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The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization

FUS (fused in sarcoma) is an abundant, predominantly nuclear protein involved in RNA processing. Under various conditions, FUS functionally associates with RNA and other macromolecules to form distinct, reversible phase-separated liquid structures. Persistence of the phase-separated state and increa...

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Autores principales: Rhoads, Shannon N., Monahan, Zachary T., Yee, Debra S., Leung, Andrew Y., Newcombe, Cameron G., O’Meally, Robert N., Cole, Robert N., Shewmaker, Frank P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Cell Biology 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6085830/
https://www.ncbi.nlm.nih.gov/pubmed/29897835
http://dx.doi.org/10.1091/mbc.E17-12-0735
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author Rhoads, Shannon N.
Monahan, Zachary T.
Yee, Debra S.
Leung, Andrew Y.
Newcombe, Cameron G.
O’Meally, Robert N.
Cole, Robert N.
Shewmaker, Frank P.
author_facet Rhoads, Shannon N.
Monahan, Zachary T.
Yee, Debra S.
Leung, Andrew Y.
Newcombe, Cameron G.
O’Meally, Robert N.
Cole, Robert N.
Shewmaker, Frank P.
author_sort Rhoads, Shannon N.
collection PubMed
description FUS (fused in sarcoma) is an abundant, predominantly nuclear protein involved in RNA processing. Under various conditions, FUS functionally associates with RNA and other macromolecules to form distinct, reversible phase-separated liquid structures. Persistence of the phase-separated state and increased cytoplasmic localization are both hypothesized to predispose FUS to irreversible aggregation, which is a pathological hallmark of subtypes of amyotrophic lateral sclerosis and frontotemporal dementia. We previously showed that phosphorylation of FUS’s prionlike domain suppressed phase separation and toxic aggregation, proportionally to the number of added phosphates. However, phosphorylation of FUS’s prionlike domain was previously reported to promote its cytoplasmic localization, potentially favoring pathological behavior. Here we used mass spectrometry and human cell models to further identify phosphorylation sites within FUS’s prionlike domain, specifically following DNA-damaging stress. In total, 28 putative sites have been identified, about half of which are DNA-dependent protein kinase (DNA-PK) consensus sites. Custom antibodies were developed to confirm the phosphorylation of two of these sites (Ser-26 and Ser-30). Both sites were usually phosphorylated in a subpopulation of cellular FUS following a variety of DNA-damaging stresses but not necessarily equally or simultaneously. Importantly, we found DNA-PK–dependent multiphosphorylation of FUS’s prionlike domain does not cause cytoplasmic localization.
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spelling pubmed-60858302018-10-16 The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization Rhoads, Shannon N. Monahan, Zachary T. Yee, Debra S. Leung, Andrew Y. Newcombe, Cameron G. O’Meally, Robert N. Cole, Robert N. Shewmaker, Frank P. Mol Biol Cell Articles FUS (fused in sarcoma) is an abundant, predominantly nuclear protein involved in RNA processing. Under various conditions, FUS functionally associates with RNA and other macromolecules to form distinct, reversible phase-separated liquid structures. Persistence of the phase-separated state and increased cytoplasmic localization are both hypothesized to predispose FUS to irreversible aggregation, which is a pathological hallmark of subtypes of amyotrophic lateral sclerosis and frontotemporal dementia. We previously showed that phosphorylation of FUS’s prionlike domain suppressed phase separation and toxic aggregation, proportionally to the number of added phosphates. However, phosphorylation of FUS’s prionlike domain was previously reported to promote its cytoplasmic localization, potentially favoring pathological behavior. Here we used mass spectrometry and human cell models to further identify phosphorylation sites within FUS’s prionlike domain, specifically following DNA-damaging stress. In total, 28 putative sites have been identified, about half of which are DNA-dependent protein kinase (DNA-PK) consensus sites. Custom antibodies were developed to confirm the phosphorylation of two of these sites (Ser-26 and Ser-30). Both sites were usually phosphorylated in a subpopulation of cellular FUS following a variety of DNA-damaging stresses but not necessarily equally or simultaneously. Importantly, we found DNA-PK–dependent multiphosphorylation of FUS’s prionlike domain does not cause cytoplasmic localization. The American Society for Cell Biology 2018-08-01 /pmc/articles/PMC6085830/ /pubmed/29897835 http://dx.doi.org/10.1091/mbc.E17-12-0735 Text en © 2018 Rhoads et al. “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology. http://creativecommons.org/licenses/by-nc-sa/3.0 This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License.
spellingShingle Articles
Rhoads, Shannon N.
Monahan, Zachary T.
Yee, Debra S.
Leung, Andrew Y.
Newcombe, Cameron G.
O’Meally, Robert N.
Cole, Robert N.
Shewmaker, Frank P.
The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization
title The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization
title_full The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization
title_fullStr The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization
title_full_unstemmed The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization
title_short The prionlike domain of FUS is multiphosphorylated following DNA damage without altering nuclear localization
title_sort prionlike domain of fus is multiphosphorylated following dna damage without altering nuclear localization
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6085830/
https://www.ncbi.nlm.nih.gov/pubmed/29897835
http://dx.doi.org/10.1091/mbc.E17-12-0735
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