INHIBITION OF AKT KINASE BY CELL-PERMEABLE CERAMIDE AND ITS IMPLICATIONS FOR CERAMIDE-INDUCED APOPTOSIS

Ceramide is an important lipid messenger involved in mediating a varie ty of cell functions including apoptosis. However, mechanisms responsi ble for ceramide-induced apoptosis remain unclear. We investigated the possibility that ceramide may decrease antiapoptotic signaling in cel ls by inhibiting Akt kinase activity. Our data show that C-2-ceramide induces apoptosis in HMN1 motor neuron cells and decreases both basal and insulin- or serum-stimulated Akt kinase activity 65-70%. These res ults are consistent with decreased Akt kinase activity being involved in the apoptotic effects of ceramide, This possibility is further supp orted by studies showing that constitutively active Akt kinase decreas es C-2-ceramide-induced death of HMN1 cells as well as COS-7 cells. De creased Akt activity is not due to ceramide activating the ceramide-ac tivated protein phosphatase or to a direct inhibition of Akt kinase by ceramide, suggesting that ceramide acts upstream of Akt kinase to dec rease its activity. Treating cells with C-2-ceramide does not affect p hosphorylation of insulin receptor substrate-1, interactions between i nsulin receptor substrate-1 and p85, or insulin-stimulated phosphatidy linositol 3-kinase activity, suggesting that the effects of C-2-cerami de on Akt kinase are not mediated through modulating phosphatidylinosi tol 3-kinase. In sum, our results suggest that inhibition of the key a ntiapoptotic kinase, Akt, may play an important role in ceramide-induc ed apoptosis.