Distinct Forms of Mitochondrial TOM-TIM Supercomplexes Define Signal-Dependent States of Preprotein Sorting

Abstract

Mitochondrial import of cleavable preproteins occurs at translocation contact sites, where the translocase of the outer membrane (TOM) associates with the presequence translocase of the inner membrane (TIM23) in a supercomplex. Different views exist on the mechanism of how TIM23 mediates preprotein sorting to either the matrix or inner membrane. On the one hand, two TIM23 forms were proposed, a matrix transport form containing the presequence translocase-associated motor (PAM; TIM23-PAM) and a sorting form containing Tim21 (TIM23^SORT). On the other hand, it was reported that TIM23 and PAM are permanently associated in a single-entity translocase. We have accumulated distinct transport intermediates of preproteins to analyze the translocases in their active, preprotein-carrying state. We identified two different forms of active TOM-TIM23 supercomplexes, TOM-TIM23^SORT and TOM-TIM23-PAM. These two supercomplexes do not represent separate pathways but are in dynamic exchange during preprotein translocation and sorting. Depending on the signals of the preproteins, switches between the different forms of supercomplex and TIM23 are required for the completion of preprotein import.

We thank Agnes Schulze-Specking, Inge Perschil, Birgit Schönfisch, and Klaus Paal for expert technical assistance.

This work was supported by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 746, Excellence Initiative of the German Federal & State Governments (EXC 294), Landesforschungspreis Baden-Württemberg, Gottfried Wilhelm Leibniz Program, and the Fonds der Chemischen Industrie.