Bid-induced cytochrome c release is mediated by a pathway independent of mitochondrial permeability transition pore and bax

Bid, a pro-apoptosis "BH3-only" member of the Bcl-2 family, can be cleaved by caspase-8 after Fas/TNF-R1 engagement. The p15 form of truncated Bid (tB id) translocates to mitochondria and induces cytochrome c release, leading to the activation of downstream caspases and apoptosis. In the current stud y, we investigated the mechanism by which tBid regulated cytochrome c relea se in terms of its relationship to mitochondrial permeability transition an d Bax, another Bcl-2 family protein. We employed an in vitro reconstitution system as well as cell cultures and an animal model to reflect the physiol ogical environment where Bid could be functional. We found that induction o f cytochrome c release by tBid was not accompanied by a permeability transi tion even at high doses. Indeed, inhibition of permeability transition did not suppress the activity of tBid in vitro nor could they block Pas activat ion-induced, Bid-dependent hepatocyte apoptosis in cultures. Furthermore, M g2+, although inhibiting permeability transition, actually enhanced the abi lity of tBid to induce cytochrome c release. We also found that tBid did no t require Bax to induce cytochrome c release in vitro. In addition, mice de ficient in bat were still highly susceptible to anti-Fas-induced hepatocyte apoptosis, in which cytochrome c release was unaffected. Moreover, althoug h Bar-induced cytochrome c release was not dependent on tBid, the two prote ins could function synergistically. We conclude that Bid possesses the bioc hemical activity to induce cytochrome c release through a mechanism indepen dent of mitochondrial permeability transition pore and Bax.