12-O-tetradecanoylphorbol-13-acetate inhibits aminophospholipid translocase activity and modifies the lateral motions of fluorescent phospholipid analogs in the plasma membrane of bovine aortic endothelial cells.
The tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) is a potent mitogenic factor which can replace the growth promoting activity of basic fibroblast growth factor (bFGF) on bovine aortic endothelial cells. However, TPA-treated cells lose their strict contact inhibition at confluence, which is a characteristic of cells grown in the presence of bFGF. We have examined whether these changes could be related to modifications of the transbilayer and lateral motions of fluorescent lipids, namely 1-acyl-2-[6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]caproyl]-p hosphatidylcholine (C6-NBD-PC), -phosphatidylserine (C6-NBD-PS), and -phosphatidylethanolamine (C6-NBD-PE) inserted in the outer leaflet of the cell plasma membrane. In TPA-treated cells, the three fluorescent phospholipids remained located in the outer leaflet for at least 1 h at 20 degrees C after their insertion, indicating a blockade of the aminophospholipid translocase activity which is normally present in the plasma membrane of bFGF-treated cells. TPA also induced a large increase in the percentage of C6-NBD-PC and C6-NBD-PE probes which were free to diffuse laterally. The mobile fractions M reached values of approximately 100% for the two lipids, while for bFGF-treated cells they were found around 85 and 75%, respectively. For the C6-NBD-PS probe, M remained unchanged in bFGF and TPA-treated cells, at around 85%. TPA treatment also induced a twofold increase in the lateral diffusion coefficients of C6-NBD-PC and C6-NBD-PE, while that of C6-NBD-PS remained nearly unchanged. These effects of TPA may be related to the observed loss of differentiated properties of vascular endothelial cells and not to its mitogenic properties.
Julien
M
M
Institut de Pharmacologie et de Biologie Structurale du CNRS, 118, Toulouse, France.
Millot
C
C
Tocanne
J F
JF
Tournier
J F
JF
D009327:4-Chloro-7-nitrobenzofurazan / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000166:cytology; D002273:Carcinogens / Q000494:pharmacology*; D002352:Carrier Proteins / Q000037:antagonists & inhibitors*; D002417:Cattle; D002462:Cell Membrane / Q000737:chemistry / Q000201:enzymology*; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D005456:Fluorescent Dyes / Q000378:metabolism / Q000494:pharmacology; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D008565:Membrane Proteins / Q000037:antagonists & inhibitors*; D010713:Phosphatidylcholines / Q000378:metabolism / Q000494:pharmacology; D010714:Phosphatidylethanolamines / Q000378:metabolism / Q000494:pharmacology; D010718:Phosphatidylserines / Q000378:metabolism / Q000494:pharmacology; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids / Q000037:antagonists & inhibitors; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
C092351:1-palmitoyl-2-(6-((7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)caproyl)glycero-3-phosphoserine; D002273:Carcinogens; D002352:Carrier Proteins; D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D008565:Membrane Proteins; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010718:Phosphatidylserines; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids; D016222:Fibroblast Growth Factor 2; C052267:1,2-(palmitoyl-NBD-aminocaproyl)phosphatidylethanolamine; C044745:1-palmitoyl-2-(6-(N-(7-nitrobenz)-2-oxa-1,3-diazol-4-yl)aminocaproyl)phosphatidylcholine; D009327:4-Chloro-7-nitrobenzofurazan; D013755:Tetradecanoylphorbol Acetate