Figure 1.

Phb1 is required for mitochondrial inner membrane integrity in the absence of Gep1. (A) Multiple sequence alignment (score matrix: Blosum62) of Gep1, Gep2, Ups1, and human homologues. The conserved MSF1′/PRELI domain is depicted. Numbers refer to amino acids. Black highlighting indicates full conservation across all species; gray highlighting represents amino acids with similar properties conserved in at least three of the analyzed sequences. (B) Synthetic lethal interaction of Δgep1 and Δphb1. Fivefold serial dilutions of cell suspensions were spotted on glucose (Glc)- or galactose (Gal)-containing YP plates, which were supplemented with 2 µg/ml doxycycline (Dox) when indicated and incubated at 30°C. WT, wild type. (C) Steady-state levels of mitochondrial proteins in Δgep1Δphb1[PHB1] cells. Mitochondria were isolated from cells grown on glucose-containing media in the presence of doxycycline for different time periods. Mitochondrial proteins were analyzed by SDS-PAGE and immunoblotting. IM, inner membrane; OM, outer membrane; M, matrix; L, long isoform of Mgm1; S, short isoform of Mgm1. (D) Dissipation of the membrane potential in mitochondria lacking Gep1 and Phb1. Mitochondria were isolated from cells grown for 12 h on glucose-containing media in the presence or absence of doxycycline and stained with the potential-sensitive dye 3,3′-dipropylthiadicarbocyanine iodide (DiDC₃(5)).