MITOCHONDRIAL DYNAMICS--MORPHOLOGY/DIVISION/SEGREGATION

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Mitochondria are dynamic organelles found in virtually all eukaryotic cells and show striking variations in their location, number and structure in different cell types. Mitochondrial abnormalities are also associated with a variety of human diseases. Virtually nothing is known about the molecular mechanisms that mediate mitochondrial dynamics. In the yeast Saccharomyces cerevisiae, mitochondria are usually elongated organelles which form a reticulum around the cell periphery. Yeast dramatically regulates the shape, size and number of its mitochondria according to the physiological state of the cell. We propose four approaches to identify and analyze the molecules that mediate mitochondrial morphology, mitochondrial number and mitochondrial location. WHAT IS THE FUNCTION OF THE MMM1 PROTEIN IN MEDIATING MITOCHONDRIAL SHAPE AND MITOCHONDRIAL SEGREGATION? mmm1 mutants are temperature-sensitive for the external shape of their mitochondria. At the restrictive temperature, elongated mitochondria appear to quickly collapse into large, spherical organelles. The Mmm1 protein is located in the mitochondrial outer membrane, with a large carboxyl-terminal domain facing the cytosol. MMM1 is a new gene and our results raise the possibility that the Mmm1 protein maintains mitochondria in an elongated conformation by mediating the binding of the organelle to the cytoskeleton or some other external framework. mmm1 mutants are temperature-sensitive for viability, and the lethality appears to result from the inability to segregate the aberrant- shaped mitochondria into daughter cells. Hence mitochondrial shape is essential for the normal segregation of mitochondria to daughter cells during cell division. To determine the role of Mmm1p, we propose to: (1) examine single yeast cells defective in Mmm1p function using time-lapse fluorescence imaging, (2) determine the topology of Mmm1p in the mitochondrial outer membrane, (3) ask if Mmm1p is located in contact sites, (4) determine if Mmm1p is part of a multisubunit complex, and (5) ask whether the binding of mitochondria to cytoskeletal elements, such as actin, or tubulin, are disrupted in mmm1 mutants. WHAT IS THE ROLE OF THE BOM1P, A POTENTIAL MMM1P-INTERACTING PROTEIN? Bom1p was identified as a molecular genetic screen for proteins that potentially interact with Mmm1p. We have shown that the Bom1 protein is essential for yeast cell viability, and that Bom1p is located in the mitochondrial inner membrane. Furthermore, when Bom1p is depleted from yeast cells, elongated mitochondria collapse into sphere-shaped organelles similar to those seen in mmm1 mutants. We propose to (1) isolate temperature-sensitive bom1 mutants to determine the role of the Bom1 protein, and (2) test whether Bom1p directly interacts with Mmm1p in the yeast cell. WHAT ROLE DOES A YEAST HOMOLOGUE OF BACTERIAL FTSZ PLAY IN MITOCHONDRIAL DIVISION? We have recently identified in yeast mitochondria a cognate of ftsZ, a protein required for E. coli cell division. Since mitochondria are thought to have arisen from bacteria, we will test whether the yeast ftsZ-related protein plays a role in mitochondrial division. CHARACTERIZE ADDITIONAL YEAST MUTANTS DEFECTIVE IN MITOCHONDRIAL DYNAMICS. In addition to mmm1, we identified 23 mutants defective in normal mitochondrial morphology, mitochondrial number, and mitochondrial distribution. We propose to further analyze these mutants to determine their role in mitochondrial dynamics.