Phosphatidylinositol 3-kinase/Akt positively regulates Fas (CD95)-mediated apoptosis in epidermal Cl41 cells.
Fas (CD95)-mediated apoptosis is an essential mechanism for the maintenance of homeostasis, and disruption of this death pathway contributes to many human diseases. The cell survival protein kinase Akt/protein kinase B (PKB) is a known regulator of apoptosis, but its role in Fas-mediated cell death and its regulatory mechanisms are unclear. In this study, we show that stimulation of the Fas receptor by its ligand (FasL) induces rapid phosphorylation of Akt/PKB and a parallel increase in cell apoptosis in epidermal Cl41 cells. Inhibition of PI3K/Akt by dominant-negative overexpression of PI3K (Deltap85) and Akt (Akt-T308A/S473A) protects the cells from apoptosis, indicating an unexpected proapoptotic role of PI3K/Akt in the Fas signaling process. Treatment of the cells with pharmacological inhibitors of PI3K, wortmannin and 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-1 (LY294002), similarly inhibits FasL-induced apoptosis and Akt/PKB phosphorylation, indicating that PI3K is an upstream mediator of Akt/PKB and is involved in Fas-mediated cell death. Electron spin resonance studies show that FasL treatment induces rapid generation of reactive oxygen species, and inhibition of ROS by antioxidants effectively inhibits Akt/PKB signaling, suggesting that FasL activation of Akt/PKB is redox sensitive. In cells transfected with dominant-negative PI3K/Akt, Fas expression is down-regulated, but FLIP expression is unaffected. Reporter gene and mRNA expression assays show that FasL activates fas transcriptional activity and this effect is inhibited by PI3K/Akt suppression. Together, our results indicate that the PI3K/Akt, in addition to its normal prosurvival role, also plays an apoptotic role in Fas-mediated cell death through a mechanism that involves transcriptional activation of Fas receptor.
Lu
Bin
B
Department of Pharmaceutical Sciences, West Virginia University, Morgantown, WV 26506, USA.
Wang
Liying
L
Stehlik
Christian
C
Medan
Djordje
D
Huang
Chuanshu
C
Hu
Shuiying
S
Chen
Fei
F
Shi
Xianglin
X
Rojanasakul
Yon
Y
D000730:Androstadienes / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000276:immunology*; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects / Q000276:immunology; D000078404:Epidermal Cells; D004817:Epidermis / Q000201:enzymology* / Q000276:immunology* / Q000378:metabolism; D053222:Fas Ligand Protein; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism / Q000502:physiology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D017382:Reactive Oxygen Species / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000276:immunology; D000079424:Tumor Necrosis Factor Inhibitors; D048069:Tumor Necrosis Factors / Q000378:metabolism / Q000502:physiology; D000077191:Wortmannin; D019014:fas Receptor / Q000096:biosynthesis / Q000378:metabolism* / Q000502:physiology
D000730:Androstadienes; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D002867:Chromones; D053222:Fas Ligand Protein; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D000079424:Tumor Necrosis Factor Inhibitors; D048069:Tumor Necrosis Factors; D000077191:Wortmannin; D019014:fas Receptor; C506386:CFLAR protein, human; C505661:FASLG protein, human; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one