TAR DNA-binding protein (TDP-43, also known as TARDBP) is the major pathological protein in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Large TDP-43 aggregates that are decorated with degradation adaptor proteins are seen in the cytoplasm of remaining neurons in ALS and FTD patients post mortem. TDP-43 accumulation and ALS-linked mutations within degradation pathways implicate failed TDP-43 clearance as a primary disease mechanism. Here, we report the differing roles of the ubiquitin proteasome system (UPS) and autophagy in the clearance of TDP-43. We have investigated the effects of inhibitors of the UPS and autophagy on the degradation, localisation and mobility of soluble and insoluble TDP-43. We find that soluble TDP-43 is degraded primarily by the UPS, whereas the clearance of aggregated TDP-43 requires autophagy. Cellular macroaggregates, which recapitulate many of the pathological features of the aggregates in patients, are reversible when both the UPS and autophagy are functional. Their clearance involves the autophagic removal of oligomeric TDP-43. We speculate that, in addition to an age-related decline in pathway activity, a second hit in either the UPS or the autophagy pathway drives the accumulation of TDP-43 in ALS and FTD. Therapies for clearing excess TDP-43 should therefore target a combination of these pathways.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder. We screened 751 familial ALS patient whole-exome sequences and identified six mutations including p.D40G in the ANXA11 gene in 13 individuals. The p.D40G mutation was absent from 70,000 control whole-exome sequences. This mutation segregated with disease in two kindreds and was present in another two unrelated cases (P = 0.0102), and all mutation carriers shared a common founder haplotype. Annexin A11–positive protein aggregates were abundant in spinal cord motor neurons and hippocampal neuronal axons in an ALS patient carrying the p.D40G mutation. Transfected human embryonic kidney cells expressing ANXA11 with the p.D40G mutation and other N-terminal mutations showed altered binding to calcyclin, and the p.R235Q mutant protein formed insoluble aggregates. We conclude that mutations in ANXA11 are associated with ALS and implicate defective intracellular protein trafficking in disease pathogenesis

Following the mutation screening of genes known to cause amyotrophic lateral sclerosis (ALS) in index cases from 107 familial ALS (FALS) kindred, a point mutation was identified in vesicle-associated membrane protein-associated protein B (VAPB), or VAMP-associated protein B, causing an amino acid change from threonine to isoleucine at codon 46 (T46I) in one FALS case but not in 257 controls. This is an important finding because it is only the second mutation identified in this gene that causes ALS. In order to investigate the pathogenic effects of this mutation, we have used a motor neuron cell line and tissue-specific expression of the mutant protein in Drosophila. We provide substantial evidence for the pathogenic effects of this mutation in abolishing the effect of wild type VAPB in the unfolded protein response, promoting ubiquitin aggregate formation, and activating neuronal cell death. We also report that expression of the mutant protein in the Drosophila motor system induces aggregate deposition, endoplasmic reticulum disorganization, and chaperone up-regulation both in neurons and in muscles. Our integrated analysis of the pathogenic effect of the T46I mutation and the previously identified P56S mutation indicate extensive commonalities in the disease mechanism for these two mutations. In summary, we show that this newly identified mutation in human FALS has a pathogenic effect, supporting and reinforcing the role of VAPB as a causative gene of ALS.