The role of Valosin Containing Protein (VCP) in the degradation of neurotoxic protein aggregates in Amyotrophic Lateral Sclerosis

Valosin Containing protein (VCP) is a AAA+ ATPase protein that has chaperon-like functions. VCP acts as an homohexamer and it is constituted of two ATPase sub-units and a N-terminal domain. The N-terminal domain interacts with a large number of adaptors and it confers VCP various roles in the Protein Quality Control (PQC) system. VCP function is mainly to extract misfolded proteins, refold them through its ATPase domains and, if it fails, to route them to the ubiquitin proteasome system (UPS). VCP is also found involved in the formation of autophagosomes and in routing misfolded proteins and more complex structures to the autophagic pathway. VCP is mutated in some neurodegenerative disease (NDs) including Amyotrophic Lateral Sclerosis (ALS). ALS is a proteinpathy: in fact, a hallmark of the disease is the formation of aggregates of misfolded protein like the mutant Superoxide Dismutase 1 (SOD1), or wild type TAR-DNA binding protein 43 (TDP-43). These aggregates alter cellular homeostasis by sequestering essential components and could impair the PQC system. The mutation of VCP is correlated with the presence of TDP-43 aggregates and the alteration of the PQC system. This study is aimed to define VCP role in removing protein aggregates present in ALS by modulating VCP expression in cell models of ALS. The models used transiently express SOD1-G93A, a mutation found in ALS patient, and TDP-43. We first confirmed the aggregation of these proteins through Immunofluorescence (IF) and Filter Retardation Assay (FRA). Then we studied these aggregates in the presence of overexpressed wild type VCP. By performing a FRA, we could observe that VCP enhances the clearance of SOD1-G93A aggregation. Moreover, by chemically inhibiting the proteasome with MG132 and the autophagic pathway with 3MA we defined that VCP routes SOD1-G93A aggregates mainly to UPS and partially to the autophagic pathway. In fact, we could observe through FRA that the inhibition of the autophagic pathway does not affect VCP functioning. These data were all confirmed in Western Blot. We also observe through FRA, that the treatment with a chemical inhibitor of VCP (DBeQ), does not alter the levels of aggregation of mutated-SOD1 but it increases the levels of TDP-43 aggregation. These data show an involvement of VCP in the clearance of both mutated-SOD1 and TDP-43. The identification of VCP role could make it a future target for ALS.