Identification of a large protein network involved in epigenetic transmission in replicating DNA of embryonic stem cells.
Pluripotency of embryonic stem cells (ESCs) is maintained by transcriptional activities and chromatin modifying complexes highly organized within the chromatin. Although much effort has been focused on identifying genome-binding sites, little is known on their dynamic association with chromatin across cell divisions. Here, we used a modified version of the iPOND (isolation of proteins at nascent DNA) technology to identify a large protein network enriched at nascent DNA in ESCs. This comprehensive and unbiased proteomic characterization in ESCs reveals that, in addition to the core replication machinery, proteins relevant for pluripotency of ESCs are present at DNA replication sites. In particular, we show that the chromatin remodeller HDAC1-NuRD complex is enriched at nascent DNA. Interestingly, an acute block of HDAC1 in ESCs leads to increased acetylation of histone H3 lysine 9 at nascent DNA together with a concomitant loss of methylation. Consistently, in contrast to what has been described in tumour cell lines, these chromatin marks were found to be stable during cell cycle progression of ESCs. Our results are therefore compatible with a rapid deacetylation-coupled methylation mechanism during the replication of DNA in ESCs that may participate in the preservation of pluripotency of ESCs during replication.
Unit of Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, 17177 Stockholm, Sweden sergi.aranda@crg.eu.
Sergi
S
Aranda
Proteomics Karolinska, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden.
Dorothea
D
Rutishauser
Unit of Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, 17177 Stockholm, Sweden Patrik.Ernfors@ki.se.
Patrik
P
Ernfors
D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D053843:DNA Mismatch Repair; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D053595:Embryonic Stem Cells / Q000378:metabolism*; D044127:Epigenesis, Genetic; D006570:Heterochromatin / Q000378:metabolism; D056284:Histone Deacetylase 1 / Q000378:metabolism; D056245:Mi-2 Nucleosome Remodeling and Deacetylase Complex / Q000378:metabolism; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism; D025941:Protein Interaction Mapping; D060066:Protein Interaction Maps; D044767:Ubiquitin-Protein Ligases
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D006570:Heterochromatin; D009687:Nuclear Proteins; D004247:DNA; D044767:Ubiquitin-Protein Ligases; C117054:Uhrf1 protein, mouse; D056284:Histone Deacetylase 1; D056245:Mi-2 Nucleosome Remodeling and Deacetylase Complex