The goal of the proposed research is to define the mechanisms of translational activation within mitochondria. Alterations in mitochondrial gene expression can compromise cellular energy production and generate reactive oxygen species that promote degenerative disease, aging, and cancer. Thus, a clearer understanding of how this system is regulated is necessary to better understand mitochondrial disease. To explore this process further, the first aim will elucidate the mechanism by which the newly discovered yeast translational activator Mam33 activates the translation of COX1, a core subunit of the cytochrome c oxidase complex. These experiments will determine 1) the region of COX1 mRNA sufficient for Mam33-dependent activation, 2) whether Mam33 directly binds the COX1 transcript, 3) if Mam33 associates with the mitochondrial ribosome, and 4) if Mam33 is rate-limiting. This information will advance our understanding of Mam33 and mitochondrial activators in general, which can be later tested with human homologues.
The second aim will employ biochemical and genetic strategies to identify and characterize factors that physically interact with - or are functionally related to Mam33. These results will allow us to place Mam33 within the COX1 expression pathway and better define its translation activation mechanism. Additionally, unanticipated Mam33 activities may be revealed.
The third aim i s to understand the structural requirements for Mam33 activity. A mutational analysis based upon crystal structure features and amino acid conservation found in homologs from other organisms will be performed. Since both Cox1 and Mam33 are evolutionarily conserved, information gained in yeast will likely be applicable to human mitochondrial disorders. Furthermore, recent data also implicates the human homologue p32 in cancer. Thus, a better understanding of yeast Mam33 could also enhance our understanding of cancer.
This proposed research seeks to better understand the causes of mitochondrial disorders; a group of diseases affecting about 1 in 4,000 children by the age of 10 years. The yeast model organism will be used to study a feature of mitochondrial genome expression conserved in humans. This information is necessary for discovering the underlying causes of mitochondrial disorders and may help identify potential therapies that could prevent, or even reverse, mitochondrial diseases.
