Global and Site-Specific Effect of Phosphorylation on Protein Turnover
Language English Country United States Media print-electronic
Document type Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't
Grant support
R01 GM137031
NIGMS NIH HHS - United States
PubMed
33238149
PubMed Central
PMC7855865
DOI
10.1016/j.devcel.2020.10.025
PII: S1534-5807(20)30875-3
Knihovny.cz E-resources
- Keywords
- DeltaSILAC, data-independent acquisition, mass spectrometry, phosphomodiform, phosphorylation, protein lifetime, protein turnover, proteomics, pulse SILAC,
- MeSH
- Cell Cycle physiology MeSH
- Cyclin-Dependent Kinases genetics metabolism MeSH
- Phosphoproteins chemistry metabolism MeSH
- Phosphorylation MeSH
- Glutamates metabolism MeSH
- Mass Spectrometry methods MeSH
- Isotope Labeling methods MeSH
- Humans MeSH
- Cell Line, Tumor MeSH
- Peptides metabolism MeSH
- Peroxiredoxin VI chemistry metabolism MeSH
- Proteolysis * MeSH
- Proteome genetics metabolism MeSH
- Proteomics methods MeSH
- Amino Acid Sequence MeSH
- RNA Splicing Factors chemistry metabolism MeSH
- Signal Transduction genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Research Support, N.I.H., Extramural MeSH
- Names of Substances
- Cyclin-Dependent Kinases MeSH
- Phosphoproteins MeSH
- Glutamates MeSH
- Peptides MeSH
- Peroxiredoxin VI MeSH
- PRDX6 protein, human MeSH Browser
- Proteome MeSH
- RNA Splicing Factors MeSH
- SF3B1 protein, human MeSH Browser
To date, the effects of specific modification types and sites on protein lifetime have not been systematically illustrated. Here, we describe a proteomic method, DeltaSILAC, to quantitatively assess the impact of site-specific phosphorylation on the turnover of thousands of proteins in live cells. Based on the accurate and reproducible mass spectrometry-based method, a pulse labeling approach using stable isotope-labeled amino acids in cells (pSILAC), phosphoproteomics, and a unique peptide-level matching strategy, our DeltaSILAC profiling revealed a global, unexpected delaying effect of many phosphosites on protein turnover. We further found that phosphorylated sites accelerating protein turnover are functionally selected for cell fitness, enriched in Cyclin-dependent kinase substrates, and evolutionarily conserved, whereas the glutamic acids surrounding phosphosites significantly delay protein turnover. Our method represents a generalizable approach and provides a rich resource for prioritizing the effects of phosphorylation sites on protein lifetime in the context of cell signaling and disease biology.
Department of Systems Biology Columbia University New York NY USA
German Cancer Research Center DKFZ 69120 Heidelberg Germany
Yale Cancer Biology Institute Yale University West Haven CT 06516 USA
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