Phosphorylation regulates arginine-rich RNA-binding protein solubility and oligomerization.
RNA-binding proteins
SC35
SRSF2
mass spectrometry
phosphorylation
posttranslational modifications (PTMs)
protein interactions
Journal
The Journal of biological chemistry
ISSN: 1083-351X
Titre abrégé: J Biol Chem
Pays: United States
ID NLM: 2985121R
Informations de publication
Date de publication:
11 2021
11 2021
Historique:
received:
25
06
2021
revised:
05
10
2021
accepted:
13
10
2021
pubmed:
22
10
2021
medline:
18
12
2021
entrez:
21
10
2021
Statut:
ppublish
Résumé
Posttranslational modifications (PTMs) such as phosphorylation of RNA-binding proteins (RBPs) regulate several critical steps in RNA metabolism, including spliceosome assembly, alternative splicing, and mRNA export. Notably, serine-/arginine- (SR)-rich RBPs are densely phosphorylated compared with the remainder of the proteome. Previously, we showed that dephosphorylation of the splicing factor SRSF2 regulated increased interactions with similar arginine-rich RBPs U1-70K and LUC7L3. However, the large-scale functional and structural impact of these modifications on RBPs remains unclear. In this work, we dephosphorylated nuclear extracts using phosphatase in vitro and analyzed equal amounts of detergent-soluble and -insoluble fractions by mass-spectrometry-based proteomics. Correlation network analysis resolved 27 distinct modules of differentially soluble nucleoplasm proteins. We found classes of arginine-rich RBPs that decrease in solubility following dephosphorylation and enrich the insoluble pelleted fraction, including the SR protein family and the SR-like LUC7L RBP family. Importantly, increased insolubility was not observed across broad classes of RBPs. We determined that phosphorylation regulated SRSF2 structure, as dephosphorylated SRSF2 formed high-molecular-weight oligomeric species in vitro. Reciprocally, phosphorylation of SRSF2 by serine/arginine protein kinase 2 (SRPK2) in vitro decreased high-molecular-weight SRSF2 species formation. Furthermore, upon pharmacological inhibition of SRPKs in mammalian cells, we observed SRSF2 cytoplasmic mislocalization and increased formation of cytoplasmic granules as well as cytoplasmic tubular structures that associated with microtubules by immunocytochemical staining. Collectively, these findings demonstrate that phosphorylation may be a critical modification that prevents arginine-rich RBP insolubility and oligomerization.
Identifiants
pubmed: 34673031
pii: S0021-9258(21)01112-1
doi: 10.1016/j.jbc.2021.101306
pmc: PMC8569591
pii:
doi:
Substances chimiques
LUC7L3 protein, human
0
Nuclear Proteins
0
Ribonucleoprotein, U1 Small Nuclear
0
SNRNP70 protein, human
0
SRSF2 protein, human
147153-65-9
Serine-Arginine Splicing Factors
170974-22-8
Protein Serine-Threonine Kinases
EC 2.7.11.1
SRPK2 protein, human
EC 2.7.11.1
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
101306Subventions
Organisme : NIGMS NIH HHS
ID : R35 GM138123
Pays : United States
Organisme : NIA NIH HHS
ID : R01 AG053960
Pays : United States
Organisme : NIA NIH HHS
ID : U01 AG061357
Pays : United States
Organisme : NINDS NIH HHS
ID : T32 NS007480
Pays : United States
Organisme : NIA NIH HHS
ID : U01 AG046161
Pays : United States
Organisme : NIGMS NIH HHS
ID : T32 GM008367
Pays : United States
Organisme : NIA NIH HHS
ID : R01 AG061800
Pays : United States
Organisme : NIA NIH HHS
ID : RF1 AG062181
Pays : United States
Informations de copyright
Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.
Déclaration de conflit d'intérêts
Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article.
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