Protein quality control is of critical importance in maintaining cellular protein homeostasis, especially during stress, disease states and aging. These networks are comprised of: 1) chaperones that assist in protein folding and stabilization of intermediates to prevent aggregation, and 2) degradation pathways, such as the ubiquitin proteasome system and autophagy, to eliminate proteins and organelles with sub-optimal function. A decline in protein quality control in aging cells contributes to the development of numerous genetically and clinically distinct neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS), Alzheimer's, Parkinson's, Huntington's and prion diseases. Valosin containing protein (VCP, also known as p97) is a AAA-ATPase that plays an important role in the ubiquitin proteasome system to capture ubiquitylated substrates via designated adaptors to mediate substrate degradation. Mutation of VCP causes several neurodegenerative disorders including ALS and a rare multi-system disorder, inclusion body myopathy, Paget?s disease of the bone and frontotemporal dementia (IBMPFD). The most commonly mutated region of VCP resides at the site of adaptor binding and in certain instances these mutations alter the constellation of adaptors that are bound to VCP skewing its targeting to substrates. A common cellular feature of VCP mutations is the formation and persistence of protein aggregates that fail to be cleared. We have identified a specific VCP-adaptor complex that is targeted to the aggresome, a structure that sequesters misfolded aggregated proteins formed during cell stress. While aggresomes are generally cyto-protective, under conditions where they persist, for example in aging neurons with declining protein quality control pathways, they eventually become cytotoxic. Depletion of the VCP adaptor leads to a deficit in aggresome formation, stabilization of ubiquitylated substrates and triggers ER stress. We hypothesize that VCP mediates aggresome clearance by adaptor specific targeting to enable clearance of specific misfolded or aggregated substrates. We will use a combination of targeted molecular studies, quantitative proteomics and high-resolution imaging to achieve the main objectives of this proposal. We will (1) characterize the VCP aggresome targeted complex to determine mechanisms of targeting and its functions within the aggresome (2) identify ubiquitylated cellular targets of the VCP complex targeted to the aggresome using ubiquitin remnant capture proteomics and (3) determine whether disease-relevant VCP mutations impact its association with the aggresome complex and test if the VCP adaptor can recognize and enable clearance of disease relevant protein aggregates. Successful implementation of this proposal will identify new targets that can be pursued for the development of agents that enable aggregate clearance in a number of neurodegenerative disorders.
The formation of protein aggregates in aging neurons is an underlying cause of a variety of neurodegenerative disorders. A decline in the optimal functioning of pathways that enable clearance of potentially toxic protein aggregates is linked to a variety of neurodegenerative diseases, and boosting these pathways in aging neurons may have therapeutic benefit. Our goal is to characterize these cellular pathways in mechanistic detail to identify new therapeutic targets, thereby enabling the development of agents to treat several protein aggregation disorders.
