ED Sciences de la Vie et de la Santé
Yeast modeling and functional consequences of mitochondrial DNA mutations associated with pathologies
by Camille CHARLES (Institut de Biochimie et Génétique Cellulaires)
The defense will take place at 14h00 - Salle de conférence de l'IBGC Institut de Biochimie et Génétique Cellulaires (IBGC) - UMR5095 CNRS 1, Rue Camille Saint Saëns, CS61390 - 33077 Bordeaux cedex, France
in front of the jury composed of
- Nathalie BONNEFOY - Directrice de recherche - Université Paris-Saclay, Institute for Integrative Biology of the Cell (I2BC) - Rapporteur
- Vincent PROCACCIO - Professeur des universités - Université d'Angers, Institut MitoVasc - Rapporteur
- Véronique PAQUIS-FLUCKLINGER - Professeure des universités - praticienne hospitalière - Université de Nice, CHU de Nice - Examinateur
- Frédéric BOUILLAUD - Directeur de recherche - Université Paris Cité, Institut Cochin - Examinateur
- Déborah TRIBOUILLARD-TANVIER - Directrice de recherche - Institut de Biochimie et Génétique Cellulaires (IBGC), INSERM - Directeur de these
Adenosine Triphosphate (ATP), a small energy-rich molecule, is mainly produced in the mitochondria in non-photosynthesizing eukaryotes though the oxidative phosphorylation process. This highly complex process depends on numerous systems, including the ATP synthase complex. This enzyme exploits a proton gradient across the inner mitochondrial membrane to synthesize ATP from ADP and inorganic phosphate. ATP synthase is composed of two functional domains, Fo and F1. The Fo carries protons across the inner membrane and generates mechanical energy that promotes ATP synthesis in the F1 through conformational changes. The chemical energy stored in ATP is made available to cellular reactions as needed through hydrolysis of a phosphate group. Mitochondrial ATP synthase stands out due to its mixed genetic origin, a very rare characteristic in the eukaryotic world (only 4 to 5 enzymatic systems share this trait). The structural genes encoding its different protein subunits (numbering about twenty) are indeed partially located in the nucleus, while others are carried by a small DNA molecule present in the mitochondria. Subunits with a nuclear origin are synthesized in the cytosol and then imported into the organelle while those encoded by mitochondrial DNA are translated within the mitochondria itself. Due to this genetic compartmentalization, the biogenesis of ATP synthase is an extremely complex process involving hundreds of factors required for the synthesis of its components and their oligomerization. Genetic defects affecting the formation or function of ATP synthase are responsible for human diseases that mainly affect organs and tissues with high-energy demands, such as the brain, heart and skeletal muscles. These diseases still await truly curative treatments despite numerous pharmacological trials using various compounds, such as vitamins, cofactors, or metabolic intermediates. Given these challenges, the laboratory where I conducted my PhD developed an approach based on modeling ATP synthase-related disease mutations in the yeast Saccharomyces cerevisiae. Thanks to its exceptional fermentative ability, it can survive ATP synthase inactivation, and it is possible to modify its ATP synthase subunits at will, both those encoded in the nucleus and those of mitochondrial origin (it is actually impossible to insert specific mutations into human mitochondrial DNA). It is worth noting that pathogenic mutations in mitochondrial DNA often affect only a fraction of the numerous copies of this DNA (several thousand per cell), making it difficult to study their functional consequences in patient-derived cells. Yeast overcomes this obstacle due to its inability to stably maintain different mitochondrial genotypes within the same cell. Taking advantage of the properties of the yeast system, I modeled about ten mutations found in the mitochondrial ATP6 gene detected in patients suffering from various neuromuscular disorders. My studies led us to conclude that some of these mutations are merely neutral polymorphisms with no significant impact on mitochondrial function, whereas others exhibited particularly deleterious effects, leaving no doubt about their pathogenic nature. Unexpectedly, the analyses revealed a new mechanism that contributes to coordinating the expression of nuclear and mitochondrial subunits of ATP synthase to prevent the accumulation of potentially harmful assembly intermediates.