Function and regulation of FEZ1, a Kinesin-1 adaptor protein, in transporting presynaptic proteins

• Applicant: Dr. John Jia En Chua
• Subject Area: Molecular Biology and Physiology of Neurons and Glial Cells
• Term: from 2013 to 2018
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 249372430

Axonal transport delivers proteins and membranes to presynaptic nerve terminals and is thus essential for formation and maintenance of active synapses. Illustrating its importance, mutations affecting components of axonal transport were identified in neurodegenerative disorders. The Kinesin adaptor FEZ1 appears to transport a subset of presynaptic cargo but neither the range of cargoes nor mechanistic details and regulation of FEZ1-dependent transport are known. Here I plan to shed more light on the role of FEZ1 in the synaptic delivery of protein complexes or vesicles required for presynaptic function. Key objectives of the research proposal are: (1) Which kinases and signaling pathways regulate FEZ1-mediated transport?We observed that phosphorylation of serine-58 on FEZ1 is needed for activating axonal transport of the presynaptic SNARE protein Syntaxin-1. Using in vitro kinase assays, we identified several FEZ1 kinases phosphorylating serine-58. Here I plan to further characterize the role of these kinases in regulating FEZ-1-dependent axonal transport using cultured mammalian neurons and Caenorhabditis elegans as model systems. I will also investigate whether there are additional signaling kinases or kinase cascades involved in the regulation of FEZ1.(2) Which cargoes are transported by FEZ1/Kinesin-1 complexes and how does FEZ1 dysfunction affect synaptic development and function?Various cargo adapters have been identified for a selected range of cargoes. In particular, 2 synaptic cargoes have been studied in detail, namely, synaptic vesicle protein transport vesicles (STVs) and Piccolo-Bassoon transport vesicles (PTVs). I plan to determine the composition of cargoes transported by FEZ1/Kinesin-1 complexes using immunoisolation-quantitative mass spectrometry. By comparing this against other axonal transport vesicles, I hope to clarify the relationships between FEZ1-vesicles, STVs and PTVs and assess the contribution of FEZ1 to presynaptic development and function.(3) To what extent do FEZ1 defects lead to synaptic malfunction and neurodegeneration?Impaired axonal transport occurs in Alzheimers disease (AD), a neurodegenerative disorder associated with synaptic loss, and likely contributes to AD pathogenesis. Impairing Kinesin-1 activity by reducing kinesin light chain 1 levels suffices to induce transport defects observed in AD. As FEZ1 binding activates Kinesin-1, FEZ1 malfunction may cause similar defects. Preliminary data reveal that FEZ1 is abnormally aggregating in model transgenic AD mice. Using this as a starting point, I plan to further investigate the involvement of FEZ1 in synaptic malfunction and neurodegeneration in relation to its function in synaptic cargo delivery. I expect that the data generated in this project will contribute to a better understanding of how axonal transport and its regulation participates in synaptogenesis and synapse maintenance.

• DFG Programme: Research Grants