Pairing Modeling and Experiment to Understand Microtubule Behavior in Healthy and Injured Neurons

Project Summary Neurons rely on polarity and stability of the microtubule cytoskeleton to support long-range directed transport and long-term survival. However, both polarity and stability can be rapidly altered in response to injury and these rearrangements are critical for neuronal resilience. It is becoming clear that rather than a single master mechanism controlling neuronal microtubule organization through time and space, multiple mechanisms operate in parallel. This complexity makes it challenging to understand how each mechanism contributes to filament organization and how the system works as a whole. To overcome this challenge, a mathematical framework that incorporates known mechanisms will be built. This framework will be invaluable for understanding the dendrite microtubule system, and how it responds to perturbations induced by injury. Aim 1. Polarized organization of microtubules in neurons is critical for correct cargo delivery to axons and dendrites. A spatial stochastic model of the polarized array of dendritic microtubules will be constructed using known mechanisms of polarity control in Drosophila dendrites. This model will also incorporate known parameters for microtubule growth dynamics. Model validation will be carried out using experimental perturbations of polarity control mechanisms as well as using measurements of microtubule dynamics. This model will provide a framework for understanding how individual microtubule dynamics and local polarity mechanisms influence microtubule spatial organization and polarity. Aim 2. Neurons normally maintain the same polarized arrangement of microtubules for a lifetime. However, if the axon is removed, a dendrite can reverse polarity and become a regenerating axon. How polarity reversal occurs is not understood. The hypothesis that increased entry of microtubules from the cell body drives reversal will be tested in vivo and in silico. The role of other control mechanisms in polarity reversal will also be systematically addressed by testing the mathematical model and informing new experimental directions. Aim 3. Most neurons have several dendrites emerging from the cell body. After axon damage, only one dendrite switches polarity whereas the others revert to their pre-injury orientation. This selection bias is hypothesized to depend on branching patterns of the dendrites. The combination of axon removal experiments in neurons with distinct branching features and a reduced mathematical description of microtubule behavior will provide insights on how dendrite geometry influences polarity control, robustness, and regeneration. The combination of sophisticated live imaging of microtubules in neurons in vivo with new mathematical modeling of microtubule behavior will drive new insights on how neurons maintain a polarized, yet dynamic and flexible microtubule cytoskeleton for a lifetime. The challenges posed by axonal injury require radical cytoskeletal reorganization. Models developed and validated with measurements from healthy neurons will be used to gain deep understanding of microtubule control mechanisms that are critical for axonal regeneration.