Endolysosomal Regulation of Aggregation-Prone Proteins in ALS

Abstract

Many neurodegenerative diseases are characterized by the misfolding and aggregation of proteins within neurons. In the motor neurons of amyotrophic lateral sclerosis (ALS) patients, the RNA-binding protein, TDP-43, mislocalizes to the cytoplasm where it misfolds and aggregates. This aggregation potentially causes the death of motor neurons giving rise to the clinical symptoms of ALS. Aggregation of proteins can arise due to age-related decline in the proteostasis network. In this thesis, we focus on understanding the regulation of TDP-43 by the proteostasis network and identify a mechanism and novel regulators of TDP-43 degradation.In Chapter 2, we identified Cdc48/VCP as a regulator of TDP-43 degradation by endolysosomal flux and demonstrated that another aggregation-prone ALS-associated protein, FUS, utilizes a similar mechanism of clearance. Perturbations to endolysosomal flux cause the accumulation and aggregation of TDP-43/FUS and inactivation of Cdc48/VCP mirrored these phenotypes. Additionally, TDP-43/FUS accumulation caused defects endolysosomal flux thereby potentially inhibiting in their own degradation. In Chapter 3, we further characterized the mechanism of endolysosomal degradation that targets TDP-43 for clearance. Using an unbiased screening method to determine regulators of TDP-43 abundance, we found that TDP-43 is subject to ESCRT-dependent internalization at the multivesicular body (MVB) and that ubiquitination of TDP-43 by the E3 ligase, Rsp5/NEDD4, destabilizes and promotes TDP-43 degradation. Further work demonstrated that TDP-43 accumulation causes defects in MVB morphology which could underly the accumulation of TDP-43 seen in ALS patients and the inhibition of endolysosomal flux caused by TDP-43 accumulation.