Analysis of MICOS independent Contact Sites between the inner and the outer membrane of mitochondria

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Mitochondria are essential in eukaryotic cells. They produce energy in from of ATP and are involved in a multitude of additional functions. For instance, Fe-S-cluster, heme, lipids and proteins are synthesized in mitochondria. In order to perform all these functions, mitochondria have to generate and maintain a complex ultrastructure. This has to be adapted in response to the different metabolic states of the cell. Mitochondrial ultrastructure represents the shape of the membrane system. In contrast to most of the other organelles, mitochondria are surrounded by two membranes, the mitochondrial inner and outer membrane. Structural elements, the contact sites, connect these two membranes. These contact sites consist of membrane protein complexes present in both, the mitochondrial inner and outer membrane. Proteins, signal molecules and metabolites have to be exchanged across both membranes. Therefore, contact sites are supposed to be highly important for the functionality of mitochondria and thus for the cell. Contact sites are observed upon electron microscopy analysis for a long time. However, the molecular nature and the functions of contact sites are largely unknown, in contrast to those mediated by the MICOS complex, which is essential for generation of mitochondrial ultrastructure.The data we obtained so far show that in addition to MICOS further contact sites exist. Moreover, we already identified a set of six mitochondrial proteins, which qualify for contact site proteins. This set includes three proteins of the inner mitochondrial membrane and three of the outer mitochondrial membrane. We identified these candidates in our previous screen for contact site proteins, which led to the identification of MICOS. However, these proteins are not subunits of MICOS. Thus, they are in likely components of new, yet uncharacterized contact sites. To extend the list of candidate proteins, we will repeat the screen for contact site proteins with significant modifications, such as analysis of MICOS deficient mitochondria. This guarantees that the new candidates are components of contact sites, which exist in addition to MICOS. The candidates will be a basis for the identification of new contact sites by co-isolation analyses. Finally, we will functionally characterize the new contact sites with a combination of biochemical, cell-biological and biophysical methods. All contact site proteins, which were identified in yeast so far, are evolutionary highly conserved. Therefore, we are convinced that the results obtained in the proposed project are highly important also for higher eukaryotes, just as it is the case with MICOS complex.