Developmentally Programmed 3′ CpG Island Methylation Confers Tissue- and Cell-Type-Specific Transcriptional Activation

ABSTRACT During development, a small but significant number of CpG islands (CGIs) become methylated. The timing of developmentally programmed CGI methylation and associated mechanisms of transcriptional regulation during cellular differentiation, however, remain poorly characterized. Here, we used genome-wide DNA methylation microarrays to identify epigenetic changes during human embryonic stem cell (hESC) differentiation. We discovered a group of CGIs associated with developmental genes that gain methylation after hESCs differentiate. Conversely, erasure of methylation was observed at the identified CGIs during subsequent reprogramming to induced pluripotent stem cells (iPSCs), further supporting a functional role for the CGI methylation. Both global gene expression profiling and quantitative reverse transcription-PCR (RT-PCR) validation indicated opposing effects of CGI methylation in transcriptional regulation during differentiation, with promoter CGI methylation repressing and 3′ CGI methylation activating transcription. By studying diverse human tissues and mouse models, we further confirmed that developmentally programmed 3′ CGI methylation confers tissue- and cell-type-specific gene activation in vivo. Importantly, luciferase reporter assays provided evidence that 3′ CGI methylation regulates transcriptional activation via a CTCF-dependent enhancer-blocking mechanism. These findings expand the classic view of mammalian CGI methylation as a mechanism for transcriptional silencing and indicate a functional role for 3′ CGI methylation in developmental gene regulation.

[1]  David A. Orlando,et al.  CpG island structure and trithorax/polycomb chromatin domains in human cells. , 2012, Genomics.

[2]  Peter A. Jones.  Functions of DNA methylation: islands, start sites, gene bodies and beyond , 2012, Nature Reviews Genetics.

[3]  Zachary D. Smith,et al.  A unique regulatory phase of DNA methylation in the early mammalian embryo , 2012, Nature.

[4]  Yutaka Suzuki,et al.  Contribution of Intragenic DNA Methylation in Mouse Gametic DNA Methylomes to Establish Oocyte-Specific Heritable Marks , 2012, PLoS genetics.

[5]  R. Sandberg,et al.  CTCF-promoted RNA polymerase II pausing links DNA methylation to splicing , 2011, Nature.

[6]  J. Issa,et al.  Epigenetic variation and cellular Darwinism , 2011, Nature Genetics.

[7]  A. Prentice,et al.  Season of Conception in Rural Gambia Affects DNA Methylation at Putative Human Metastable Epialleles , 2010, PLoS genetics.

[8]  Michael Weber,et al.  Targets and dynamics of promoter DNA methylation during early mouse development , 2010, Nature Genetics.

[9]  Robert S. Illingworth,et al.  Orphan CpG Islands Identify Numerous Conserved Promoters in the Mammalian Genome , 2010, PLoS genetics.

[10]  Yi Zhang,et al.  Dnmt3a-Dependent Nonpromoter DNA Methylation Facilitates Transcription of Neurogenic Genes , 2010, Science.

[11]  Allen D. Delaney,et al.  Conserved Role of Intragenic DNA Methylation in Regulating Alternative Promoters , 2010, Nature.

[12]  Anne H. O'Donnell,et al.  Chromatin and sequence features that define the fine and gross structure of genomic methylation patterns. , 2010, Genome research.

[13]  J. Espinosa,et al.  Gene-specific repression of the p53 target gene PUMA via intragenic CTCF-Cohesin binding. , 2010, Genes & development.

[14]  D. Zilberman,et al.  Genome-Wide Evolutionary Analysis of Eukaryotic DNA Methylation , 2010, Science.

[15]  Robert S. Illingworth,et al.  CpG islands influence chromatin structure via the CpG-binding protein Cfp1 , 2010, Nature.

[16]  M. Pellegrini,et al.  Conservation and divergence of methylation patterning in plants and animals , 2010, Proceedings of the National Academy of Sciences.

[17]  Chia-Lin Wei,et al.  Dynamic changes in the human methylome during differentiation. , 2010, Genome research.

[18]  Daniel G. Miller,et al.  Characterization of microRNAs involved in embryonic stem cell states. , 2010, Stem cells and development.

[19]  C. D. Wright,et al.  The Involvement of Proline-Rich 15 in Early Conceptus Development in Sheep1 , 2009, Biology of reproduction.

[20]  Lee E. Edsall,et al.  Human DNA methylomes at base resolution show widespread epigenomic differences , 2009, Nature.

[21]  Lanlan Shen,et al.  Epigenomic profiling indicates a role for DNA methylation in early postnatal liver development. , 2009, Human molecular genetics.

