Work from this lab and others has established that mitochondrial motility is governed by a motor/adaptor complex that couples both kinesin and dynein motors to the mitochondrial surface. Much, however, remains unknown about how regulation of that complex determines when and where a mitochondrion will stop, or why some mitochondria are long-term stationary, while others only pause briefly before continuing to move. Anchoring proteins are known that can hold mitochondria in place, but the interplay of anchoring proteins and mitochondrial motors is poorly understood. This proposal seeks to understand mechanisms that regulate the assembly of the motor/adaptor complex and signaling pathways that can turn off those motors. The proposal also seeks to understand when mitochondria are merely passively stationary and when they are being actively held in place by an anchor. The exceptional shapes of neurons makes mitochondrial motility and its regulation particularly important so that the branches of axons and dendrites can be properly supplied with mitochondria and certain locations with high energetic demand, such as synapses, can have increased density of mitochondria. This proposal therefore places special emphasis on how mitochondrial dynamics are regulated in axons and on signals that may localize mitochondria to synapses. To better understand mitochondrial regulation, the proposal introduces two novel approaches. One approach is the misdirection of the mitochondrial motor/adaptor complex to peroxisomes so that assembly and regulation of the complex can be studied and mutated away from the endogenous mitochondrial proteins. The other new approach is to use a heterodimerizing agent to attach a constitutively active kinesin motor to mitochondria and thereby test whether mitochondria are being actively held in place by an anchor. In addition, this proposal introduces two new factors that govern the motility of mitochondria, a protein that anchors mitochondria to the actin cytoskeleton, and a kinase that regulates the localization of mitochondria at synapses. In Aim 1 we propose to study how a GTPase domain of Miro regulates the ability of the complex to assemble and what physiological significance there is to the GTPase activity. In Aim 2 we investigate competing models for how elevated Ca2+and the PINK1/Parkin pathway cause mitochondria to stop and ask whether or not they immobilize mitochondria with anchoring proteins. In Aim 3 we investigate the mechanism by which Aurora kinase B inhibits mitochondrial movement and thereby promotes the localization of mitochondria to synapses.
NIH ExPorter
NIH ExPorter