasts in the absence of growth factor stimulation. Reciprocally, lamellipodium formation induced by plateletderived growth factor was significantly inhibited upon expression of constitutively active DGK␥. Moreover, the constitutively active DGK␥ mutant suppressed integrinmediated cell spreading. These effects are isoform-specific because, in the same experiments, none of the corresponding mutants of DGK␣ and DGK␤, closely related isoforms, affected cell morphology. These results suggest that DGK␥ specifically participates in the Rac1-mediated signaling pathway leading to cytoskeletal reorganization. In support of this, DGK␥ co-localized with dominant-active Rac1 especially in lamellipodia. Moreover, we found that endogenous DGK␥ was physically associated with cellular Rac1. Dominant-negative Rac1 expression blocked the lamellipodium formation induced by kinase-dead DGK␥, indicating that DGK␥ acts upstream of Rac1. This model is supported by studies demonstrating that kinase-dead DGK␥ selectively activated Rac1, but not Cdc42. Taken together, these results strongly suggest that DGK␥ functions through its catalytic action as an upstream suppressor of Rac1 and, consequently, lamellipodium/ruffle formation.It is well recognized that a variety of lipid second messengers in low abundance carry out specific tasks for a wide range of biological processes in eukaryotic cells. The cellular concentrations of such signaling lipids must be strictly regulated by the action of metabolic enzymes. Diacylglycerol kinase (DGK) 1 phosphorylates diacylglycerol (DAG) to yield phosphatidic acid (PA) (1). DAG is an established activator of conventional and novel protein kinases C (PKCs), Unc-13, and Ras guanyl nucleotide-releasing protein (2-6). PA has also been reported to regulate a number of signaling proteins such as phosphatidylinositol-4-phosphate 5-kinase, Ras GTPase-activating protein, Raf-1 kinase, atypical PKC, and chimaerins (6 -8). Moreover, specific PA-binding sites have recently been identified in several important proteins such as Raf-1 (9), mTOR (mammalian target of rapamycin) (10), and p47 phox (11). Thus, DGK can potentially participate in a diverse range of cellular events through modulating the balance between two bioactive lipids, DAG and PA.Mammalian DGK is known to exist as a large protein family consisting of nine isozymes classified into five subtypes according to their structural features (12-15). These subfamilies can be characterized by the presence of a variety of regulatory domains of known and/or predicted functions, clearly indicating their distinct functions and regulatory mecha...

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Selectivity of the diacylglycerol kinase inhibitor 3-{2-(4-[bis-(4-fluorophenyl)methylene]-1-piperidinyl)ethyl}-2,3-dihydro-2-thioxo-4(1H)quinazolinone (R59949) among diacylglycerol kinase subtypes

Cited by 118 publications

References 39 publications

AtDGK2, a Novel Diacylglycerol Kinase from Arabidopsis thaliana, Phosphorylates 1-Stearoyl-2-arachidonoyl-sn-glycerol and 1,2-Dioleoyl-sn-glycerol and Exhibits Cold-inducible Gene Expression

AtDGK2, a Novel Diacylglycerol Kinase from Arabidopsis thaliana, Phosphorylates 1-Stearoyl-2-arachidonoyl-sn-glycerol and 1,2-Dioleoyl-sn-glycerol and Exhibits Cold-inducible Gene Expression

Activation of diacylglycerol kinase α is required for VEGF-induced angiogenic signaling in vitro

Diacylglycerol Kinase γ Serves as an Upstream Suppressor of Rac1 and Lamellipodium Formation

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