It is not known if or how mammalian cells induce selective translation of mitochondrial proteins and if this differs by cell/tissue type. The long-trm goal of this research is to identify strategies that can be used to increase mitochondrial protein synthesis in humans for the purpose of increasing healthspan. The overall objective is to identify post-transcriptional mechanisms that are involved in increasing mitochondrial protein synthesis during periods of energetic stress in mammalian cells/tissues. The central hypothesis is that the selective translation of mitochondrial proteins observed in liver, skeletal muscle, and heart of mammals during periods of energetic stress is dictated by the complexity of mRNA sequence elements and/or subcellular localization to the mitochondrial reticulum. This hypothesis has been formulated based on evidence generated in the applicants' laboratories as well as extensive published evidence in lower eukaryotes. The rationale for the proposed research is that the successful completion of this research would provide new opportunities to identify therapeutic targets to slow aging and prolong healthspan by targeting mechanisms that have been virtually unstudied in mammalian cells. The research has two specific aims: 1) to determine in vitro the mechanisms responsible for the preferential translation and synthesis of mammalian mitochondrial proteins during the signaling of energetic stress, and the extent of the contribution of autophagy/mitophagy to these increases, and 2) to identify strategies that activate pathways of energetic stress in vivo to preserve mitochondrial protein synthesis and slow aging. To accomplish these specific aims, multiple cell types and tissues will be analyzed. Methods used in vitro include polysome profiles with analysis of rapidly translating mRNA, bioinformatic analysis of mRNA sequences, and novel stable isotope techniques for the determination of mitochondrial protein synthesis and autophagic flux. Methods used in vivo include additional novel isotope techniques for measuring protein synthesis at the tissue, organelle, and individual protein level as well as cellular proliferation. The contribution of the proposed research is significant because it is designed to elucidate an important, but unstudied, regulatory step of mitochondrial biogenesis. Further, it is designed to translate in vitro mechanisms to in vivo studies to determine if these mechanisms can be targeted therapeutically. The proposed research is innovative because it considers post-transcriptional mechanisms of mitochondrial biogenesis, which have previously been largely overlooked. Further, the laboratory is among the very few groups capable of making long-term protein synthetic and cellular proliferation measurements that allow for the translation of in vitro studie to in vivo outcomes. Finally, the in vivo assessment of the synthesis of individual mitochondrial proteins has until now not been feasible, thus the project is technically innovative. The results are expected to have a positive impact because they focus on mechanisms that may offer novel therapeutic targets that are alternatives to a therapy (lifelong caloric restriction) that has provn efficacy, but extremely limited adherence in humans.
The proposed research is relevant to public health because it supports the National Institute of Aging mission of addressing mechanisms of aging and then develops interventions to target them therapeutically. The project is designed to translate in vitr mechanisms into possible in vivo treatments using pharmacologic approaches that are already in human use for other age-related diseases. Although the current proposal is focused on aging, it is likely that information gained will also be relevant to chronic diseases (e.g. diabetes and heart disease) in which mitochondrial dysfunction is thought to be causative.
