Figure 7. Chaperone binding by Tom70 prevents mitoprotein-induced toxicity.

(A) Proteins that are enriched by the expression of mt-Tah1 compared with Δtom70/71 are shown. Only proteins with masses smaller than 18 kDa and positive enrichment factors were considered. Mitochondrial proteins are indicated in blue.

(B) Schematic representations of small inner membrane proteins for which information about their overall structure and targeting information exists. Blue regions show presequences, and black boxes indicate transmembrane domains.

(C and D) Cox5a-HA, Tim11-HA, Atp17-HA, and Atp18-HA were expressed under GAL1 control from multi-copy plasmids in wild-type and Δtom70/71 cells. The times indicate how long cells were shifted to 0.5% galactose-containing medium.

(E) Radiolabeled Cox5a was incubated with isolated mitochondria for the times indicated at 30°C. The membrane potential (Δψ) was depleted in control samples by addition of CCCP. Mitochondria were reisolated and incubated with or without proteinase K.

(F and G) The indicated strains were transformed with plasmids to express Atp17-HA, Tim11-HA, Cox5a-HA, and Pet9-HA under the control of the GAL1 promoter. All cultures were grown on lactate medium to mid-log phase, induced with 0.5% galactose for 4.5 h, and dropped onto galactose plates.

(H) Rpn4-driven gene expression was measured using a yellow fluorescent protein (YFP) reporter system (Boos et al., 2019).

(I) Tom70 supports the biogenesis of aggregation-prone mitochondrial membrane proteins by recruiting cytosolic chaperones to the mitochondrial surface, thereby generating a “mitochondria-associated proteophilic zone.”