Spatio-temporal Proteomic Analysis of Stress Granule disassembly using APEX Reveals Regulation by SUMOylation and links to ALS pathogenesis

ABSTRACT: Cytoplasmic stress granules (SG) are membraneless organelles that form in response to a variety of cellular stresses by phase-separation of proteins associated with non-translating mRNAs. SG have recently been linked with neurodegeneration, including ALS, however there has been no systematic investigation of SG composition in normal and disease contexts. We created a proximity proteomics platform by using multiple ascorbate peroxidase (APEX2) baits to comprehensively characterize the spatio-temporal organization of SG in normal and disease (ALS-like) conditions. Multi-bait APEX enhanced sensitivity and enabled cross-validation, leading to the mass spectrometric discovery of 109 additional SG proteins, and internal SG substructures at proteomic resolution. A proteomic analysis of SG disassembly over time revealed a group of 349 proteins recruited specifically during SG disassembly, which we named disassembly-engaged proteins (DEPs), including SUMO ligases. A parallel study of SG disassembly in the presence of C9ORF72-associated dipeptides, which are found in patients with ALS and frontotemporal dementia revealed impaired SG disassembly and DEP recruitment, which may be linked to neurodegeneration. Finally, broad SUMOylation of SG proteins was required for SG disassembly, and impaired by C9ORF72-associated dipeptides. Altogether, our study provides a valuable resource, and dissects the SG spatio-temporal proteomic landscape, revealing basic and disease-relevant mechanisms of SG dynamics.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Tamar Geiger  

LAB HEAD: Professor Tamar Geiger

PROVIDER: PXD017330 | Pride | 2020-10-19

REPOSITORIES: Pride