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Identifying chromatin readers using a SILAC-based histone peptide pull-down approach
1Molecular Cancer Research, University Medical Center Utrecht, 3584 CG, Utrecht, The Netherlands. m.vermeulen-3@umcutrecht.nl
Methods in Enzymology
|August 23, 2012
Summary
We developed a SILAC-based method to identify proteins that bind to histone posttranslational modifications (PTMs). This approach helps discover new "chromatin readers" involved in nuclear processes like transcription and DNA repair.
Area of Science:
- Molecular Biology
- Epigenetics
- Proteomics
Background:
- Histone posttranslational modifications (PTMs) are crucial epigenetic marks regulating nuclear processes.
- These PTMs recruit specific proteins, known as chromatin readers, to modulate gene expression and DNA dynamics.
- Identifying these readers is essential for understanding chromatin regulation.
Purpose of the Study:
- To present a generic SILAC-based peptide pull-down method for unbiased identification of histone PTM readers.
- To detail the workflow of this novel approach.
Main Methods:
- Stable Isotope Labeling by Amino acids in Cell culture (SILAC) based peptide pull-down assay.
- Unbiased identification of protein interactors (chromatin readers) for specific histone PTMs.
Main Results:
- The presented method enables the discovery of novel chromatin readers.
- The workflow is detailed for reproducible application.
Conclusions:
- This SILAC-based approach provides a powerful and unbiased tool for identifying histone PTM readers.
- Understanding these reader proteins advances knowledge of epigenetic regulation in transcription, replication, and DNA repair.
