Spatio-temporal Proteomic Analysis of Stress Granule Disassembly Using APEX Reveals Regulation by SUMOylation and Links to ALS Pathogenesis

Stress-granules (SG) are cytoplasmic assemblies of proteins associated with non-translating mRNAs, which form in response to a variety of cellular stresses. Whereas much has been learned about SG formation, a major gap remains in understanding the compositional changes SG undergo during normal disassembly and under disease conditions. Here, we address this gap by proteomic dissection of the SG temporal disassembly sequence using multi-bait APEX proximity proteomics. We discover 109 novel SG-proteins and characterize biophysically distinct SG substructures at proteomic resolution. We reveal dozens of disassembly-engaged proteins (DEPs) that are specifically recruited, some of which play functional roles in SG disassembly, including small ubiquitin-like modifier (SUMO) ligases. We further demonstrate that SUMOylation regulates SG disassembly and SG formation. Parallel proteomics with amyotrophic lateral sclerosis (ALS)-associated C9ORF72 dipeptides uncovered attenuated recruitment of DEPs during SG disassembly and impaired SUMOylation. Accordingly, SUMO-ligase overexpression ameliorated C9ORF72-ALS-related neurodegeneration in Drosophila. Altogether, our study amplifies the knowledge of SG composition in human cells by dissecting the SG spatio-temporal proteomic landscape, provides an in-depth resource for future work on SG function, and reveals basic and disease-relevant mechanisms of SG disassembly.