Brefeldin A-mediated apoptosis requires the activation of caspases and is inhibited by Bcl-2

Brefeldin A (BFA) has recently been shown to induce apoptosis in human tumo r cells in a p53-independent fashion. in this study, BFA-induced apoptosis was analyzed in the human Jurkat T-cell line. Apoptosis occurred 8 h after treatment with BFA and at concentrations as low as 10 ng/ml and increased w ith the duration of BFA exposure. Forskolin, an inhibitor of BFA-induced de aggregation of the Golgi-microtubular complex in some cell lines, failed to reverse BFA-mediated apoptosis. Further study of the mechanism of BFA-indu ced apoptosis was conducted by using a series of peptide protease inhibitor s. Complete inhibition of cell death was achieved with benzyloxyearbonyl-va l-Ala-Asp-fluromethylketone, a peptide inhibitor of the caspase protease fa mily, and Z-Asp-Glu-Val-Asp-FMK, a specific inhibitor of caspase-3. Both Ac etyl-Tyr-Val-Ala-Asp-chloromethyl-ketone and Acetyl-Tyr-Val-Ala-Asp-aldehyd e, selective caspase-1 (interleukin-lp converting enzyme) inhibitors, exert ed only partial protection of cells from apoptosis at higher concentrations . Z-Phe-Ala-FMK, a cysteine protease inhibitor lacking aspartate at the pi position, did not have any impact on BFA-induced apoptosis. Furthermore, Ju rkat cells transfected with the proto-oncoprotein Bcl-2, which is able to b lock various apoptotic conditions, showed remarkable resistance to the apop totic effect of BFA. Thus, the data indicate that EFA-induced apoptosis req uires caspase(s) activation, primarily the activation of caspase-3, and is inhibited by overexpression of Bcl-2. (C) 1998 Academic Press.