Autophagy and Neurodegeneration: Insights from a Cultured Cell Model of ALS

Abstract

Autophagy plays a major role in the elimination of cellular waste components, the renewal of intracellular proteins and the prevention of the build-up of redundant or defective material. It is fundamental for the maintenance of homeostasis and especially important in post-mitotic neuronal cells, which, without competent autophagy, accumulate protein aggregates and degenerate. Many neurodegenerative diseases are associated with defective autophagy; however, whether altered protein turnover or accumulation of misfolded, aggregate-prone proteins is the primary insult in neurodegeneration has long been a matter of debate. Amyotrophic lateral sclerosis (ALS) is a fatal disease characterized by selective degeneration of motor neurons. Most of the ALS cases occur in sporadic forms (SALS), while 10%-15% of the cases have a positive familial history (FALS). The accumulation in the cell of misfolded/abnormal proteins is a hallmark of both SALS and FALS, and altered protein degradation due to autophagy dysregulation has been proposed to contribute to ALS pathogenesis. In this review, we focus on the main molecular features of autophagy to provide a framework for discussion of our recent findings about the role in disease pathogenesis of the ALS-linked form of the VAPB gene product, a mutant protein that drives the generation of unusual cytoplasmic inclusions.

Keywords: ALS; UPS; VAPB; aggregates; autophagy; autophagy receptors; cytoplasmic inclusions; neurodegeneration; protein degradation; proteostasis.

Figure 1

Molecular links between the two main protein degradation pathways in the cell: the proteasomal and autophagolysosomal systems. The cartoon illustrates the involvement of receptors/adaptors/chaperones common to both systems in the process of the degradation of misfolded proteins, as an example of a function highly relevant to neurodegenerative diseases, such as ALS. See the text for further explanation. UBD, ubiquitin-binding domain; LIR, LC3-interacting region; MVB, multivesicular body.

Figure 2

Mutant VAPB inclusions in a model motor neuronal cell line (NSC34 cells) (A) and in HeLa cells (B). (A) NSC34 cells stably transfected with wild-type (left) or P56S-VAPB (right) under a Tet-repressible promoter were grown in the absence of the antibiotic to induce expression of the transgene. Shown are fluorescence images (top) and fluorescence superimposed on the phase contrast images of the same two fields (bottom). The inset of the upper left panel shows a two-fold enlargement of the boxed area and illustrates the web-like distribution of wt VAPB, typical of an ER protein, as opposed to the clusters of mutant protein that congregate around the nucleus. Modified from [160]. Scale bar, 15 µm. (B) HeLa cell lines, expressing P56S-VAPB under a Tet-repressible promoter, were grown in the absence of the antibiotic (left) or at low concentration (right, 0.1 ng/mL) and then analyzed by transmission EM. Arrows in the upper panels indicate the paired cisternae with interposed dense cytosolic layer; such structures can be visualized both in moderately (left) and low (right) expressing cells. The lower panels show the inclusions at higher magnification. The arrowheads and asterisks indicate the dense cytosolic layer and attached ribosomes, respectively. RER, rough endoplasmic reticulum, M, mitochondria, N, nucleus. Scale bars: upper panels, 500 nm; lower panels 100 nm. Modified from [159].