Extended Data Fig. 6: Decreased mitochondrial PE promotes YME1L-mediated proteolysis.

a, b, Phospholipid analysis of mitochondrial fractions from HeLa cells treated with Torin1 for 4 h (n = 3 independent experiments); a, relative distribution; b, absolute abundance. c, Acyl chain composition of mitochondrial PE from HeLa cells treated with Torin1 for 4 h (n = 3 independent experiments). d, Phospholipid analysis of mitochondrial fractions from WT, PRELID3B^−/− and PRELID3B^−/− HeLa cells expressing PRELID3B–Flag or PRELID3B(T57K)–Flag (n = 3 independent experiments). e, Quantification of protein levels from Fig. 2h (n = 4 independent experiments). f, g, Immunoblot and mitochondrial phospholipid analysis of scrambled control (Scr) or Prelid3b siRNA-transfected HeLa cells. Quantification of indicated protein levels is shown (immunoblot, n = 6 independent experiments; phospholipid analysis, n = 3 independent experiments). h, NBD-PS transfer by PRELID3B (black circles), PRELID3B(T57K) (red circles) or without addition of protein (white circles). PRELID3B(T57K) contains a mutation in the PS-binding site, which abolishes PS transfer but does not interfere with the assembly of PRELID3B into lipid transfer complexes. Average of n = 3 independent experiments. i, SDS–PAGE analysis of recombinant PRELID3B–TRIAP1 complexes and the T57K variant (40 pmol) by CBB staining (n = 1 experiment). j, Quantification of protein levels from Fig. 2i (n = 5 independent experiments). k, l, Phospholipid analysis of the mitochondrial fraction from HeLa cells treated with scrambled control (Scr) or Pisd siRNA (n = 3 independent experiments) (k), WT and Tsc2^−/− MEFs (n = 3 independent experiments) (l). Mean ± s.e.m.; two tailed t-test (a, b, f, g, j–l), one-way ANOVA with Dunnett’s multiple comparisons test (d, e). fc, fold change.

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