SELECTIVE DISRUPTION OF THE CELLULAR MACHINERY TO DISSECT NEURONAL DIFFERENTIATION

Precise growth cone responses to guidance cues require the coordinated action of cellular effectors, namely actin reorganization and membrane dynamics, to steer the axon. Previous findings demonstrated that second messenger modulation in restricted microdomains of the plasma membrane encode the blueprint of an axon response to a set guidance cue. This showed that membrane compartmentalization is a key feature regulating neuronal responses to guidance cues. Membrane exocytosis and actin reorganization have a strong role in the establishment and maintenance of neuronal cytoarchitecture. We hypothesized that local confinement of these effectors could be defining the structure and physiology of the neuron.
We engineered a set of molecular tools to selectively disrupt SNARE-mediated exocytosis and actin polymerization in specific microdomains of the plasma membrane, to assess whether cell-effector compartmentalization regulates neuronal cytoarchitecture. NSF700 and DeAct-GS1 were targeted to, or excluded from, lipid rafts, to analyze the role of localized exocytosis and actin polymerization in key aspects of neuronal differentiation in culture. We show the effects of selective disruption of membrane dynamics and actin polymerization on neuronal polarization, axon elongation, dendrite arborization, and spine formation in vitro, and highlight how membrane microdomains help define the complexity of the neuron.
Lipid raft-targeted and -excluded NSF700 and DeAct-GS1 enable understanding the role of membrane compartmentalization in the development of neuronal cytoarchitecture and provide a new viewpoint on the acquisition of neuronal features. Manipulating these features “a la carte” opens new avenues on therapeutic interventions and regeneration.