|dc.description.abstracteng||Mitochondria are essential components of eukaryotic cells. They consist of an outer and inner membrane, which itself is divided into the inner boundary membrane and the cristae. The transition from the inner boundary membrane and the cristae is termed cristae junctions. Essential for the maintenance of these cristae junctions is the multi-protein complex MICOS (mitochondrial contact site and cristae organizing system). Previous studies have focused on the characterization of MICOS and on the identification of its components and their interaction partners, but to date little is known about alterations of the submitochondrial protein distribution as a reaction to external stimuli. Even though electron microscopy has shown that the interior architecture of mitochondria adapts to the energy demand of the cells, it is not known whether the submitochondrial protein distribution of the MICOS complex is altered. In order to investigate the protein clusters of certain MICOS components, we selected two processes in which mitochondria play a central role: first, the energy supply of the neuromuscular junctions after increased physical strain and the alterations of mitochondria during apoptosis. By investigating the submitochondrial protein distribution using fluorescence microscopy and quantitative image analysis of the protein clusters, I showed the adaptation of the protein distribution of MICOS components.
In the first part of this work I used fluorescence microscopy to depict alterations of the neuromuscular junctions in general and of the submitochondrial protein distribution of Mic60 in particular, in reaction to enhanced physical strain. The cross-sectional area of motor endplates were slightly enlarged in the Soleus but not in the Tibialis anterior following enhanced activity of the innervated muscle. The submitochondrial protein distribution of Mic60 was more heterogeneous in the Soleus but was unchanged in the Tibialis anterior in reaction to the endurance exercises. The observed changes after the training stimuli, both to the area of the endplates as well as to the submitochondrial protein distribution of Mic60, were extremely subtle, but could be detected due to the high sensitivity both of the imaging and of the analysis procedures. The data shows muscle-specific alterations of the submitochondrial protein distribution of Mic60 and of the total area of motor endplate in reaction to external signals.
The second process to be studied was the programmed cell death, or apoptosis. I observed highly significant alterations of the submitochondrial protein distribution of all investigated proteins, among them three components of the MICOS complex, after induction of apoptosis. A loss of distinct protein clusters took place before the release of cytochrome c, a pro-apoptotic factor which residues in the cristae. For the release of cytochrome c to take place, it is essential that the outer mitochondrial membrane is permeabilized (MOMP, mitochondrial outer membrane permeabilization) by proteins of the Bcl-2 family, among them Bax. MOMP is the point in time after which the onset of apoptosis is normally irreversible. By investigating Bax using 2D and 3D STED microscopy, I showed that the Bax-proteins form ring-like structures that colocalize with fragmented mitochondria.
In summary the data shows subtle, muscle-specific changes of the submitochondrial protein distribution of Mic60 following endurance exercises in mice, but massive alterations of MICOS components as well as other mitochondrial proteins occurring during apoptosis. In this dissertation I show that the examined stimuli evoke different changes to the submitochondrial protein distribution of the MICOS components.||de