H3K9 methylation is a barrier during somatic cell reprogramming into iPSCs

The induction of pluripotent stem cells (iPSCs) by defined factors is poorly understood stepwise. Here, we show that histone H3 lysine 9 (H3K9) methylation is the primary epigenetic determinant for the intermediate pre-iPSC state, and its removal leads to fully reprogrammed iPSCs. We generated a panel of stable pre-iPSCs that exhibit pluripotent properties but do not activate the core pluripotency network, although they remain sensitive to vitamin C for conversion into iPSCs. Bone morphogenetic proteins (BMPs) were subsequently identified in serum as critical signaling molecules in arresting reprogramming at the pre-iPSC state. Mechanistically, we identified H3K9 methyltransferases as downstream targets of BMPs and showed that they function with their corresponding demethylases as the on/off switch for the pre-iPSC fate by regulating H3K9 methylation status at the core pluripotency loci. Our results not only establish pre-iPSCs as an epigenetically stable signpost along the reprogramming road map, but they also provide mechanistic insights into the epigenetic reprogramming of cell fate.

[1]  J. Inman,et al.  Bmc Cell Biology a Rapid and Sensitive Bioassay for the Simultaneous Measurement of Multiple Bone Morphogenetic Proteins. Identification and Quantification of Bmp4, Bmp6 and Bmp9 in Bovine and Human Serum , 2022 .

[2]  Riitta Lahesmaa,et al.  Copy number variation and selection during reprogramming to pluripotency , 2011, Nature.

[3]  Roland Rad,et al.  Rapid and efficient reprogramming of somatic cells to induced pluripotent stem cells by retinoic acid receptor gamma and liver receptor homolog 1 , 2011, Proceedings of the National Academy of Sciences.

[4]  Jennifer Nichols,et al.  Promotion of Reprogramming to Ground State Pluripotency by Signal Inhibition , 2008, PLoS biology.

[5]  Scott Horowitz,et al.  Structure and Function of Histone H3 Lysine 9 Methyltransferases and Demethylases. , 2011 .

[6]  G. Pan,et al.  The histone demethylases Jhdm1a/1b enhance somatic cell reprogramming in a vitamin-C-dependent manner. , 2011, Cell stem cell.

[7]  Marcos J. Araúzo-Bravo,et al.  Chromatin-Remodeling Components of the BAF Complex Facilitate Reprogramming , 2010, Cell.

[8]  Karl Mechtler,et al.  Methylation of histone H3 lysine 9 creates a binding site for HP1 proteins , 2001, Nature.

[9]  Eran Meshorer,et al.  Global epigenetic changes during somatic cell reprogramming to iPS cells. , 2011, Journal of molecular cell biology.

[10]  Zachary D. Smith,et al.  Reprogramming factor expression initiates widespread targeted chromatin remodeling. , 2011, Cell stem cell.

[11]  N. Benvenisty,et al.  Epigenetic memory and preferential lineage-specific differentiation in induced pluripotent stem cells derived from human pancreatic islet beta cells. , 2011, Cell stem cell.

[12]  A. Childs,et al.  BMP Signaling in the Human Fetal Ovary is Developmentally Regulated and Promotes Primordial Germ Cell Apoptosis , 2010, Stem cells.

[13]  Thomas Lufkin,et al.  Reprogramming of fibroblasts into induced pluripotent stem cells with orphan nuclear receptor Esrrb , 2009, Nature Cell Biology.

[14]  T. Mikkelsen,et al.  Dissecting direct reprogramming through integrative genomic analysis , 2008, Nature.

[15]  P. Park,et al.  Ascorbic acid prevents loss of Dlk1-Dio3 imprinting and facilitates generation of all-iPS cell mice from terminally differentiated B cells , 2012, Nature Genetics.

[16]  Krishanu Saha,et al.  Pluripotency and Cellular Reprogramming: Facts, Hypotheses, Unresolved Issues , 2010, Cell.

[17]  Eric S. Lander,et al.  Chromatin modifying enzymes as modulators of reprogramming , 2012, Nature.

[18]  J. Wrana,et al.  Functional genomics reveals a BMP-driven mesenchymal-to-epithelial transition in the initiation of somatic cell reprogramming. , 2010, Cell stem cell.

[19]  S. Berger,et al.  IDH mutation impairs histone demethylation and results in a block to cell differentiation , 2012, Nature.

[20]  H. Redl,et al.  Vitamin C enhances the generation of mouse and human induced pluripotent stem cells. , 2010, Cell stem cell.

[21]  Brad T. Sherman,et al.  Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources , 2008, Nature Protocols.

[22]  Annick Harel-Bellan,et al.  A subset of the histone H3 lysine 9 methyltransferases Suv39h1, G9a, GLP, and SETDB1 participate in a multimeric complex. , 2010, Molecular cell.

[23]  H. Blau,et al.  Nuclear reprogramming to a pluripotent state by three approaches , 2010, Nature.

[24]  Tomohiro Kono,et al.  Aberrant silencing of imprinted genes on chromosome 12qF1 in mouse induced pluripotent stem cells , 2010, Nature.

[25]  L. David,et al.  Bone Morphogenetic Protein-9 Is a Circulating Vascular Quiescence Factor , 2008, Circulation research.

[26]  R. Lechleider,et al.  Suv39h histone methyltransferases interact with Smads and cooperate in BMP-induced repression , 2004, Oncogene.

[27]  B. Thiers.  Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors , 2008 .

[28]  S. Yamanaka,et al.  Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined Factors , 2006, Cell.

[29]  Hidenori Akutsu,et al.  A small-molecule inhibitor of tgf-Beta signaling replaces sox2 in reprogramming by inducing nanog. , 2009, Cell stem cell.

[30]  D. Sinclair,et al.  The role of nuclear architecture in genomic instability and ageing , 2007, Nature Reviews Molecular Cell Biology.

[31]  Xi Chen,et al.  Jmjd1a and Jmjd2c histone H3 Lys 9 demethylases regulate self-renewal in embryonic stem cells. , 2007, Genes & development.

[32]  Kathrin Plath,et al.  Progress in understanding reprogramming to the induced pluripotent state , 2011, Nature Reviews Genetics.

[33]  Jing Chen,et al.  BMPs functionally replace Klf4 and support efficient reprogramming of mouse fibroblasts by Oct4 alone , 2011, Cell Research.

[34]  Mike J. Mason,et al.  Role of the Murine Reprogramming Factors in the Induction of Pluripotency , 2009, Cell.

[35]  Jonathan M. Monk,et al.  Wdr5 Mediates Self-Renewal and Reprogramming via the Embryonic Stem Cell Core Transcriptional Network , 2011, Cell.

[36]  Jeroen S. van Zon,et al.  Direct cell reprogramming is a stochastic process amenable to acceleration , 2009, Nature.

[37]  Jialiang Liang,et al.  A mesenchymal-to-epithelial transition initiates and is required for the nuclear reprogramming of mouse fibroblasts. , 2010, Cell stem cell.

[38]  Sheng Ding,et al.  Induction of pluripotent stem cells from mouse embryonic fibroblasts by Oct4 and Klf4 with small-molecule compounds. , 2008, Cell stem cell.

[39]  Jing Liu,et al.  Towards an Optimized Culture Medium for the Generation of Mouse Induced Pluripotent Stem Cells* , 2010, The Journal of Biological Chemistry.