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Chromatin Immunoprecipitation from Human Embryonic Stem Cells
Published on: July 22, 2008
Changes in the distributions and dynamics of polycomb repressive complexes during embryonic stem cell differentiation
Xiaojun Ren1, Claudius Vincenz, Tom K Kerppola
1Howard Hughes Medical Institute and Department of Biological Chemistry, University of Michigan, Ann Arbor, MI 48109-0650. kerppola@umich.edu.
Molecular and Cellular Biology
|March 5, 2008
Summary
Polycomb group (PcG) proteins maintain cell identity. During embryonic stem cell differentiation, CBX protein complexes change localization and dynamics, impacting gene regulation and epigenetic reprogramming.
Area of Science:
- Epigenetics
- Molecular Biology
- Cell Biology
Background:
- Polycomb group (PcG) proteins are crucial for maintaining cell identity through gene repression.
- Embryonic stem (ES) cell differentiation involves significant genome-wide changes in gene expression targeted by PcG proteins.
Purpose of the Study:
- To investigate how differentiation and protein interactions affect the localization and dynamics of CBX family PcG proteins.
- To understand the role of specific protein domains in CBX protein behavior during ES cell differentiation.
Main Methods:
- Utilized fluorescence imaging techniques to observe CBX protein localization and mobility in mouse ES cells.
- Employed bimolecular fluorescence complementation (BiFC) analysis to detect CBX-RING1B complexes.
- Generated CBX protein variants with deletions in the chromobox or chromodomain for functional analysis.
Main Results:
- CBX proteins displayed distinct distributions and mobilities in ES cells, often enriched in Polycomb bodies.
- Polycomb body formation did not impede CBX protein mobility, and these bodies dispersed upon differentiation.
- CBX protein mobility increased during initial differentiation and decreased as differentiation progressed.
- Deletion of the chromobox domain, crucial for RING1B interaction, prevented CBX protein immobilization.
- Deletion of the chromodomain, involved in histone H3 binding, had minimal impact on CBX protein dynamics.
Conclusions:
- Epigenetic reprogramming during ES cell differentiation is linked to global alterations in the subnuclear distribution and dynamics of CBX protein complexes.
- The interaction between CBX proteins and RING1B, mediated by the chromobox domain, is critical for regulating CBX protein dynamics during differentiation.
- CBX protein dynamics are sensitive to differentiation status and protein complex formation, reflecting changes in epigenetic regulation.
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