Involvement of myosin V and associated proteins in protein trafficking and neuronal morphogenesis

• Main Author: Lisé, Marie-France
• Format: Others
• Language: English
• Published: University of British Columbia 2009
• Online Access: http://hdl.handle.net/2429/15250

Proper neuronal development and function requires precise sorting and delivery of various elements from the soma to the synapse. Important mediators of intracellular transport events are the actin-based class V myosin motors, which are involved in organelle transport in various cell types. Two myosin V family members, myosin Va and Vb, are present in the brain, however, the identity of cargoes transported by these motors is unknown. The objective of this thesis was to conduct molecular and cell biological studies to identify and characterize novel myosin V cargoes in neurons. The first approach I used was to characterize the distribution of candidate protein cargoes after blocking the function of endogenous myosin Va and Vb with dominant-negative (DN) versions. I found that in developing neurons, expression of DN myosin Vb, but not DN myosin Va, resulted in the accumulation in the soma of the AMPA-type glutamate receptor subunit, GluR1, and a reduction of its surface expression. I also found that myosin Vb-mediated trafficking of GluR1 required an interaction with the GTPase Rab11. These results reveal a novel mechanism for the transport of a specific glutamate receptor subunit mediated by myosin Vb and Rab11. As an alternative approach to identify myosin Va binding partners in the brain, we conducted a yeast-two hybrid screen of a rat brain cDNA library using the cargo binding domain of myosin Va. Among the proteins identified in our screen, I selected a protein of unknown function previously identified as Rab-lysosomal-interacting protein like 2 (RILPL2) and further assessed its function. I found that RILPL2 expression in non-neuronal cells resulted in morphological changes and activation of the Rho GTPase Rac1. In developing neurons, gain or loss of RILPL2 function altered the density of dendritic spine protrusions and increased phosphorylation of the Rac1 effector Pak. These findings uncover a novel role for the myosin Va-interacting protein, RILPL2, in regulating dendritic spine development, possibly through Rac signaling. Taken together, the work presented in this thesis provides novel insights into the function of class V myosins in neurons, and into the critical machinery involved in trafficking of AMPARs and dendritic spine morphogenesis. === Medicine, Faculty of === Graduate