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Updated: Apr 23, 2026
Phosphoproteomic Strategy for Profiling Osmotic Stress Signaling in Arabidopsis
Published on: June 25, 2020
Quantitative phosphoproteomics reveals crosstalk between phosphorylation and O-GlcNAc in the DNA damage response
Jun Zhong1, Marissa Martinez, Srona Sengupta
1McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Abstract:
The modification of intracellular proteins by monosaccharides of O-linked β-N-acetylglucosamine (O-GlcNAc) is an essential and dynamic PTM of metazoans. The addition and removal of O-GlcNAc is catalyzed by the O-GlcNAc transferase (OGT) and O-GlcNAcase, respectively. One mechanism by which O-GlcNAc is thought to mediate proteins is by regulating phosphorylation. To provide insight into the pathways regulated by O-GlcNAc, we have utilized SILAC-based quantitative proteomics to carry out comparisons of site-specific phosphorylation in OGT wild-type and Null cells. Quantitation of the phosphoproteome demonstrated that of 5529 phosphoserine, phosphothreonine, and phosphotyrosine sites, 232 phosphosites were upregulated and 133 downregulated in the absence of O-GlcNAc. Collectively, these data suggest that deletion of OGT has a profound effect on the phosphorylation of cell cycle and DNA damage response proteins. Key events were confirmed by biochemical analyses and demonstrate an increase in the activating autophosphorylation event on ATM (Ser1987) and on ATM's downstream targets p53, H2AX, and Chk2. Together, these data support widespread changes in the phosphoproteome upon removal of O-GlcNAc, suggesting that O-GlcNAc regulates processes such as the cell cycle, genomic stability, and lysosomal biogenesis. All MS data have been deposited in the ProteomeXchange with identifier PXD001153 (http://proteomecentral.proteomexchange.org/dataset/PXD001153).
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