| Description |
Mitochondrial morphology maintenance is a highly dynamic process regulated by multiple different pathways. The two main pathways that regulate dynamics are fission and fusion. In this thesis, I describe studies designed to further our understanding of both pathways. Fission requires the coordinated interaction of three proteins, Fis1p, Mdv1p, and Dnm1p. Fis1p is a tail-anchored, outer mitochondrial membrane protein. Mdv1p and Dnm1p are soluble, cytoplasmic proteins. The mechanism by which these three molecules assemble on the outer mitochondrial membrane to stimulate fission remains unclear even though multiple models have been proposed. In this thesis, I show that Fis1p and Mdv1p interact directly and it is this interaction that forms a platform for Dnm1p assembly into higher-ordered structures to stimulate fission. Related experiments also describe the molecular details of the Fis1p-Mdv1p interaction at an unprecedented resolution. In addition, I describe preliminary experiments required for future structural and biochemical studies to further define the interactions of the fission complex. Taken together, these studies have revised and refined existing models of mitochondrial fission. The mitochondrial fusion pathway also requires three proteins, Fzo1p, Mgm1p, and Ugo1p. Fzo1p and Mgm1p are both GTPases, with Fzo1p embedded in the outer mitochondrial membrane and Mgm1p found in the intermembrane space. Ugo1p is an integral, outer membrane protein with a large domain facing the cytoplasm and a slightly smaller domain localized to the intermembrane space. These three molecules form a complex, but at this point, little is known about the role that each protein plays in fusion. In this thesis I explore the role of Ugo1p in fusion. In addition to the transmembrane domain, the 502 amino acid Ugo1 protein contains only two recognizable motifs. These small motifs are characteristic of mitochondrial carrier domains found in mitochondrial carrier proteins. In this thesis, I test whether these putative carrier domains (PCDs) are required for Ugo1p function and find that while one of them is dispensable, the second one is required for efficient fusion. In addition, this thesis describes studies that begin to uncover the mechanism of Ugo1p targeting to the mitochondrial outer membrane. These studies show that the transmembrane domain of Ugo1p is not required for targeting, but that a 50 amino acid region at the C-terminus of the protein can be targeted efficiently to mitochondria. |
