Selective Activation of Rolipram-Sensitive, cAMP-Specific Phosphodiesterase Isoforms by Phosphatidic Acid
- 1Unité 352 Institut National de la Santé et de la Recherche Médicale, Biochimie et Pharmacologie Institut National des Sciences Appliquées-Lyon, 69621 Villeurbanne, France (G.N.), and2Division of Reproductive Biology, Department of Gynecology and Obstetrics, Stanford University School of Medicine, Stanford, California 94305 (C.S., M.C.)
Abstract
In rat thymic lymphocytes, accumulation of phosphatidic acid (PA) occurs at the same time as decrease in cAMP levels and activation of a cAMP-specific phosphodiesterase (PDE) [type 4, EC 3.1.4.17 (PDE4)]. We investigated the nature of the PDE activated by PA and the mechanism of activation by using recombinant cAMP-specific PDE4 isoforms derived from three different genes (PDE4A, PDE4B, andPDE4D). The “long” variants expressed from each gene (PDE4A5, PDE4B1, and PDE4D3) were activated by PA, whereas the “short” variants (PDE4A1, PDE4B2, PDE4D1, and PDE4D2) were not. Phosphatidylserine was an activator that was as effective as PA, whereas phosphatidylcholine was ineffective, indicating that activation was restricted to anionic phospholipids. PA caused an increase in theVmax value of PDE4D3 without affecting theKm value of the enzyme for the cAMP substrate. PA also caused a change in the Mg2+requirement for hydrolysis. Half-maximal stimulation of the PDE was obtained with ∼10 μg/ml PA. Although protein kinase A-mediated phosphorylation of PDE4D3 produces effects similar to those elicited by PA, the mechanism of PA-induced activation was not found to involve a phosphorylation. Instead, several observations suggest that PA may directly interact with the enzyme. The stimulation of cAMP PDEs by PA and other acidic phospholipids may be a mechanism by which growth factors and hormones modulate the cAMP-dependent signal transduction pathway during cell stimulation.
Footnotes
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Send reprint requests to: Dr. G. Némoz, Laboratoire de Biochimie et Pharmacologie, Unité INSERM 352, Batiment 406, INSA, 69621 Villeurbanne Cedex, France. E-mail:nemoz{at}insa.insa-lyon.fr
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↵1 G. Némoz, unpublished observations.
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This work was supported by the Institut National de la Santé et de la Recherche Médicale and by National Institutes of Health Grant HD20788 (M.C.).
- Abbreviations:
- PA
- phosphatidic acid
- PDE
- phosphodiesterase
- LPA
- lysophosphatidic acid
- PC
- phosphatidylcholine
- PS
- phosphatidylserine
- DAG
- diacylglycerol
- PKA
- cAMP-dependent protein kinase
- PKI
- cAMP-dependent protein kinase synthetic inhibitor peptide
- SDS
- sodium dodecyl sulfate
- PAGE
- polyacrylamide gel electrophoresis
- PKC
- protein kinase C
- EGTA
- ethylene glycol bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid
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- Received July 15, 1996.
- Accepted October 22, 1996.
- The American Society for Pharmacology and Experimental Therapeutics
