Pioneer factor interactions and unmethylated CpG dinucleotides mark silent tissue-specific enhancers in embryonic stem cells.
Recent studies have suggested that, in ES cells, inactive genes encoding early developmental regulators possess bivalent histone modification domains and are therefore poised for activation. However, bivalent domains were not observed at typical tissue-specific genes. Here, we show that windows of unmethylated CpG dinucleotides and putative pioneer factor interactions mark enhancers for at least some tissue-specific genes in ES cells. The unmethylated windows expand in cells that express the gene and contract, disappear, or remain unchanged in nonexpressing tissues. However, in ES cells, they do not always coincide with common histone modifications. Genomic footprinting and chromatin immunoprecipitation demonstrated that transcription factor binding underlies the unmethylated windows at enhancers for the Ptcra and Alb1 genes. After stable integration of premethylated Ptcra enhancer constructs into the ES cell genome, the unmethylated windows readily appeared. In contrast, the premethylated constructs remained fully methylated and silent after introduction into Ptcra-expressing thymocytes. These findings provide initial functional support for a model in which pioneer factor interactions in ES cells promote the assembly of a chromatin structure that is permissive for subsequent activation, and in which differentiated tissues lack the machinery required for gene activation when these ES cell marks are absent. The enhancer marks may therefore represent important features of the pluripotent state.
Xu
Jian
J
Howard Hughes Medical Institute, Molecular Biology Institute, University of California, Los Angeles, CA 90095, USA.
Pope
Scott D
SD
Jazirehi
Ali R
AR
Attema
Joanne L
JL
Papathanasiou
Peter
P
Watts
Jason A
JA
Zaret
Kenneth S
KS
Weissman
Irving L
IL
Smale
Stephen T
ST
D000418:Albumins / Q000235:genetics; D000818:Animals; D002460:Cell Line; D018899:CpG Islands / Q000235:genetics*; D019175:DNA Methylation; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D020868:Gene Silencing; D005819:Genetic Markers; D006410:Hematopoiesis; D006657:Histones / Q000378:metabolism; D006801:Humans; D018664:Interleukin-12 / Q000235:genetics; D008562:Membrane Glycoproteins / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000235:genetics*; D011485:Protein Binding; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics; D016415:Sequence Alignment; D013950:Thymus Gland / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics
D000418:Albumins; D005819:Genetic Markers; D006657:Histones; D008562:Membrane Glycoproteins; D009711:Nucleotides; D016693:Receptors, Antigen, T-Cell, alpha-beta; D014157:Transcription Factors; C090377:pre-T cell receptor alpha; D018664:Interleukin-12