Leaky Protons

REFERENCE: Alavian et al. “Bcl-X_L regulated metabolic efficiency of neurons through interaction with the mitochondrial F₁F₀ ATP synthase” Nature Cell Biology (2011) 13(10) pgs 1224 – 1233.

                B-cell lymphoma-extra large (Bcl-X_L), a member of the programmed cell death Bcl-2 family proteins, is the major anti-apoptotic protein in adult neurons.  When Bcl-X_L is overexpressed, mitochondria translocate to presynaptic sites, the number and size of synpases increase, and the overall mitochondrial biomass goes up.  Synaptic strengthening requires high metabolism but the exact involvement of Bcl-X_L in these events is not clear.

                In this month’s edition of Nature Cell Biology, Alavian et al. detail a role for Bcl-X_L in binding to F₁F₀ ATP synthase.  Previous subcellular localization studies have placed Bcl-X_L within the mitochondrial outer membrane, but new immunoelectron microscopy data published here supports various other studies that suggest the protein is at the mitochondrial inner membrane, as well.  Further analysis showed that Bcl-X_L was present with the F₁F₀ ATP synthase complex and endogenous Bcl-X_L was co-immunoprecipitated with the ß subunit of ATP synthase.

                To clarify the role of Bcl-X_L binding, the authors studied ATP hydrolysis by submitochondrial vesicles enriched with F₁F₀ ATP synthase protein complexes (Figure 6.1).  The experiment used the H⁺ fluorescent indicator ACMA, which is unable to be transported into the vesicles.  Upon ATP hydrolysis, fluorescence of ACMA dropped as H⁺ is pumped out of the buffer and into the vesicles by the F₁F₀ ATPase.  Treatment of the vesicles with proton pump inhibitors or compounds that create vesicle pores (and subsequent proton leaks) resulted in higher levels of fluorescence upon ATP hydrolysis.  Interestingly, these same results were also seen when vesicles were treated with Bcl-X_L inhibitors.

                The hypothesis that Bcl-X_L is acting at F₁F₀ ATPase to prevent proton leak was further supported by patch-clamp studies where the authors measured leak conductance.  Conductance dropped dramatically in the Bcl-X_L overexpressing vesicles when either ATP or ADP was added to the buffer.  Reducing the amount of Bcl-X_L in these vesicles by way of knockdown studies showed the conductance to be higher across these membranes.

                The authors also showed that in Bcl-X_L overexpressing neurons, the uptake of oxygen is lower but ATP levels are higher than in wild type neurons and that recombinant Bcl-X_L can directly increase the rate of ATP hydrolysis by F₁F₀ ATPase.  Taken together, Jonas and colleagues conclude that Bcl-X_L reduces proton leak during ATP synthesis, which thereby increases the neurons’ ATP synthesis efficiency and improves their metabolism.