Biochemical and genetic analysis of the mitochondrial response of yeast toBAX and BCL-X-L

The BCL-2 family includes both proapoptotic (e.g., BAX and BAK) and antiapo ptotic (e.g., BCL-2 and BCL-X-L) molecules. The cell death-regulating activ ity of BCL-2 members appears to depend on their ability to modulate mitocho ndrial function, which may include regulation of the mitochondrial permeabi lity transition pore (PTP). We examined the function of BAX and BCL-X-L usi ng genetic and biochemical approaches in budding yeast because studies with yeast suggest that BCL-2 family members act upon highly conserved mitochon drial components. In this study we found that in wild-type yeast, BAX induc ed hyperpolarization of mitochondria, production of reactive oxygen species , growth arrest, and cell death; however, cytochrome c was not released det ectably despite the induction of mitochondrial dysfunction. Coexpression of BCL-X-L prevented all BAX-mediated responses. We also assessed the functio n of BCL-X-L and BAX in the same strain of Saccharomyces cerevisiae with de letions of selected mitochondrial proteins that have been implicated in the function of BCL-2 family members. BAX-induced growth arrest was independen t of the tested mitochondrial components, including voltage-dependent anion channel (VDAC), the catalytic beta subunit or the delta subunit of the F0F 1-ATP synthase, mitochondrial cyclophilin, cytochrome c, and proteins encod ed by the mitochondrial genome as revealed by [rho(0)] cells. In contrast, actual cell killing was dependent upon select mitochondrial components incl uding the beta subunit of ATP synthase and mitochondrial genome-encoded pro teins but not VDAC. The BCL-X-L protection from either BAX-induced growth a rrest or cell killing proved to be independent of mitochondrial components. Thus, BAX induces two cellular processes in yeast which can each be abroga ted by BCL-X-L: cell arrest, which does not require aspects of mitochondria l biochemistry, and cell killing, which does.