6th ESACP Congress, Heidelberg, April 7-11, 1999

A112
WANDERING IN APOPTOSIS WITH FLOW CYTOMETRY: FROM CELL TO MITOCHONDRIA.
Petit PX

INSERM U129, ICGM, Paris, France

Both physiological cell death (apoptosis) and at least some cases of accidental cell death involve a two-step-process. At the first level, numerous physiological or pathological stimuli can trigger mitochondrial permeability transition which constitutes a ratelimiting event and initiates the common phase of the death process. Mitochondrial permeability transition (PT) involves the formation of proteaceous, regulated pore, probably by apposition of inner and outer mitochondrial membrane proteins which cooperate to form the PT pore complex (reconstituted into liposomes). Inhibition of the PT by pharmacological intervention on mitochondrial structures or mitochondrial expression of the apoptosisinhibitory oncoprotein Bcl-2 thus can prevent cell death (all the steps are followed by flow cytometric analysis). At a second level, the consequences of mitochondrial dysfunction (studied on isolated mitochondria or in a cell-free system) can entail a bioenergetic catastrophy and/or the activation and action of apoptogenic proteases with secondary endonuclease activation and consequent oligonucleosomal DNA fragmentation (apoptosis). The acquisition of the biochemical and ultrastructural features of apoptosis relies on the liberation of apoptogenic proteins (AIF) or protease activators (cytochrome c) from the inter membrane space. The notion that the mitochondrial events control cell death has major implications for the developpement of death-inhibitory drugs. In conclusion, AIF provides a new molecular link between mitochondrial membrane permeabilization and nuclear apoptosis, and that the caspases, DFF/CAD and AIF are probably engaged in complementary cooperative or redundant pathways that lead to nuclear apoptosis.