[22]  V. Corces,et al.  CTCF: Master Weaver of the Genome , 2009, Cell.

[23]  Robert S Illingworth,et al.  CpG islands – ‘A rough guide’ , 2009, FEBS letters.

[24]  H. Cedar,et al.  Epigenetic silencing during early lineage commitment , 2009 .

[25]  Israel Steinfeld,et al.  Developmental programming of CpG island methylation profiles in the human genome , 2009, Nature Structural &Molecular Biology.

[26]  B. Lowry.  Faculty Opinions recommendation of Human induced pluripotent stem cells free of vector and transgene sequences. , 2009 .

[27]  Paul Pavlidis,et al.  Histone deacetylase inhibition elicits an evolutionarily conserved self-renewal program in embryonic stem cells. , 2009, Cell stem cell.

[28]  Israel Steinfeld,et al.  BMC Bioinformatics BioMed Central , 2008 .

[29]  A. Riggs,et al.  A human B cell methylome at 100-base pair resolution. , 2009, Proceedings of the National Academy of Sciences of the United States of America.

[30]  Rafael A. Irizarry,et al.  Genome-wide methylation analysis of human colon cancer reveals similar hypo- and hypermethylation at conserved tissue-specific CpG island shores , 2008, Nature Genetics.

[31]  T. Mikkelsen,et al.  Genome-scale DNA methylation maps of pluripotent and differentiated cells , 2008, Nature.

[32]  Michael B. Stadler,et al.  Lineage-specific polycomb targets and de novo DNA methylation define restriction and potential of neuronal progenitors. , 2008, Molecular cell.

[33]  D. Gold,et al.  Gene silencing in cancer by histone H3 lysine 27 trimethylation independent of promoter DNA methylation , 2008, Nature Genetics.

[34]  A. Bird,et al.  DNA methylation landscapes: provocative insights from epigenomics , 2008, Nature Reviews Genetics.

[35]  E. Snyder,et al.  Preparation of autogenic human feeder cells for growth of human embryonic stem cells. , 2008, Current protocols in stem cell biology.

[36]  Jim Stalker,et al.  A Novel CpG Island Set Identifies Tissue-Specific Methylation at Developmental Gene Loci , 2008, PLoS biology.

[37]  Minoru Toyota,et al.  Integrated genetic and epigenetic analysis identifies three different subclasses of colon cancer , 2007, Proceedings of the National Academy of Sciences.

[38]  Mi Zhou,et al.  CTCFBSDB: a CTCF-binding site database for characterization of vertebrate genomic insulators , 2007, Nucleic Acids Res..

[39]  H. Clevers,et al.  Identification of stem cells in small intestine and colon by marker gene Lgr5 , 2007, Nature.

[40]  Pearlly S Yan,et al.  High-throughput methylation profiling by MCA coupled to CpG island microarray. , 2007, Genome research.

[41]  G. Pan,et al.  Whole-genome analysis of histone H3 lysine 4 and lysine 27 methylation in human embryonic stem cells. , 2007, Cell stem cell.

[42]  Li Zhang,et al.  Genome-Wide Profiling of DNA Methylation Reveals a Class of Normally Methylated CpG Island Promoters , 2007, PLoS genetics.

[43]  T. Mikkelsen,et al.  Genome-wide maps of chromatin state in pluripotent and lineage-committed cells , 2007, Nature.

[44]  W. Reik.  Stability and flexibility of epigenetic gene regulation in mammalian development , 2007, Nature.

[45]  A. Bird.  Perceptions of epigenetics , 2007, Nature.

[46]  T. Mikkelsen,et al.  Systematic discovery of regulatory motifs in conserved regions of the human genome, including thousands of CTCF insulator sites , 2007, Proceedings of the National Academy of Sciences.

[47]  Michael Q. Zhang,et al.  Analysis of the Vertebrate Insulator Protein CTCF-Binding Sites in the Human Genome , 2007, Cell.

[48]  Michael B. Stadler,et al.  Distribution, silencing potential and evolutionary impact of promoter DNA methylation in the human genome , 2007, Nature Genetics.

[49]  C Anthony Blau,et al.  A Comparison of NIH‐Approved Human ESC Lines , 2006, Stem cells.

[50]  J. Rogers,et al.  DNA methylation profiling of human chromosomes 6, 20 and 22 , 2006, Nature Genetics.

[51]  M. Pellegrini,et al.  Genome-wide High-Resolution Mapping and Functional Analysis of DNA Methylation in Arabidopsis , 2006, Cell.

[52]  M. Nakao,et al.  CTCF-dependent chromatin insulator is linked to epigenetic remodeling. , 2006, Molecular cell.

