This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Kaszkin, M. Articles by Kinzel, V. Search for Related Content PubMed PubMed Citation Articles by Kaszkin, M. Articles by Kinzel, V.

Proposed Role of Phosphatidic Acid in the Extracellular Control of the Transition from G₂ Phase to Mitosis Exerted by Epidermal Growth Factor in A431 Cells¹

Department of Pathochemistry, German Cancer Research Center, Im Neuenheimer Feld 280, D-6900 Heidelberg, Germany

Epidermal growth factor (EGF) has been shown to cause an inhibition of A431 cells in G₂ phase within approximately 10 min, i.e., shortly before mitosis (Kinzel et al., Cancer Res., 50: 7932–7936, 1990). This system has been used to study the proposed role phospholipid metabolites, particularly phosphatidic acid (PA), may play (Kaszkin et al., Cancer Res., 51: 4328–4335, 1991) in the extracellular control of cells at the physiological restriction site in G₂ phase. A431 cells responded to EGF with a dose-dependent formation of phosphatidic acid (PA) which correlated with the dose-dependent G₂ delay as well as with their time courses. The G₂ delay induced by EGF as well as PA mobilization were effected in conditioned medium or in fresh medium containing bovine serum albimun instead of serum, i.e., under the conditions necessary for precursor studies to be carried out. The major pathway of PA formation was probably via phospholipase C-mediated breakdown of phosphatidylinositol and diacylglycerol kinase: (a) the dose response of PA formation correlated with that of total inositol phosphate accumulation; (b) little diacylglycerol was found and then only at a high EGF concentration; (c) prelabeling with [1-¹⁴C]arachidonic acid resulting in a large specific labeling of phosphatidylinositol led to an EGF-induced, dose-dependent formation of radioactive arachidonyl-PA (correlated with that of total PA and inositol phosphate), but in the presence of a primary alcohol not to the formation of radioactive phosphatidylalcohol; (d) prelabeling with [1-¹⁴C]oleic acid led to the EGF-induced formation of labeled PA, which in the presence of a primary alcohol was only slightly reduced to the advantage of very low levels of labeled phosphatidyl alcohol, thus demonstrating that an EGF-effected activation of phospholipase D did occur but contributed little to the general PA level. An alternative mobilization of PA was attempted with the phorbolester 12-O-tetradecanoylphorbol-13-acetate (TPA), which was shown to activate phospholipase D in A431 cells and to elicit PA from a phospholipid pool which was not significantly labeled with radioactive arachidonic acid. The TPA-induced degree of PA formation and of the G₂ delay correlated. Both phenomena were considerably larger with fresh medium containing 0.5% bovine serum albumin instead of serum than in conditioned medium. Another attempt to increase the cellular PA level consisted in a treatment of A431 cells with exogenous phospholipase D (2 units/ml) from Streptomyces chromofuscus. It caused an increase of the PA levels and induced a parallel G₂ delay. Modulation of the EGF receptor by a 15-min pretreatment with TPA in conditioned medium diminished the EGF-effected G₂ delay and reduced at the same time the EGF-induced formation of PA. Down-regulation of TPA-sensitive protein kinase C by a 17-h pretreatment with 10^-7M TPA abolished neither the G₂ delay nor the PA formation effected by EGF, thus indicating that the two phenomena correlated and were independent of TPA-sensitive protein kinase C. Taken together, the observed correlations between the mobilization of PA and the G₂ delay effected and influenced by different extracellular means argue for a role of PA as a possible mediator involved in the extracellular control of the transition from G₂ phase to mitosis.

¹ Supported by the Deutsche Forschungsgemeinschaft. Dedicated to Gerald C. Mueller, Professor of Oncology at the McArdle Laboratory for Cancer Research at the University of Wisconsin, Madison, on the occasion of his retirement and in appreciation for his contributions to aspects of cell cycle control and of phospholipid metabolism.

² To whom requests for reprints should be addressed.

Received 4/ 1/92. Accepted 8/ 4/92.

This article has been cited by other articles:

				J. A. Jones and Y. A. Hannun Tight Binding Inhibition of Protein Phosphatase-1 by Phosphatidic Acid. SPECIFICITY OF INHIBITION BY THE PHOSPHOLIPID J. Biol. Chem., May 3, 2002; 277(18): 15530 - 15538. [Abstract] [Full Text] [PDF]

				S. Tomic, U. Greiser, R. Lammers, A. Kharitonenkov, E. Imyanitov, A. Ullrich, and F.-D. Böhmer Association of SH2 Domain Protein Tyrosine Phosphatases with the Epidermal Growth Factor Receptor in Human Tumor Cells J. Biol. Chem., September 8, 1995; 270(36): 21277 - 21284. [Abstract] [Full Text] [PDF]

				M. Grange, C. Sette, M. Cuomo, M. Conti, M. Lagarde, A.-F. Prigent, and G. Nemoz The cAMP-specific Phosphodiesterase PDE4D3 Is Regulated by Phosphatidic Acid Binding. CONSEQUENCES FOR cAMP SIGNALING PATHWAY AND CHARACTERIZATION OF A PHOSPHATIDIC ACID BINDING SITE J. Biol. Chem., October 20, 2000; 275(43): 33379 - 33387. [Abstract] [Full Text] [PDF]