Related Experiment Video
Updated: Jun 7, 2026
11:11
Identification of Novel CK2 Kinase Substrates Using a Versatile Biochemical Approach
Published on: February 21, 2019
Single-cell profiling of kinase substrate phosphorylation by single-molecule imaging
Takuya Hidaka1,2, Ryotaro Motoya2,3, Gao Jintian2,4
1RIKEN Center for Biosystems Dynamics, Suita, Osaka, Japan.
Plos One
|June 5, 2026
Summary
We developed a sensitive single-cell system to profile protein phosphorylation, revealing cell-to-cell variability in kinase signaling. This method overcomes challenges of low phosphoprotein abundance for better understanding cellular processes.
Area of Science:
- Cellular Biology
- Biochemistry
- Molecular Biology
Background:
- Protein phosphorylation is crucial for cellular processes.
- Analyzing phosphorylation at the single-cell level is difficult due to low phosphoprotein abundance.
Purpose of the Study:
- To develop a sensitive system for profiling kinase substrate phosphorylation in individual cells.
- To investigate cell-to-cell variability in kinase signaling.
Main Methods:
- Single-cell proteome fluorescence labeling
- Immunoprecipitation targeting specific phospho-motifs
- Miniaturized SDS-PAGE
- Single-molecule detection via light-sheet fluorescence microscopy
Main Results:
- Successfully profiled phosphorylation of casein kinase 2 (CK2) substrates in HeLa cells.
- Confirmed treatment-induced accumulation of phosphorylated CK2 substrates.
- Revealed heterogeneous phosphorylation patterns at the single-cell level, obscured in bulk analysis.
Conclusions:
- The developed platform enables sensitive, single-cell profiling of kinase substrate phosphorylation.
- Highlights the importance of single-cell analysis for understanding heterogeneous signaling dynamics.
- Provides a foundation for dissecting cell-to-cell variability in signaling pathways.
