Nearly every aspect of neuronal function depends on the polarized trafficking of membrane proteins to either the axonal or somatodendritic domains. Live-cell imaging of cultured neurons shows that selective microtubule- based transport plays a key role in ensuring the accurate delivery of membrane proteins to these domains: vesicles containing dendritic proteins are transported exclusively within the somatodendritic domain of the cell; vesicles that contain axonal proteins move in dendrites, but their transport is biased toward the axon so that their protein cargoes accumulate there. We hypothesize that different sets of motor proteins associate with these different vesicle populations and that the distinctive properties of these motor proteins and their regulators determine transport selectivity. We propose to evaluate this hypothesis using novel methods we have developed to identify the motor proteins that associate with specific vesicle populations and the adaptors that link motors to these vesicles. Axon-selective transport may be explained by the properties of the kinesin motors that associate with vesicles containing axonal proteins. To evaluate this hypothesis, we will identify the kinesins that bind axonal vesicles and determine whether mutations in these kinesins disrupt axon-selective transport. Our previous results demonstrate that dendrite-selective transport cannot be explained solely by kinesin translocation preferences. Instead, selective transport likely depends on regulation mediated by kinesin adaptors specific to dendritic vesicles or on the interplay between kinesins and other molecular motors associated with these vesicles. To evaluate these possibilities, we will determine whether the sets of kinesin adaptors and myosins and dyneins differ between axonal and dendritic vesicles. We believe the novel experimental strategy proposed will provide a definitive identification of the motors and motor adaptors associated with these important neuronal vesicle populations and enable significant strides toward elucidating the mechanisms that underlie transport selectivity.
Nearly every aspect of neuronal function depends on the accurate transport of membrane proteins to axons or dendrites; defects in long-range transport are thought to underlie the axonal degeneration characteristic of many neurodegenerative diseases. By elucidating the regulatory mechanisms that underlie the accuracy of microtubule-based transport, our work may identify a novel set of targets for pharmacologic manipulations that could enhance long-range transport and perhaps ameliorate neural degeneration.
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