FAS OR CERAMIDE INDUCE APOPTOSIS BY RAS-REGULATED PHOSPHOINOSITIDE-3-KINASE ACTIVATION

We demonstrate a rapid and transient activation of phosphoinositide-3- kinase (PI-3-K) by Fas receptor triggering or cellular treatment with synthetic CG-ceramide. The stimulation of PI-3-K is critical for Fas o r CG-ceramide-induced programmed cell death because transfection with a transdominant inhibitory PI-3-K construct or pretreatment with the P I-3-K inhibitor wortmannin almost completely prevented Fas or CG-ceram ide-mediated apoptosis. Treatment with the caspase inhibitor Ac-YVAD-c mk or cellular transfection with transdominant inhibitory N17Ras preve nted PI-3-K stimulation by Fas, suggesting that Fas activates PI-3-K v ia caspases and Ras. N17Ras expression also prevented CG-ceramide-init iated PI-S-K stimulation. The notion of a PI-3-K regulation by Ras upo n Fas receptor ligation or ceramide treatment is supported by co-immun oprecipitation experiments revealing an activation-dependent associati on of PI-3-K and Ras.