Over 100 genetic diseases are known to affect mitochondrial activity making this one of the most common classes of human pathologies. Mitochondrial function requires the coordinated expression of ~1500 nuclear-encoded genes and 37 mtDNA-encoded genes. This genomic organization has two important implications: 1) mitochondrial function is a large mutational target, and 2) nuclear-mitochondrial interactions are central to normal physiological performance. The long-term goal of this research program is to use Drosophila as a model to dissect the genetic bases of nuclear-mitochondrial interactions that affect organismal fitness. The initial phase of this research has uncovered a striking epistatic interaction where a particular mtDNA in one nuclear background (Oregon R) produces a suite of highly compromised phenotypes. When this same mtDNA is placed on an alternative set of wild chromosomes, these phenotypes are restored to near-wild type levels. The traits affected are hallmarks of metabolic disease: delayed development, reduced fecundity, locomotion, COX activity, and bristle defects. The current proposal seeks to identify the locus or loci responsible for this nuclear-mitochondrial interaction, and dissect the function of these genes. There are three specific aims: 1) Genetically map nuclear factors that modify mtDNA performance using segregation, meiotic and deficiency mapping. We will test the hypothesis that all affected phenotypes are due to a single locus, vs. independent loci for each trait;2) Molecular characterization of the mapped nuclear loci using SNP-array mapping among F2 offspring, and analyses of existing and novel alleles to identify the nucleotide changes causing specific phenotypes and, 3) Tissue specific expression of the identified genes. We will document the tissue specific expression of the newly identified genes, and their impact on mitochondrial gene expression. We will manipulate tissue specific expression to test the hypothesis that this nuclear-mitochondrial interaction effect on phenotype is distinct in different tissues. These Aims will identify the genetic bases of strong mito-nuclear epistatic interactions uncovered in the first phase of the research, and undertake new mechanistic studies of how the mitochondrial enzyme complexes function under specific genetic manipulations of nuclear and mtDNA encoded proteins. These experiments will provide fundamental information on how specific mutations in nuclear and mitochondrial genes interact to determine organismal performance. As these interactions are evolutionarily ancient and highly conserved, the dissection of genetic pathways in Drosophila will be very relevant to understanding this very common class of diseases in humans.