Neuronal trafficking: basic mechanisms and ALS pathology

Abstract

A cell is divided into different compartments and organelles, which enables the cell to create specialized environments for specific functions. To perform these functions, organelles need a unique composition of proteins and lipids. By actively controlling the trafficking of proteins and membrane lipids, the function of these organelles can be maintained and proteins get to their appropriate location. Because of their polarized and extended morphologies, intracellular transport is particularly important for neurons. Abnormalities in the function of intracellular trafficking have been implicated in multiple neurodegenerative disorders. The work presented in this thesis focusses on Amyotrophic Lateral Sclerosis (ALS), a neurodegenerative disease characterized by the degeneration of motor neurons, leading to paralysis and patient death. Mutations in the VAMP-Associated Protein B (VAPB) have been identified in patients with ALS. VAPB proteins are located in the endoplasmic reticulum (ER), an organelle important for the proper folding of proteins. VAP proteins (VAPA and VAPB) have been implicated in a variety of processes and, given that they localize to one of the secretory organelles, it is not surprising that many of the proposed functions involve regulation of the secretory machinery. First, we studied the role of VAPB in the secretory pathway in neurons. We identified a new VAPB binding partner, YIF1A, a protein present in ER-Golgi intermediate compartments (ERGIC) and show that VAPB and YIF1A are required for intracellular membrane trafficking into dendrites. Aggregation of the ALS mutant VAPB leads to the mislocalization of YIF1A and we propose that this could contribute to the pathological mechanisms observed in ALS. Second, we used transgenic mice to show that these VAPB aggregates are not neurotoxic, but rather represent protective structures associated with ER associated degradation (ERAD). Third, we studied Golgi fragmentation, a poorly understood neuropathological phenomenon identified in ALS motor neurons. Fragmentation of the Golgi is a very early event in ALS and is associated with altered intracellular trafficking. The second series of studies were aimed to obtain more fundamental insights into the regulation of transport in neurons, zooming in on the question how neuronal cargos are sorted between axons and dendrites. Using a trafficking assay in hippocampal neurons to selectively probe specific motor protein activity we found that dynein motors drive cargo, such as AMPA receptors, selectively into dendrites governed by their mixed microtubules. Another level of regulation is exerted by adaptor proteins, such as TRAK1 and 2, which link mitochondria to different microtubule-based motors. Finally, we studied the role of dynein in axonal transport by investigating NDEL1, a protein involved in the regulation of dynein. NDEL1 specifically localizes to the first part of the axon, called the axon initial segment (AIS). We propose a model in which NDEL1 controls dynein activity at the AIS and facilitates the reversal of somatodendritic transport cargos. Taken together, in this thesis we describe protein aggregation and sorting abnormalities in ALS. We also describe two novel roles for dynein in neuronal transport; targeting of dendrites and cargo reversal in the axon.