Fig. 6: A BAK-nucleated complex including PHB1/2 and STOM mediates BAK-induced autophagy and inhibits ATP secretion during apoptosis.

a Selected list of prohibitin-family proteins and VDAC1 identified as potential latch interactors. The number of peptides identified in the WT and 5M proteomics samples, and their difference between both samples (DIFF.), are indicated. b Co-immunoprecipitation assay showing interaction between activated GST-BAK and PHB2/STOM in response to BIM. DKO MEFs depleted of CASP3 and CASP9 and expressing GST-BAK (and also human PHBs and STOM for increased detectability) were retrovirally transduced with HA-BIM and, 15 h later, crosslinked with 1% paraformaldehyde and lysed. Lysates were incubated with control or GSH-agarose beads and the resulting precipitates subjected to Western blot (left panel, IPs). Expression levels of all contenders in total protein lysates are shown on the right panel (TLs). c PHBs mediate the autophagic response induced by BIM (left panel) and tBID (right panel). Bax-/- MEFs were transduced with the shown CRISPR/Cas9 constructs and, 40 h later, with the indicated BH3-only molecules. Cells were treated with 25 μM zVAD.fmk 7 h later and lysed 20 h post-transduction for Western blot. Relative densitometric quantifications of LC3-II are shown at the bottom. d PHBs inhibit ATP secretion during BH3-only-induced apoptosis. Bax-/- MEFs were transduced with the shown CRISPR/Cas9 constructs and, subsequently, with the indicated BH3-only molecules (as in c) for 22 h, and the levels of extracellular ATP were measured at 14 and 22 h. Graph shows mean values −/+ s.d. of cumulative data resulting from the sum of the luciferase activity units obtained at both time points from triplicate experimental points (n = 3 biological replicas; *P < 0.05, ***P < 0.001, two-tailed Student’s t-test). Numeric P-values are shown. Control points were lysed 48 h post-CRISPR/Cas9 transduction for Western blot to determine PHB2 depletion (right panel). Source data are provided as a Source Data file.