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| Status |
Public on Aug 24, 2017 |
| Title |
The Glycogen Synthase Kinase-3 Phosphoproteome: GSK-3 Phosphorylates Multiple Splicing Factors and Regulates Alternative Splicing |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Glycogen Synthase Kinase-3 (GSK-3) is a constitutively active, ubiquitously expressed protein kinase that regulates multiple signaling pathways. Over 100 putative GSK-3 substrates have been reported in diverse cell types based on in vitro kinase assays or genetic and pharmacological manipulation of GSK-3. Many more have been predicted based on a recurrent GSK-3 consensus motif, but this prediction has not been tested by analyzing the GSK-3 phosphoproteome. We used stable isotope labeling of amino acids in culture (SILAC) and mass spectrometry to analyze the repertoire of GSK-3 dependent substrates in mouse embryonic stem cells (ESCs). A comparison of wild-type and Gsk3a;Gsk3b knockout (DKO) ESCs revealed prominent GSK-3-dependent phosphorylation of multiple splicing factors and regulators of RNA biosynthesis, as well as proteins that regulate transcription, translation, and cell division. We demonstrate direct, GSK-3-dependent phosphorylation of the splicing factors RBM8A and PSF as well as the nucleolar protein NPM1. RNA sequencing to compare the transcriptomes of wild-type and Gsk3 DKO ESCs identified more than 210 genes that are alternatively spliced in a GSK-3-dependent manner, supporting a broad role for GSK-3 in regulating alternative splicing. Overall, this study provides the first unbiased analysis of the GSK-3 phosphoproteome and strong evidence for GSK-3 as a regulator of alternative splicing.
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| Overall design |
2 biological replicates of Gsk3a+/+;Gsk3b+/+ (Wild Type) and Gsk3a-/-;Gsk3b-/- (DKO) E15 mouse embryonic stem cells were analyzed by RNA-Seq
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| Contributor(s) |
Mansi S, Simone S, Katarzyna K, Michael MJ, Caleb RM, Amanda R, Yoseph B, Kristen LW, Benjamin GA, Peter KS |
| Citation(s) |
28916722 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| R35 GM118048 |
Molecular Mechanisms and Signal-Induced Regulation of Alternative Splicing |
UNIVERSITY OF PENNSYLVANIA |
KRISTEN W LYNCH |
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| |
| Submission date |
Dec 05, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Kristen Lynch |
| E-mail(s) |
klync@pennmedicine.upenn.edu
|
| Organization name |
University of Pennsylvania
|
| Department |
Biochemistry and Molecular Biophysics
|
| Lab |
Kristen Lynch
|
| Street address |
906 Stellar Chance Labs 422 Curie Blvd
|
| City |
Philadelphia |
| State/province |
Pennsylvania |
| ZIP/Postal code |
19104 |
| Country |
USA |
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| Platforms (1) |
| GPL21103 |
Illumina HiSeq 4000 (Mus musculus) |
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| Samples (4)
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| Relations |
| BioProject |
PRJNA356332 |
| SRA |
SRP094598 |
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