In vivo analysis of the mechanisms of axon transport.


Principal Investigator: Catherine M Drerup
Abstract: DESCRIPTION (provided by applicant): In neurons, axonal transport of proteins and organelles to and from synapses is essential for formation and maintenance of neural connectivity. Impaired axon transport is thought to contribute to numerous neurodevelopmental and neurodegenerative disorders, including Alzheimer's Disease, Amyotrophic Lateral Sclerosis, and Charcot-Marie Tooth Disease. Despite the pervasiveness of these disorders, their underlying causes are still poorly understood, which has hindered the development of effective therapies This is at least partially due to the lack of a vertebrate model system in which to study this process in vivo and test potential therapeutics. I have developed zebrafish as an in vivo model for studying axon transport by 1) developing a novel imaging approach to visualize movement of fluorescently labeled cargo in an intact animal;and 2) participating in a forward genetic screen to isolate mutants with axonal transport defects. One of these mutant strains (rogue) has a phenotype typical of disruptions in axon transport, i.e. nerve truncation, nerve thinning and distal axonal swellings in long sensory axons. Live imaging revealed that rogue has reduced density and altered transport parameters of some actively transported cargos. Positional cloning identified the underlying genetic lesion in the gene encoding jnk interacting protein 3 (jip3). Previous studies in vitro revealed that Jip3 binds both the microtubule motor Kinesin-1 and axonal cargo. Jip3 has also been shown to modulate cJun N- terminal kinase (Jnk) activity in cell culture, which could potentially have downstream effects on the microtubule cytoskeleton and cargo-motor binding. However, which axonal cargos, if any, are directly Jip3- dependent and which of these Jip3-dependent molecular interactions regulate axonal transport during axon extension and synapse formation is not known. To address these questions, I will first use the live embryo imaging approach I developed to determine if microtubule dynamics and axon transport of specific cargos are disrupted in rogue. Second, I will determine if Jip3 interaction with Jnk and/or Kinesin-1 are necessary for proper regulation of the microtubule cytoskeleton or axonal transport of specific cargos, thus promoting axon extension and synapse formation. The proposed experiments will define the Jip3-dependent cellular and molecular processes which mediate axon transport in vivo. Long-term, the system I have developed can be used to analyze axon transport in vivo to fully understand how abnormalities in this process disrupt nervous system formation and function in normal and disease states. PUBLIC HEALTH RELEVANCE: The transport of proteins and organelles from the neuronal cell body to axon terminals and vice versa is critical both to maintain the health of the cell body and support the formation of functional nervous system connections. Defects in this process are associated with numerous developmental and neurodegenerative diseases such as Spinal Muscular Atrophy, Alzheimer's Disease, and Amyotrophic Lateral Sclerosis. The knowledge gained from these studies will advance our understanding of the basic mechanisms required for axonal transport in vivo. Additionally, they will establish zebrafish as a model system that can be used to investigate the function of genes associated with axonal diseases to determine if disease etiology lies in interruptions of this basic cellular process.
Funding Period: 2011-05-01 - 2012-04-30
more information: NIH RePORT