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Illuminating the mechanisms of mitochondrial DNA quality control and inheritance

Periodic Reporting for period 2 - IlluMitoDNA (Illuminating the mechanisms of mitochondrial DNA quality control and inheritance)

Reporting period: 2019-02-01 to 2020-07-31

Mitochondria are known as the power plants of our cells, because they provide our cells with a large amount of energy through a process known as oxidative phosphorylation (OXPHOS). Several crucial proteins required for OXPHOS are encoded on the mitochondrial genome (mtDNA), which is present in multiple copies in cells. Cells must therefore ensure maintenance of functional mtDNA to secure their efficient energy supply and health. Accordingly, mutations in mtDNA can have dire consequences and can lead to mitochondrial diseases and ageing.

Quite surprisingly, fundamental questions regarding the biology of mtDNA are currently poorly understood: How is mtDNA faithfully passed on to daughter cells during cell division? How are mtDNA copies distributed throughout the mitochondria, that form a tubular and reticulated network in cells? How is the integrity of mtDNA maintained over generations, despite an elevated level of reactive oxygen species within mitochondria? It is these mysteries that we aim to unravel in this project, down to a deep mechanistic understanding. The answers to these question also hold the key for a better understanding of human mitochondrial disorders and will pave the way for the development of new therapeutical approaches. To study mitochondria and mtDNA, we combine unique experimental advantages in the model organism S. cerevisiae with a wide variety of cell biological techniques, including fluorescent live-cell and super-resolution microscopy, next-generation sequencing, protein biochemistry and yeast genetics.
We know very little about how cells faithfully maintain mtDNA over generations. A central aspect to this process is the regulation of the correct number of mtDNA copies within the cell. We have performed a systematic analysis of ~5000 mutant yeast strains to identify genes required for maintenance of normal cellular mtDNA levels. This screen revealed several mutants that displayed elevated levels of mtDNA, including a strain with a deletion of the previously uncharacterized gene MRX6. Cells lacking MRX6 display normal mitochondrial morphology, but contain clusters of mtDNA copies, suggesting that segregation of replicated mtDNA copies is defective in such cells. We found that the Mrx6 protein physically interacts with the evolutionary conserved protease Pim1, a central player in mitochondrial protein quality control. Similar to cell lacking MRX6, depletion of Pim1 resulted in elevated cellular mtDNA levels, which were not further elevated by additional absence of Mrx6. These results support the model that Mrx6 regulates mtDNA levels through modulation of Pim1 mediated protein degradation. It will be an exciting task for future studies to elucidate the detailed mechanism that underlie regulation of mtDNA levels through the Mrx6-Pim1 complex.
Besides the regulation of mtDNA levels, it is of utmost importance to understand how cells coordinate mitochondrial dynamics with faithful maintenance of mtDNA and its transmission to daughter cells during the cell cycle. Our current, yet unpublished, efforts focus on this question. Furthermore, we examine how cells prevent the accumulation of mutated mtDNA. It is our goal to answer the following questions: Are cells able to detect mutant mitochondrial genomes and if yes, what are the signals and what are the molecular mechanisms that underlie this process?