Quantification of SAHA-Dependent Changes in Histone Modifications Using Data-Independent Acquisition Mass Spectrometry

Histone
post-translational modifications (PTMs) are important regulators
of chromatin structure and gene expression. Quantitative analysis
of histone PTMs by mass spectrometry remains extremely challenging
due to the complex and combinatorial nature of histone PTMs. The most
commonly used mass spectrometry-based method for high-throughput histone
PTM analysis is data-dependent acquisition (DDA). However, stochastic
precursor selection and dependence on MS1 ions for quantification
impede comprehensive interrogation of histone PTM states using DDA
methods. To overcome these limitations, we utilized a data-independent
acquisition (DIA) workflow that provides superior run-to-run consistency
and postacquisition flexibility in comparison to DDA methods. In addition,
we developed a novel DIA-based methodology to quantify isobaric, co-eluting
histone peptides that lack unique MS2 transitions. Our method enabled
deconvolution and quantification of histone PTMs that are otherwise
refractory to quantitation, including the heavily acetylated tail
of histone H4. Using this workflow, we investigated the effects of
the histone deacetylase inhibitor SAHA (suberoylanilide hydroxamic
acid) on the global histone PTM state of human breast cancer MCF7
cells. A total of 62 unique histone PTMs were quantified, revealing
novel SAHA-induced changes in acetylation and methylation of histones
H3 and H4.