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Detecting protein and post-translational modifications in single cells with iDentification and qUantification
Yandong Zhang1, Changho Sohn1, Seoyeon Lee1
1Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA, USA.
Communications Biology
|August 5, 2020
Summary
This study introduces DUET, a novel method for detecting and quantifying protein and post-translational modification (PTM) isoforms in single cells. DUET reveals cell-cycle-specific heterogeneities in histone H2B ubiquitination previously hidden by bulk measurements.
Area of Science:
- Proteomics
- Molecular Biology
- Cell Biology
Background:
- Single-cell analysis technologies for transcriptomes are mature, but protein and post-translational modification (PTM) detection in single cells remains challenging due to the inability to amplify proteins.
- Detecting minute protein quantities in single cells requires sensitive and specific methods.
Purpose of the Study:
- To develop a strategy for detecting and quantifying targeted protein and PTM isoforms within individual cells.
- To address the limitations of current protein analysis methods in single-cell studies.
Main Methods:
- Developed a novel strategy called iDentification and qUantification sEparaTion (DUET).
- Proteins from single cells are barcoded with oligonucleotides.
- Barcoded cells are pooled, and proteins/PTM isoforms are separated using bulk gel electrophoresis.
- Abundances are quantified by sequencing the associated oligonucleotides.
Main Results:
- Successfully applied DUET to measure histone H2B and its monoubiquitination isoform (H2Bub) in single yeast cells.
- Revealed significant heterogeneities in H2B ubiquitination levels across different cell-cycle stages.
- Demonstrated that DUET can uncover single-cell variations obscured by traditional ensemble measurements.
Conclusions:
- DUET provides a powerful new approach for single-cell proteomic and PTM analysis.
- The method enables the study of protein isoform heterogeneity at the single-cell level.
- DUET facilitates a deeper understanding of cellular processes by resolving cell-to-cell variations in protein modifications.
