Regulatory principles of human mitochondrial gene expression revealed by kinetic analysis of the RNA life cycle

Mitochondria play a critical role in cellular metabolism primarily through hosting the oxidative phosphorylation (OXPHOS) machinery that is encoded by mitochondrial DNA (mtDNA) and nuclear DNA, with each genome separately regulated in their respective compartments. To unravel how the two gene expression systems collaborate to produce the OXPHOS complexes, the regulatory principles controlling the production of mtDNA-encoded proteins need to be elucidated. Here, we performed a quantitative analysis of the mitochondrial messenger RNA (mt-mRNA) life cycle to gain insight into which steps of gene expression experience the most regulatory control. Our analysis revealed a unique balance between the rapid turnover and high accumulation of mt-mRNA, leading to a 700-fold higher transcriptional output than nuclear-encoded OXPHOS genes. Additionally, we observed that mt-mRNA processing and its association with the mitochondrial ribosome occur rapidly and that these processes are linked mechanistically. These data resulted in a model of mtDNA expression that is predictive across human cell lines, revealing that differential turnover and translation efficiencies are the major contributors to mitochondrial-encoded protein synthesis. Applying this framework to a disease model of Leigh syndrome, French-Canadian type, we found that disrupting the responsible nuclear-encoded gene, LRPPRC, perturbs OXPHOS biogenesis predominantly through altering mt-mRNA stability. Our findings provide a comprehensive view of the intricate regulatory mechanisms governing mtDNA-encoded protein synthesis, highlighting the importance of quantitatively analyzing the mitochondrial RNA life cycle for decoding the regulatory principles of mtDNA expression. ### Competing Interest Statement The authors have declared no competing interest.