Project Details
Projekt
Early steps in the biogenesis of mitochondrial inner membrane proteins: The role of cytosolic factors in synthesis, targeting and quality control
Applicants
Professor Dr. Johannes M. Herrmann; Dr. Stefan Pfeffer; Professorin Dr. Konstanze Winklhofer
Subject Area
Biochemistry
Term
since 2024
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 541210481
Mitochondria consist of hundreds of proteins which initially are synthesized as precursor proteins in the cytosol. Most of these precursors are imported post-translationally after their synthesis was completed. Since the translocation reaction is much faster (seconds) than protein synthesis (minutes), the post-translational import mode is assumed to reduce the number of required import sites and to decrease the risk of dissipating the inner membrane potential by proton leakage on translocation intermediates. However, there is one class of mitochondrial precursor proteins for which a co-translational import mode was demonstrated by different experimental approaches: Proteins which are inserted into the inner membrane by lateral insertion of stop-anchor sequences often have large domains following their transmembrane domain. These C-terminal domains must traverse the outer membrane without access to the import motor which resides in the matrix. There is accumulating evidence that these C-terminal domains are imported co-translationally and that cytosolic ribosomes facilitate their translocation across the outer membrane. The details of this translocation process are however still unknown. For this project, we will combine the expertise of three laboratories representing three different fields of biology to study the co-translational import of stop-anchored inner membrane proteins. We will address three major aspects: First, we will identify the mechanisms of the translocation process and study the features of these proteins that prevent post-translational import. Preliminary data show that poly-proline stretches in the sequence of these proteins serve as arrest peptides which require the activity of a specific translation factor, called eIF5A in humans or Hyp2 in yeast, before translation can commence. Second, we will use cellular cryo-electron tomography to study the molecular structure and specific arrangement of ribosomes on the mitochondrial surface under wild type situations as well as upon conditions at which the stalling of precursor proteins is induced (such as upon expression of poly-proline mutant proteins or in eIF5A/Hyp2 deficient cells). Third, we will study the removal of stalled translocation intermediates by cytosolic quality control components. This project is only possible because the three groups bring together very complementary expertise in mitochondrial biology, proteostasis research, and structural biology of ribosomes. With this team, we will study the situation in both yeast and human cells to identify evolutionary conserved basic mechanisms in the biogenesis of stop-anchored mitochondrial proteins. The integration of this project into the general expertise of the priority program provides a unique opportunity to study this so far uncharacterized aspect of mitochondrial biology.
DFG Programme
Priority Programmes
