Cellular and molecular mechanisms of endosomal membrane trafficking in neuronal systems.
Complex brain processes from developmental wiring to synaptic plasticity rely on vectorial cell communication. Vectoriality is possible through the asymmetry of neurons. Nerve cells are organized in segregated membrane domains, dendrites and axonal terminals, each with its own functional and molecular identity. The axonal and dendritic membrane proteins that provide that singularity are synthesized in the cell body before being targeted to their proper compartment, a process that uses protein packaging in vesicular carriers. After they reach destination proteins are in a continuous equilibrium between the plasma membrane and internal vesicular compartments. A major example is the exocytic -endocytic cycle of synaptic vesicles. Sorting choices need to be made when proteins leave the cell body and iterative ones at their final destination because of the back and forth traffic from and to the cell surface. Sorting is achieved by a choreographic array of signals on the membrane proteins and cytosolic receptors that pack those proteins on vesicle carriers of unique composition. Understanding how these vesicles of unique composition are made and the molecules involved is currently one of the most active fields in cell biology and an emergent one in neuroscience. My lab is interested in understanding how neuronal organization emerges from sorting and vesicle formation processes. I have focused my research to the analysis of synaptic vesicle formation, since it is the major membrane traffic pathway in neurons. My work has led to the identification of a new pathway of synaptic vesicle biogenesis from endosomes. Using in vitro reconstitution assays with purified organelles, bacterially produced recombinant proteins, I have identified the first layer of components required to generate synaptic vesicles: 1) A regulatory GTPase, ARF1. 2) A neuronal cytosolic complex, AP-3, presumably involved in selective vesicle formation and sorting. 3) The membrane receptor for that complex, the synaptic vesicle protein synaptobrevin II. 4) and a kinase involved in the regulation of the AP-3. Our current work is exploring each one of these topics with the goal to identify interacting molecules and develop stage-specific reagents able to perturb, both in vitro and in vivo, the formation of synaptic vesicles. Exciting new developments in my lab are the possibility to assembly in fibroblast parts of a neuronal vesicle trafficking pathway using neuronal-specific genes and preliminary evidence suggest that this vesicle production pathway could be selectively affected in prevalent brain diseases. My long-term goal is to engineer neuron and organism-models defective in this and other pathways to understand their contribution to normal and pathological processes that depend in the asymmetric nature of neurons.