Regulation of mtDNA gene expression and the role of respiratory chain supercomplexes in mammalian mitochondria

Abstract

Mitochondria are cellular organelles found in nearly all eukaryotic cells, where they fulfill a plethora of functions, including energy conversion. They harness energy from carbon sources to synthesize adenosine triphosphate (ATP) through the process of oxidative phosphorylation (OXPHOS). Mammalian mitochondria contain ~1200 proteins and only 13 of these are encoded by mitochondrial DNA (mtDNA). Despite the small contribution to the mitochondrial proteome, expression of mtDNA is nevertheless critically important for biogenesis and normal function of the OXPHOS system. The expression of mtDNA is dependent on ~200 nucleus-encoded proteins that are imported into mitochondria to control maintenance, replication, and transcription of mtDNA, as well as translation of the mtDNA-encoded messenger RNAs (mRNAs). Mitochondrial gene expression is thus controlled at many different levels. In the first part of this thesis, molecular mechanisms are reported for initiation and completion of mtDNA replication. Furthermore, mechanisms controlling the switch from transcription initiation to elongation, as well as a putative link between transcription elongation and RNA processing are reported. These results were based on extensive analysis of conditional knockout mouse models for RNase H1, TEFM, and mtSSB. In the second part of the thesis, the organization of the OXPHOS system is studied. It consists of the respiratory chain enzyme complexes, two electron shuttles and the ATP synthase. A wide range of studies have shown that the different respiratory chain enzyme complexes can interact with each other to form higher order assemblies, so called supercomplexes. The respirasome is a particularly interesting supercomplex as it consists of a complete respiratory chain. Results from multiple studies have suggested that the respirasome plays a crucial role in cellular bioenergetics by facilitating translocation and routing of electrons. To address this question, we extensively characterized a knockin mouse model engineered to have normal levels of respiratory chain complexes that cannot interact to form respirasomes. Surprisingly, mice with drastically reduced levels of respirasomes are healthy without any clear impact on bioenergetics or whole animal physiology. Our results thus challenge the multiple proposed roles for respirasomes in physiology and disease.