Abstract
Fused in sarcoma (FUS) is a nuclear RNA-binding protein. Mutations in FUS lead to the mislocalization of FUS from the nucleus to the cytosol and formation of pathogenic aggregates in neurodegenerative diseases including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD), yet with unknown molecular mechanisms. Using mutant and stress conditions, we visualized FUS localization and aggregate formation in cells. We used single-molecule pull-down (SiMPull) to quantify the native oligomerization states of wildtype (WT) and mutant FUS in cells. We demonstrate that the NLS mutants exhibited the highest oligomerization (>3) followed by other FUS mutants (>2) and WT FUS which is primarily monomeric. Strikingly, the mutant FUS oligomers are extremely stable and resistant to treatment by high salt, hexanediol, RNase, and Karyopherin-β2 and only soluble in GdnHCl and SDS. We propose that the increased oligomerization units of mutant FUS and their high stability may contribute to ALS/FTLD pathogenesis.
Keywords:
Biophysics; Molecular interaction; Properties of biomolecules.