Protein: DNA Interactions in Drosophila Mitchondrial Regulation

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9600681 Kaguni The long term objective of the proposed work is to understand the regulation of mitochondrial function, in terms of the structure of the mitochondrial chromosome and its expression and replication. Mitochondria are required in higher animal cells, and mitochondrial biogenesis requires expression and duplication of the mitochondrial DNA (mtDNA) genome. Because both transcription and replication are influenced by the structure of the DNA template, the role of nucleoprotein complexes that are involved in maintaining DNA conformation in vivo on these functions warrants detailed study. To begin to elucidate the role of proteins associated with the mitochondrial chromosome in maintaining its structure and to examine their influence on its function, we will isolate the mitochondrial nucleoid from Drosophila melanogaster, purify and characterize its protein components, and initiate efforts to clone them. Our long-term goal will be to develop in vitro transcription and replication systems from Drosophila mitochondria that employ mitochondrial enzyme fractions and mitochondrial DNA in the form of nucleoprotein complexes. The isolation and characterization of the mitochondrial nucleoid is an integral part in the development of the Drosophila model for animal mtDNA function. %%% Mitochondria are the energy-producing units in the cell and are essential to life. Because of the limited size of the mitochondrial chromosome, all of the proteins required for mitochondrial biogenesis are encoded outside the mitochondrion in the nucleus. These proteins are comparatively rare in the animal cell, and are therefore difficult to study. The fruit fly Drosophila is a higher animal that has proven to be an excellent model for the study of cellular function by biochemical, genetic and developmental approaches. The structure and organization of mitochondrial DNA (mtDNA) is highly conserved in animals, and current data suggest that the mechanisms of mtDNA packaging and gene expression will also be. Our lo ng-term goal is to develop Drosophila as an animal model system to study these processes. The isolation and characterization of the mitochondrial chromosome and its protein components represents the first step in the development of the Drosophila model for animal mtDNA function. ***