@article{a1886d7c8236480e9eff13db527ec1ca,
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.",
keywords = "amyotrophic lateral sclerosis, frontotemporal dementia, intrinsically disordered protein, prion, ribonucleoprotein granule",
author = "Zachary Monahan and Ryan, {Veronica H.} and Janke, {Abigail M.} and Burke, {Kathleen A.} and Rhoads, {Shannon N.} and Zerze, {G{\"u}l H.} and Robert O'Meally and Dignon, {Gregory L.} and Conicella, {Alexander E.} and Wenwei Zheng and Best, {Robert B.} and Cole, {Robert N.} and Jeetain Mittal and Frank Shewmaker and Fawzi, {Nicolas L.}",
note = "Funding Information: We thank Michael Clarkson for helpful discussions. We thank Geoff Williams at the Leduc Bioimaging Facility at Brown University for microscopy assistance. We would like to thank Drs. Rachel Cox, USU Department of Biochemistry, and Dennis McDaniel, USU Biomedical Instrumentation Center, for help with fluorescence microscopy, and Mr. Michael Panagos, USU Department of Pharmacology, for help with project organization. Research reported in this publication was supported in part by the National Institute Of General Medical Sciences (NIGMS) of the National Institutes of Health (NIH) under Award Numbers R01GM118530 (to N.L.F) and R35GM119790 (to F.S.), a subproject as part of an Institutional Development Award (IDeA) from NIGMS (P20GM104937), a grant from the DEARS Foundation (to N.L.F.), and Medical Research Grant no. 20133966 from the Rhode Island Foundation (to N.L.F). V.H.R. was supported in part by an NIMH training grant to the Neuroscience Graduate Program at Brown University (T32 MH020068). A.E.C. was supported in part by an NIGMS training grant to the graduate program in Molecular Biology, Cell Biology and Biochemistry (MCB) at Brown University (T32 GM07601) and a BIBS Graduate Award in Brain Science from the Brown Institute for Brain Science Reisman Fund. Work at Lehigh University was supported by the U.S. Department of Energy (DOE), Office of Science, Basic Energy Sciences (BES), Division of Material Sciences and Engineering, under Award DE-SC0013979 (to J.M.). Use of the high-performance computing capabilities of the Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by the National Science Foundation (NSF) Grant TG-MCB-120014, is gratefully acknowledged. This research used resources of the National Energy Research Scientific Computing Center, a DOE Office of Science User Facility supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231. This research is based in part on data obtained at the Brown University Structural Biology Core Facility supported by the Division of Biology and Medicine, Brown University. We thank Christoph Schorl and the Brown Genomics Core Facility supported by NIGMS P30GM103410, NCRR P30RR031153, P20RR018728, and S10RR02763, National Science Foundation EPSCoR 0554548. This research is based in part upon work conducted using the Rhode Island NSF/EPSCoR Proteomics Share Resource Facility, which is supported in part by the National Science Foundation EPSCoR Grant No. 1004057, National Institutes of Health Grant No. 1S10RR020923, S10RR027027, a Rhode Island Science and Technology Advisory Council grant, and the Division of Biology and Medicine, Brown University. We are grateful to Pfizer for generously providing calicheamicin. The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding agencies. Publisher Copyright: {\textcopyright} 2017 The Authors. Published under the terms of the CC BY NC ND 4.0 license",
year = "2017",
month = oct,
day = "16",
doi = "10.15252/embj.201696394",
language = "English (US)",
volume = "36",
pages = "2951--2967",
journal = "EMBO Journal",
issn = "0261-4189",
publisher = "Nature Publishing Group",
number = "20",
}