Date

April 2009

Document Type

Dissertation

Degree Name

Ph.D.

Institution

Oregon Health & Science University

Abstract

This dissertation presents studies concerning two specific aspects of protein trafficking in neurons. First, live-cell imaging of GFP-tagged dendritic proteins in cultured hippocampal neurons was used to study the selective transport of dendritic proteins, which are specifically blocked from entering the axon. This selective transport is likely the primary mechanism by which dendritic proteins are restricted to the somatodendritic domain, and is therefore critical to neuronal function. The block of dendritic transport into the axon was found to have two components; selective entry, which reduced the probability that a dendritic vesicle would enter the axon, and the transport filter, which blocked dendritic vesicles that did enter the proximal axon from advancing more than a few microns into the axon. In mature neurons the transport filter aligned with the cell-body proximal edge of the initial segment, a specialized domain of the axon that forms the boundary between axonal and somatodendritic domains. However, the transport filter was detected in the axons of newly formed axons of stage 3 cells, before the initial segment developed. Furthermore, selective dendritic transport was detected in neurites of stage 2 neurites, at which time it was dynamically regulated. Second, live-cell imaging was used to study the transport of two types of axonal proteins—synaptic vesicle proteins and dense core granule proteins—both of which must be transported into the far reaches of the axon to support synaptic and trophic signaling. A Kinesin-3 family member, KIF1A, has been implicated in the transport of both of these proteins in the axon. Two-color live-cell imaging showed that upon Golgi departure synaptic vesicle and dense core granule proteins, as well as axonal plasma membrane proteins are initially transported in a common transport tubule. KIF1A did not associate with this transport tubule but did label spherical dense core granule-containing organelles at later time points, suggesting that more specialized vesicle populations that arise from the common tubule are moved by KIF1A. These studies have revealed important new aspects of dendritic and axonal transport, which will be useful in future studies that seek to understand the regulation of protein trafficking in neurons.

Identifier

doi:10.6083/M40V89SH

Division

Neuroscience Graduate Program

School

School of Medicine

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