Significance: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that leads to death of spinal motor neurons and paralysis. Genetic studies of ALS patients have identified mutations in autophagy pathway genes including p62 / SQSTM1 , OPTN , TBK1 , VCP , and C9ORF72 . However, the mechanisms by which these mutations cause ALS are not understood. Here we investigated the role of autophagy in a mouse model of ALS by specifically disrupting the critical autophagy gene Atg7 in motor neurons. We found that inhibition of autophagy in motor neurons accelerated disease onset but prolonged lifespan. This increase in longevity was associated with a reduction in glial inflammation and interneuron pathology, indicating a non–cell-autonomous role for motor neuron autophagy in disease pathogenesis.

A common feature of non‐coding repeat expansion disorders is the accumulation of RNA repeats as RNA foci in the nucleus and/or cytoplasm of affected cells. These RNA foci can be toxic because they sequester RNA‐binding proteins, thus affecting various steps of post‐transcriptional gene regulation. However, the precise step that is affected by C9orf72 GGGGCC (G4C2) repeat expansion, the major genetic cause of amyotrophic lateral sclerosis (ALS), is still poorly defined. In this work, we set out to characterise these mechanisms by identifying proteins that bind to C9orf72 RNA. Sequestration of some of these factors into RNA foci was observed when a (G4C2)₃₁ repeat was expressed in NSC34 and HeLa cells. Most notably, (G4C2)₃₁ repeats widely affected the distribution of Pur‐alpha and its binding partner fragile X mental retardation protein 1 (FMRP, also known as FMR1), which accumulate in intra‐cytosolic granules that are positive for stress granules markers. Accordingly, translational repression is induced. Interestingly, this effect is associated with a marked accumulation of poly(A) mRNAs in cell nuclei. Thus, defective trafficking of mRNA, as a consequence of impaired nuclear mRNA export, might affect translation efficiency and contribute to the pathogenesis of C9orf72 ALS.

Recent years have witnessed a renewed interest in the pathogenic mechanisms of amyotrophic lateral sclerosis (ALS), a late-onset progressive degeneration of motor neurons. The discovery of new genes associated with the familial form of the disease, along with a deeper insight into pathways already described for this disease, has led scientists to reconsider previous postulates. While protein misfolding, mitochondrial dysfunction, oxidative damage, defective axonal transport, and excitotoxicity have not been dismissed, they need to be re-examined as contributors to the onset or progression of ALS in the light of the current knowledge that the mutations of proteins involved in RNA processing, apparently unrelated to the previous "old partners," are causative of the same phenotype. Thus, newly envisaged models and tools may offer unforeseen clues on the etiology of this disease and hopefully provide the key to treatment.

Significance: DNA sequence variants in the Tbk1 gene associate with both sporadic and familial amyotrophic lateral sclerosis (ALS). We report a functional study of 25 missense TBK1 mutations scattered across all structural domains. These mutations negatively impact TBK1 functions ranging from kinase activity, substrate specificity, dimerization, and protein stability to interactions with critical autophagy receptors. Contrary to previous studies, we find that TBK1 dimerization increases protein stability and improves catalytic efficiency, but it is not required for its activation. Our results show that different TBK1 ALS mutations, which are associated with the same pathological outcome, affect distinct signaling pathways in which TBK1 is critically involved. TBK1-associated ALS provides a remarkable example of the genetic and functional complexity of ALS disease mechanisms.