Protocol

Eukaryotic Transcriptional and Post-Transcriptional Gene Expression Regulation

Volume 1507 of the series Methods in Molecular Biology pp 153-168

Date:

Profiling Changes in Histone Post-translational Modifications by Top-Down Mass Spectrometry

  • Mowei ZhouAffiliated withEnvironmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory
  • , Si WuAffiliated withEnvironmental Molecular Sciences Laboratory, Pacific Northwest National LaboratoryDepartment of Chemistry and Biochemistry, University of Oklahoma
  • , David L. StenoienAffiliated withEnvironmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory
  • , Zhaorui ZhangAffiliated withEnvironmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory
  • , Lanelle ConnollyAffiliated withDepartment of Biochemistry and Biophysics, Center for Genome Research and Biocomputing, Oregon State University
  • , Michael FreitagAffiliated withDepartment of Biochemistry and Biophysics, Center for Genome Research and Biocomputing, Oregon State University
  • , Ljiljana Paša-TolićAffiliated withEnvironmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory Email author 

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Abstract

Top-down mass spectrometry is a valuable tool for understanding gene expression through characterization of combinatorial histone post-translational modifications (i.e., histone code). In this protocol, we describe a top-down workflow that employs liquid chromatography (LC) coupled to mass spectrometry (MS), for fast global profiling of changes in histone proteoforms, and apply LCMS top-down approach for comparative analysis of a wild-type and a mutant fungal species. The proteoforms exhibiting differential abundances can be subjected to further targeted studies by other MS or orthogonal (e.g., biochemical) assays. This method can be generally adapted for screening of changes in histone modifications between samples such as wild type vs. mutant or healthy vs. diseased.

Key words

Histone Post-translational modification Liquid chromatography Mass spectrometry Top-down Screening