Membrane Binding by tBid Initiates an Ordered Series of Events Culminating in Membrane Permeabilization by Bax

Abstract

In normal circumstances, the Bcl-2 family dutifully governs when cells die. However, the rules of engagement between the pro- and antiapoptotic family members are still contested, and how Bax is transformed from a cytosolic monomer to an outer mitochondrial membrane-permeabilizing oligomer is unclear. With fluorescence techniques and an in vitro system, the combination of tBid and Bax produced dramatic membrane permeabilization. The membrane is not a passive partner in this process beause membranes are required for the protein-protein interactions to occur. Simultaneous measurements of these interactions revealed an ordered series of steps required for outer membrane permeabilization: (1) tBid rapidly binds to membranes, where (2) tBid interacts with Bax, causing (3) Bax insertion into membranes and (4) oligomerization, culminating in (5) membrane permeabilization. Bcl-XL prevents membrane-bound tBid from binding Bax. Bad releases tBid from Bcl-XL, restoring both tBid binding to Bax and membrane permeabilization.

Bax activation by Bim?

“…Do other BH3-only proteins, such as tBid, which has also been proposed to be an activator of Bax and interacts with Bax in a lipid environment, 30 trigger Bax and/or Bak in a similar manner?…”

Bcl-xL phosphorylation at Ser49 by polo kinase 3 during cell cycle progression and checkpoints

“…Bax insertion and oligomerization into membranes require activation, i.e. structural reorganization by a BH3-only activating protein, events that lead to outer mitochondrial membrane permeabilization; in contrast, the ability of anti-apoptotic proteins, such as Bcl-xL, to trap and inhibit these BH3-only activating proteins, prevents membrane permeabilization [17][18][19][20].…”

“…Structural similarities between Bcl-xL, particularly its α5-and α6-helices and the pore-forming domains of some bacterial toxins that act as channels for either ions or proteins, suggest that Bcl-2 members could function by constituting pores in intracellular organelles, including mitochondria [5,[17][18][19][20][21]. Whether these channel activities function by themselves, or in association with other megachannels, such as components of mitochondrial permeability transition pores, or others, is still not completely elucidated [3,4,19,[22][23][24][25][26][27][28][29][30][31].…”

Dissociation of Bak α1 helix from the core and latch domains is required for apoptosis

“…A carboxy (C)-terminal helix, a9, anchors Bak constitutively in the mitochondrial outer membrane (MOM) 5,6 , whereas in Bax, the a9 helix is initially sequestered in the hydrophobic surface groove, rendering Bax largely cytosolic 4,7 . When cells experience sufficient stress, the BH3-only class of Bcl-2 family proteins cause Bak and Bax to undergo a series of conformation changes-collectively referred to as 'activation'-that culminate in dimerization, leading to pore formation in the MOM [8][9][10] . Pore formation releases cytochrome c into the cytosol to initiate caspase activation and dismantle the cell 11 .…”