Mammalian Bcl-x(L) protein localizes to the outer mitochondrial membrane, where it inhibits apoptosis by binding Bax and inhibiting Bax-induced outer membrane permeabilization. Contrary to expectation, we found by electron microscopy and biochemical approaches that endogenous Bcl-x(L) also localized to inner mitochondrial cristae. Two-photon microscopy of cultured neurons revealed large fluctuations in inner mitochondrial membrane potential when Bcl-x(L) was genetically deleted or pharmacologically inhibited, indicating increased total ion flux into and out of mitochondria. Computational, biochemical, and genetic evidence indicated that Bcl-x(L) reduces futile ion flux across the inner mitochondrial membrane to prevent a wasteful drain on cellular resources, thereby preventing an energetic crisis during stress. Given that F(1)F(O)-ATP synthase directly affects mitochondrial membrane potential and having identified the mitochondrial ATP synthase β subunit in a screen for Bcl-x(L)-binding partners, we tested and found that Bcl-x(L) failed to protect β subunit-deficient yeast. Thus, by bolstering mitochondrial energetic capacity, Bcl-x(L) may contribute importantly to cell survival independently of other Bcl-2 family proteins.

Following exocytosis, the rate of recovery of neurotransmitter release is determined by vesicle retrieval from the plasma membrane and by recruitment of vesicles from reserve pools within the synapse, which is dependent on mitochondrial ATP. The anti-apoptotic Bcl-2 family protein Bcl-xL also regulates neurotransmitter release and recovery in part by increasing ATP availability from mitochondria. We now find, that Bcl-xL directly regulates endocytic vesicle retrieval in hippocampal neurons through protein-protein interaction with components of the clathrin complex. Our evidence suggests that, during synaptic stimulation, Bcl-xL translocates to clathrin-coated pits in a calmodulin-dependent manner and forms a complex with the GTPase Drp1, Mff and clathrin. Depletion of Drp1 produces misformed endocytic vesicles. Mutagenesis studies suggest that formation of the Bcl-xL-Drp1 complex is necessary for the enhanced rate of vesicle endocytosis produced by Bcl-xL, thus providing a mechanism for presynaptic plasticity.

Anti-apoptotic Bcl2 family proteins such as Bcl-x(L) protect cells from death by sequestering apoptotic molecules, but also contribute to normal neuronal function. We find in hippocampal neurons that Bcl-x(L) enhances the efficiency of energy metabolism. Our evidence indicates that Bcl-x(L)interacts directly with the β-subunit of the F(1)F(O) ATP synthase, decreasing an ion leak within the F(1)F(O) ATPase complex and thereby increasing net transport of H(+) by F(1)F(O) during F(1)F(O) ATPase activity. By patch clamping submitochondrial vesicles enriched in F(1)F(O) ATP synthase complexes, we find that, in the presence of ATP, pharmacological or genetic inhibition of Bcl-x(L) activity increases the membrane leak conductance. In addition, recombinant Bcl-x(L) protein directly increases the level of ATPase activity of purified synthase complexes, and inhibition of endogenous Bcl-x(L) decreases the level of F(1)F(O) enzymatic activity. Our findings indicate that increased mitochondrial efficiency contributes to the enhanced synaptic efficacy found in Bcl-x(L)-expressing neurons.