Philipp Lachenmann: DELPHI Rationale


Young Investigator Award 2005

Dr. Peter Rehling

Institut für Biochemie und Molekularbiologie, Albert-Ludwigs-Universität Freiburg

Protein translocation across the inner mitochondrial membrane

Mitochondria are the powerhouses of eukaryotic cells since they fulfill a central function in cellular energy metabolism. They use more than 95% of the oxygen required by cells to generate ATP, which then serves to fuel cellular processes. In addition, mitochondria participate in many other important tasks and therefore defects in mitochondrial functions are often lethal or lead to severe diseases in man. Nuclear genes encode 99% of the 800 to 1500 proteins that constitute the mitochondrial proteome while the mitochondrial genome encodes only few proteins. Thus, proteins need to be transported from the cytosol into one of the four mitochondrial subcompartments, the outer membrane, intermembrane space, inner membrane or matrix. We are interested in how proteins are transported from the cytosol into mitochondria and especially how they pass across the inner membrane.

Work in several laboratories using yeast as a model system, has allowed some insight into how this process occurs. After synthesis of proteins in the cytosol receptors on the surface of mitochondria recognize signals in proteins destined for mitochondria. Transport across the outer membrane is mediated by a multi-protein machinery (TOM complex). Transport across the inner membrane requires additional translocation machineries and two different protein complexes mediate this transport step. Metabolite carrier proteins use the TIM22 complex for insertion into the inner membrane while proteins with amino terminal signals are transported by the TIM23 complex. To transport proteins into the matrix the TIM23 complex cooperates with the motor complex (PAM). The central component of PAM is the chaperone Hsp70, which binds precursor proteins and promotes their transport into the matrix by cycles of binding and release.

We have focused our analyses on the two TIM complexes and investigated their protein composition and the mechanism of protein translocation. Our analyses have shown that different mechanisms are used to transport precursor proteins across the inner membrane and that the translocation machineries are very dynamic protein complexes. Upon isolation and reconstitution of the TIM22 complex we have found that it contains two functionally coupled pores. This translocase uses the electrochemical gradient across the inner membrane to provide the driving force necessary to insert the carrier protein into the membrane in a multi-step process. Moreover, my group isolated the TIM23 complex together with the PAM complex and identified a number of new components. We have found that during protein transport the Tim21 protein binds to the TOM complex and allows transfer of the precursor proteins to the TIM23 complex. Depending on the precursor, molecular rearrangements can occur within the translocation machinery. For matrix transport, the PAM complex is recruited to the translocase and Tim21 is released. The identification of new proteins of the PAM complex revealed that the activity of Hsp70 in the PAM complex is tightly regulated. We are now investigating the molecular role of the different Tim and Pam proteins during protein transport and address the dynamic of the complexes during protein import.



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