If cells lose the capacity to produce or to maintain mitochondria, they die. Movement disorders, blindness, deafness, seizures, mental retardation, stunted growth and impaired neural development are all linked to mitochondrial myopathies, diseases associated with mtDNA mutations and resulting respiratory defects. This proposal focuses on one aspect of cytokinesis: the process whereby mitochondria are transferred from mother to daughter during cell division. Mitochondria can arise only from pre-existing mitochondria. Therefore, mitochondrial inheritance is an integral part of the cell division that insures that each cell contains this indispensable organelle. Early studies indicate that yeast mitochondria segregate during mitotic cell division. Thus, mitochondrial inheritance is a tightly regulated process whereby specific populations of mitochondria may be targeted for transfer into daughter cells. Our studies indicate that mitochondria are associated with the yeast actin cytoskeleton, and that these organelle-cytoskeletal interactions control mitochondrial motility and inheritance. In addition, we identified a novel myosin I protein and actin-dependent motor activity on isolated yeast mitochondria. We propose that mitochondrial segregation is produced by 1) polarized, myosin I-driven movement of mitochondria into daughter cells using actin cables as tracks, 2) preferential inheritance of organelles proximal to the developing daughter cell, and 3) down-regulation of motor activity which results in retention of some mitochondria in mother cells and retention of all newly inherited organelles in daughters. The experiments described are designed to test this model. We will examine the role of calmodulin and calcium, known regulators of myosin, on mitochondrial motor activity in vitro and in vivo. In addition, if mitochondrial motility is regulated at the level of the motor, then the organelle motor must be tightly associated with mitochondria. We obtained the first experimental evidence for a myosin receptor protein on a target membrane, and propose to identify this receptor protein as well as motor sub- domains required for receptor binding. Finally, we will examine the effect of mutations perturbing the actin cytoskeleton, mitochondrial motor function, and cell polarity determinants on mitochondrial segregation. Given the fundamental roles of cytokinesis in cell division, cell polarization in development and cell migration, and mitochondrial defects in human disease, it is surprising how little is known regarding organelle-cytoskeletal interactions which control mitochondrial inheritance. The projects proposed are designed to fill that gap.
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