[53]  James A. Cuff,et al.  A Bivalent Chromatin Structure Marks Key Developmental Genes in Embryonic Stem Cells , 2006, Cell.

[54]  P. Chuang,et al.  Mice Deficient in the Fused Homolog Do Not Exhibit Phenotypes Indicative of Perturbed Hedgehog Signaling during Embryonic Development , 2005, Molecular and Cellular Biology.

[55]  U. Bunz.  How Are Alkynes Scrambled? , 2005, Science.

[56]  Hiroki Nagase,et al.  Association of tissue-specific differentially methylated regions (TDMs) with differential gene expression. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[57]  M. Washington,et al.  Gene expression profile analysis of mouse colon embryonic development , 2005, Genesis.

[58]  J. Mcwhir,et al.  Immortalized Fibroblast‐Like Cells Derived from Human Embryonic Stem Cells Support Undifferentiated Cell Growth , 2004, Stem cells.

[59]  Shan-Mei Xu,et al.  Palmitoylation is required for the production of a soluble multimeric Hedgehog protein complex and long-range signaling in vertebrates. , 2004, Genes & development.

[60]  K A Baggerly,et al.  Sensitive and quantitative universal Pyrosequencing methylation analysis of CpG sites. , 2003, BioTechniques.

[61]  J. Herman,et al.  Heterozygous disruption of Hic1 predisposes mice to a gender-dependent spectrum of malignant tumors , 2003, Nature Genetics.

[62]  Kyoung-Mee Kim,et al.  Methylation reveals a niche: stem cell succession in human colon crypts , 2002, Oncogene.

[63]  Daiya Takai,et al.  Comprehensive analysis of CpG islands in human chromosomes 21 and 22 , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[64]  J. Mesirov,et al.  Chemosensitivity prediction by transcriptional profiling , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[65]  W. Reik,et al.  Epigenetic Reprogramming in Mammalian Development , 2001, Science.

[66]  D. Leprince,et al.  Identification in the Human Candidate Tumor Suppressor GeneHIC-1 of a New Major Alternative TATA-less Promoter Positively Regulated by p53* , 2001, The Journal of Biological Chemistry.

[67]  G. Felsenfeld,et al.  Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene , 2000, Nature.

[68]  Margaret A. Johns,et al.  Mice deficient in the candidate tumor suppressor gene Hic1 exhibit developmental defects of structures affected in the Miller-Dieker syndrome. , 2000, Human molecular genetics.

[69]  S. Baylin,et al.  p53 activates expression of HIC-1, a new candidate tumour suppressor gene on 17p13.3 , 1995, Nature Genetics.

[70]  Rudolf Jaenisch,et al.  Targeted mutation of the DNA methyltransferase gene results in embryonic lethality , 1992, Cell.

[71]  S. Tapscott,et al.  De novo methylation of the MyoD1 CpG island during the establishment of immortal cell lines. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[72]  Y. Benjamini,et al.  More powerful procedures for multiple significance testing. , 1990, Statistics in medicine.

[73]  Peter A. Jones,et al.  Multiple new phenotypes induced in 10T 1 2 and 3T3 cells treated with 5-azacytidine , 1979, Cell.

[74]  R Holliday,et al.  DNA modification mechanisms and gene activity during development , 1975, Science.

[75]  S. Cole,et al.  Sequences Human Induced Pluripotent Stem Cells Free of Vector and Transgene , 2012 .

[76]  D. Leprince,et al.  HIC1 (Hypermethylated in Cancer 1) epigenetic silencing in tumors. , 2009, The international journal of biochemistry & cell biology.

[77]  S. Henikoff,et al.  Genome-wide analysis of Arabidopsis thaliana DNA methylation uncovers an interdependence between methylation and transcription , 2007, Nature Genetics.

[78]  J. Issa,et al.  Optimizing annealing temperature overcomes bias in bisulfite PCR methylation analysis. , 2007, BioTechniques.

[79]  P. Jones,et al.  The DNA methylation paradox. , 1999, Trends in genetics : TIG.

[80]  Arthur D. Riggs,et al.  X inactivation, differentiation, and DNA methylation. , 1975, Cytogenetics and cell genetics.

[81]  Timothy J. Durham,et al.  "Systematic" , 1966, Comput. J..

[82]  Nianxiang Zhang,et al.  Widespread and Tissue Specific Age-related Dna Methylation Material Supplemental Related Content a Hallmark of Cancer Age-dependent Dna Methylation of Genes That Are Suppressed in Stem Cells Is , 2022 .

[83]  Richard M Myers,et al.  Network: from Single Conserved Sites to Genome-wide Repertoire Comparative Genomics Modeling of the Nrsf/rest Repressor Material Supplemental , 2022 .