Role of dynein in the clearance of misfolded proteins responsible for ALS

Amyotrophic lateral sclerosis (ALS) and spinal and bulbar muscular atrophy (SBMA) share several similarities including the formation of aggregates of proteins with aberrant conformation (misfolded). Mutated SOD1, TDP-43, FUS-TLS, the C9ORF72 dipeptide, etc. in ALS and mutated androgen receptor (AR) in SBMA are misfolded proteins that aggregate. In affected motoneurons, the protein quality control (PQC) system (chaperones, autophagy and ubiquitin-proteasome system (UPS)) protects from misfolded proteins toxicity. The HSPB8-BAG3-HSC70-CHIP complex promotes aggregate degradation by autophagy. BAG3 interacts with dynein transport of mutant proteins at microtubule organization center (MTOC) where they aggregate, and degraded by autophagy. However, here misfolded proteins may impair autophagy causing flux blockage. In NSC34 cells we evaluated the role of dynein into aggregates formation process. Unexpectedly, treatment with a dynein inhibitor (EHNA) drastically reduced the retention of mutSOD1, mutTDP43 and mutAR aggregates in filter retardation assay (FRA), even when autophagy was inhibits with 3-MA. Conversely, UPS blockage with MG132 counteracted the reduction induced by altered dynein transports. In addition, the silencing of dynein heavy chain resulted in a drastic reduction of mutAR retained in FRA. Moreover, dynein silencing drastically altered localization of autophagic markers (LC3 and p62). RTq-PCR on RNAs from NSC34 cells treated with EHNA showed an increased BAG1:BAG3 ratio to re-route misfolded proteins to UPS. Moreover, EHNA increased the degradation of proteasome reporter GFPu, while BAG1 overexpression reduced the level of aggregates retained in FRA. Collectively, these data suggest that when autophagy is overload by misfolded proteins, dynein inhibition restores the physiological and soluble protein pool via UPS.