Insights into the Atomistic Mechanisms of Phosphorylation in Disrupting Liquid–Liquid Phase Separation and Aggregation of the FUS Low-Complexity Domain

Fused in sarcoma (FUS), a nuclear
RNA binding protein, can not
only undergo liquid–liquid phase separation (LLPS) to form
dynamic biomolecular condensates but also aggregate into solid amyloid
fibrils which are associated with the pathology of amyotrophic lateral
sclerosis and frontotemporal lobar degeneration diseases. Phosphorylation
in the FUS low-complexity domain (FUS-LC) inhibits FUS LLPS and aggregation.
However, it remains largely elusive what are the underlying atomistic
mechanisms of this inhibitory effect and whether phosphorylation can
disrupt preformed FUS fibrils, reversing the FUS gel/solid phase toward
the liquid phase. Herein, we systematically investigate the impacts
of phosphorylation on the conformational ensemble of the FUS37–97 monomer and dimer and the structure of the FUS37–97 fibril by performing extensive all-atom molecular dynamics simulations.
Our simulations reveal three key findings: (1) phosphorylation shifts
the conformations of FUS37–97 from the β-rich,
fibril-competent state toward a helix-rich, fibril-incompetent state;
(2) phosphorylation significantly weakens protein–protein interactions
and enhances protein–water interactions, which disfavor FUS-LC
LLPS as well as aggregation and facilitate the dissolution of the
preformed FUS-LC fibril; and (3) the FUS37–97 peptide
displays a high β-strand probability in the region spanning
residues 52–67, and phosphorylation at S54 and S61 residues
located in this region is crucial for the disruption of LLPS and aggregation
of FUS-LC. This study may pave the way for ameliorating phase-separation-related
pathologies via site-specific phosphorylation.