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Title: Phosphorylation of the FUS low-complexity domain disrupts phase separation, aggregation, and toxicity

Abstract

Neuronal inclusions of aggregated RNA-binding protein fused in sarcoma (FUS) are hallmarks of ALS and frontotemporal dementia subtypes. Intriguingly, FUS's nearly uncharged, aggregation-prone, yeast prion-like, low sequence-complexity domain (LC) is known to be targeted for phosphorylation. Here we map in vitro and in-cell phosphorylation sites across FUS LC. We show that both phosphorylation and phosphomimetic variants reduce its aggregation-prone/prion-like character, disrupting FUS phase separation in the presence of RNA or salt and reducing FUS propensity to aggregate. Nuclear magnetic resonance spectroscopy demonstrates the intrinsically disordered structure of FUS LC is preserved after phosphorylation; however, transient domain collapse and self-interaction are reduced by phosphomimetics. Moreover, we show that phosphomimetic FUS reduces aggregation in human and yeast cell models, and can ameliorate FUS-associated cytotoxicity. Hence, post-translational modification may be a mechanism by which cells control physiological assembly and prevent pathological protein aggregation, suggesting a potential treatment pathway amenable to pharmacologic modulation.

Authors:
 [1];  [2];  [3];  [3];  [1];  [4];  [5]; ORCiD logo [4];  [6];  [7];  [7];  [5]; ORCiD logo [4]; ORCiD logo [1]; ORCiD logo [8]
  1. Uniformed Services Univ., Bethesda, MD (United States). Dept. of Pharmacology and Molecular Therapeutics
  2. Brown Univ., Providence, RI (United States). Neuroscience Graduate Program
  3. Brown Univ., Providence, RI (United States). Dept. of Molecular Pharmacology, Physiology, and Biotechnology
  4. Lehigh Univ., Bethlehem, PA (United States). Dept. of Chemical and Biomolecular Engineering
  5. Johns Hopkins Univ., Baltimore, MD (United States). Johns Hopkins Mass Spectrometry and Proteomic Facility
  6. Brown Univ., Providence, RI (United States). Graduate Program in Molecular Biology, Cell Biology and Biochemistry
  7. National Inst. of Health (NIH), Bethesda, MD (United States). Lab. of Chemical Physics
  8. Brown Univ., Providence, RI (United States). Neuroscience Graduate Program. Dept. of Molecular Pharmacology, Physiology, and Biotechnology. Graduate Program in Molecular Biology, Cell Biology and Biochemistry
Publication Date:
Research Org.:
Lehigh Univ., Bethlehem, PA (United States); Brown Univ., Providence, RI (United States); Uniformed Services Univ., Bethesda, MD (United States); Johns Hopkins Univ., Baltimore, MD (United States); National Inst. of Health (NIH), Bethesda, MD (United States); Lawrence Berkeley National Laboratory-National Energy Research Scientific Computing Center
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22); National Inst. of Health (NIH) (United States); Rhode Island Foundation (United States); National Science Foundation (NSF); Rhode Island Science and Technology Advisory Council (United States)
OSTI Identifier:
1547127
Alternate Identifier(s):
OSTI ID: 1499120; OSTI ID: 1547129
Grant/Contract Number:  
SC0013979; AC02‐05CH11231; S10RR027027; 1S10RR020923; S10RR02763; P20RR018728; P30RR031153; P30GM103410; R01GM118530; R35GM119790; P20GM104937; T32 MH020068; T32 GM07601; 20133966; TG‐MCB‐120014; 0554548; 1004057; DE‐AC02‐05CH11231
Resource Type:
Published Article
Journal Name:
EMBO Journal
Additional Journal Information:
Journal Volume: 36; Journal Issue: 20; Journal ID: ISSN 0261-4189
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; amyotrophic lateral sclerosis; frontotemporal dementia; intrinsically disordered protein; prion; ribonucleoprotein granule

Citation Formats

Monahan, Zachary, Ryan, Veronica H., Janke, Abigail M., Burke, Kathleen A., Rhoads, Shannon N., Zerze, Gül H., O'Meally, Robert, Dignon, Gregory L., Conicella, Alexander E., Zheng, Wenwei, Best, Robert B., Cole, Robert N., Mittal, Jeetain, Shewmaker, Frank, and Fawzi, Nicolas L. Phosphorylation of the FUS low-complexity domain disrupts phase separation, aggregation, and toxicity. United States: N. p., 2017. Web. doi:10.15252/embj.201696394.
Monahan, Zachary, Ryan, Veronica H., Janke, Abigail M., Burke, Kathleen A., Rhoads, Shannon N., Zerze, Gül H., O'Meally, Robert, Dignon, Gregory L., Conicella, Alexander E., Zheng, Wenwei, Best, Robert B., Cole, Robert N., Mittal, Jeetain, Shewmaker, Frank, & Fawzi, Nicolas L. Phosphorylation of the FUS low-complexity domain disrupts phase separation, aggregation, and toxicity. United States. doi:10.15252/embj.201696394.
Monahan, Zachary, Ryan, Veronica H., Janke, Abigail M., Burke, Kathleen A., Rhoads, Shannon N., Zerze, Gül H., O'Meally, Robert, Dignon, Gregory L., Conicella, Alexander E., Zheng, Wenwei, Best, Robert B., Cole, Robert N., Mittal, Jeetain, Shewmaker, Frank, and Fawzi, Nicolas L. Tue . "Phosphorylation of the FUS low-complexity domain disrupts phase separation, aggregation, and toxicity". United States. doi:10.15252/embj.201696394.
@article{osti_1547127,
title = {Phosphorylation of the FUS low-complexity domain disrupts phase separation, aggregation, and toxicity},
author = {Monahan, Zachary and Ryan, Veronica H. and Janke, Abigail M. and Burke, Kathleen A. and Rhoads, Shannon N. and Zerze, Gül H. and O'Meally, Robert and Dignon, Gregory L. and Conicella, Alexander E. and Zheng, Wenwei and Best, Robert B. and Cole, Robert N. and Mittal, Jeetain and Shewmaker, Frank and Fawzi, Nicolas L.},
abstractNote = {Neuronal inclusions of aggregated RNA-binding protein fused in sarcoma (FUS) are hallmarks of ALS and frontotemporal dementia subtypes. Intriguingly, FUS's nearly uncharged, aggregation-prone, yeast prion-like, low sequence-complexity domain (LC) is known to be targeted for phosphorylation. Here we map in vitro and in-cell phosphorylation sites across FUS LC. We show that both phosphorylation and phosphomimetic variants reduce its aggregation-prone/prion-like character, disrupting FUS phase separation in the presence of RNA or salt and reducing FUS propensity to aggregate. Nuclear magnetic resonance spectroscopy demonstrates the intrinsically disordered structure of FUS LC is preserved after phosphorylation; however, transient domain collapse and self-interaction are reduced by phosphomimetics. Moreover, we show that phosphomimetic FUS reduces aggregation in human and yeast cell models, and can ameliorate FUS-associated cytotoxicity. Hence, post-translational modification may be a mechanism by which cells control physiological assembly and prevent pathological protein aggregation, suggesting a potential treatment pathway amenable to pharmacologic modulation.},
doi = {10.15252/embj.201696394},
journal = {EMBO Journal},
number = 20,
volume = 36,
place = {United States},
year = {2017},
month = {8}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
DOI: 10.15252/embj.201696394

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Cited by: 88 works
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