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Evaluation of Histone-Modifying Enzymes in Stem Cell Populations

  • Leanne Stalker
  • Christopher WynderEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 809)

Abstract

The histone demethylases are a relatively novel family of histone-modifying enzymes. Their gene expression suggests that each of the subfamily members may have a discrete role in cell function. The KDM5 family of H3K4 histone demethylases has four members. Each family member has a distinct cellular role, including KDM5a, which is a tumor suppressor (Christensen et al. Cell 128: 1063–1076, 2007); KDM5b, which is an oncogene (Dey et al. Mol Cell Biol 17: 5312–5327, 2008); and KDM5c (Iwase et al. Cell 128: 1077–1088, 2007), which is expressed in terminally differentiated populations. To properly analyze how these enzymes are regulated, we interrogate their bioactivity in ES cells (ESCs) and during neural differentiation of ESCs. We evaluate the bioactivity from both affinity-purified complexes and reconstituted complexes and directly in the cell. These assays have allowed us to define a set of factors that regulate the KDM5 family of histone demethylases.

Key words

ES cells Gene regulation Histone demethylase Neural differentiation Lysine ­demethylation Enzyme kinetics 

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Department of BiochemistryMcMaster UniversityHamiltonCanada
  2. 2.Department of Biochemistry, Schulich School of Medicine and DentistryThe University of Western OntarioLondonCanada

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