Profiling post-translational modifications of histones in neural differentiation of embryonic stem cells using liquid chromatography–mass spectrometry

• Chemical derivatization of lysine to improve HPLC separation of tryptic peptides.
• An integrated HPLC–MS/MS for histone post-translational modifications (HPTMs).
• Profiling of HPTMs in neural differentiation of embryonic stem cells (ESCs).
• Identification of novel changes of combinational HPTMs in cells differentiation.

The neural differentiation of embryonic stem cells (ESCs) is of great significance for understanding of the mechanism of diseases. Histone post-translational modifications (HPTMs) play a key role in the regulation of ESCs differentiation. Here, we combined the stable isotope chemical derivatization with nano-HPLC-mass spectrometry (MS) for comprehensive analysis and quantification of histone post-translational modifications (HPTMs) in mouse embryonic stem cells (mESCs) and neural progenitor cells (mNPCs) that was derived from ESCs. We identified 85 core HPTM sites in ESCs and 78HPTM sites in NPCs including some novel lysine modifications. Our quantitative analysis results further revealed the changes of HPTMs from ESCs to NPCs and suggested effect of combinational HPTMs in the differentiation. This study demonstrates that HPLC–MS-based quantitative proteomics has a considerable advantage on quantification of combinational PTMs and expands our understanding of HPTMs in the differentiation.