Defining early lineage specification of human embryonic stem cells by the orchestrated balance of canonical Wnt/β-catenin, Activin/Nodal and BMP signaling

The canonical Wnt/β-catenin signaling has remarkably diverse roles in embryonic development, stem cell self-renewal and cancer progression. Here, we show that stabilized expression of β-catenin perturbed human embryonic stem (hES)-cell self-renewal, such that up to 80% of the hES cells developed into the primitive streak (PS)/mesoderm progenitors, reminiscent of early mammalian embryogenesis. The formation of the PS/mesoderm progenitors essentially depended on the cooperative action of β-catenin together with Activin/Nodal and BMP signaling pathways. Intriguingly, blockade of BMP signaling completely abolished mesoderm generation, and induced a cell fate change towards the anterior PS progenitors. The PI3-kinase/Akt, but not MAPK, signaling pathway had a crucial role in the anterior PS specification, at least in part, by enhancing β-catenin stability. In addition, Activin/Nodal and Wnt/β-catenin signaling synergistically induced the generation and specification of the anterior PS/endoderm. Taken together, our findings clearly demonstrate that the orchestrated balance of Activin/Nodal and BMP signaling defines the cell fate of the nascent PS induced by canonical Wnt/β-catenin signaling in hES cells.

[1]  J. Miyazaki,et al.  Quantitative expression of Oct-3/4 defines differentiation, dedifferentiation or self-renewal of ES cells , 2000, Nature Genetics.

[2]  P. Greengard,et al.  GSK-3-selective inhibitors derived from Tyrian purple indirubins. , 2003, Chemistry & biology.

[3]  Wei Li,et al.  Extracellular signal-regulated kinase 2 is necessary for mesoderm differentiation , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[4]  Y. Sasai,et al.  Endoderm induction by the organizer‐secreted factors chordin and noggin in Xenopus animal caps. , 1996, The EMBO journal.

[5]  G. Evan,et al.  A modified oestrogen receptor ligand-binding domain as an improved switch for the regulation of heterologous proteins. , 1995, Nucleic acids research.

[6]  C. Niehrs,et al.  Dickkopf-1 is a member of a new family of secreted proteins and functions in head induction , 1998, Nature.

[7]  Heiko Lickert,et al.  Formation of multiple hearts in mice following deletion of beta-catenin in the embryonic endoderm. , 2002, Developmental cell.

[8]  Tsutomu Chiba,et al.  Induction and monitoring of definitive and visceral endoderm differentiation of mouse ES cells , 2005, Nature Biotechnology.

[9]  E. Fuchs,et al.  Links between signal transduction, transcription and adhesion in epithelial bud development , 2003, Nature.

[10]  Randall T Moon,et al.  WNT and beta-catenin signalling: diseases and therapies. , 2004, Nature reviews. Genetics.

[11]  Gordon Keller,et al.  Differentiation of Embryonic Stem Cells to Clinically Relevant Populations: Lessons from Embryonic Development , 2008, Cell.

[12]  J. Brickman,et al.  Hex acts with beta-catenin to regulate anteroposterior patterning via a Groucho-related co-repressor and Nodal. , 2006, Development.

[13]  A. Pyle,et al.  Defining the Role of Wnt/β‐Catenin Signaling in the Survival, Proliferation, and Self‐Renewal of Human Embryonic Stem Cells , 2005, Stem cells.

[14]  G. Keller,et al.  Wnt, activin, and BMP signaling regulate distinct stages in the developmental pathway from embryonic stem cells to blood. , 2008, Cell stem cell.

[15]  Austin G Smith,et al.  Functional gene screening in embryonic stem cells implicates Wnt antagonism in neural differentiation , 2002, Nature Biotechnology.

[16]  Ariel J. Levine,et al.  TGFbeta/activin/nodal signaling is necessary for the maintenance of pluripotency in human embryonic stem cells. , 2005, Development.

[17]  R. Derynck,et al.  Smad-dependent and Smad-independent pathways in TGF-beta family signalling. , 2003, Nature.

[18]  Gordon Keller,et al.  Development of definitive endoderm from embryonic stem cells in culture , 2004, Development.

[19]  A. Bradley,et al.  Mice deficient for BMP2 are nonviable and have defects in amnion/chorion and cardiac development. , 1996, Development.

[20]  Norio Nakatsuji,et al.  Efficient establishment of human embryonic stem cell lines and long-term maintenance with stable karyotype by enzymatic bulk passage. , 2006, Biochemical and biophysical research communications.

[21]  A. McMahon,et al.  T (Brachyury) is a direct target of Wnt3a during paraxial mesoderm specification. , 1999, Genes & development.

[22]  Patrick J. Paddison,et al.  Wnt and TGF-β signaling are required for the induction of an in vitro model of primitive streak formation using embryonic stem cells , 2006, Proceedings of the National Academy of Sciences.

[23]  Dongxin Zhao,et al.  WNT/β-catenin pathway up-regulates Stat3 and converges on LIF to prevent differentiation of mouse embryonic stem cells , 2006 .

[24]  R. McKay,et al.  New cell lines from mouse epiblast share defining features with human embryonic stem cells , 2007, Nature.

[25]  Stephen S. Taylor,et al.  GSK-3 inhibitors induce chromosome instability , 2007, BMC Cell Biology.

[26]  Sheng Ding,et al.  Long-term self-renewal and directed differentiation of human embryonic stem cells in chemically defined conditions. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[27]  Dongxin Zhao,et al.  WNT/beta-catenin pathway up-regulates Stat3 and converges on LIF to prevent differentiation of mouse embryonic stem cells. , 2006, Developmental biology.

[28]  D. Norris,et al.  Cell fate decisions within the mouse organizer are governed by graded Nodal signals. , 2003, Genes & development.

[29]  N. Nakatsuji,et al.  Apoptosis and differentiation of human embryonic stem cells induced by sustained activation of c-Myc , 2007, Oncogene.

[30]  J. Thiery.  Epithelial–mesenchymal transitions in tumour progression , 2002, Nature Reviews Cancer.

[31]  J. Gurdon,et al.  Morphogen gradient interpretation , 2001, Nature.

[32]  A. Hall,et al.  Cdc42 regulates GSK-3β and adenomatous polyposis coli to control cell polarity , 2003, Nature.

[33]  T. Lepage,et al.  Zebrafish endoderm formation is regulated by combinatorial Nodal, FGF and BMP signalling , 2006, Development.

[34]  T. Blundell,et al.  Functional specificity of the Xenopus T-domain protein Brachyury is conferred by its ability to interact with Smad1. , 2005, Developmental cell.

[35]  P. Greengard,et al.  Maintenance of pluripotency in human and mouse embryonic stem cells through activation of Wnt signaling by a pharmacological GSK-3-specific inhibitor , 2004, Nature Medicine.

[36]  G. Keller,et al.  Embryonic stem cell differentiation: emergence of a new era in biology and medicine. , 2005, Genes & development.

[37]  K. Mostov,et al.  NH2-terminal Deletion of β-Catenin Results in Stable Colocalization of Mutant β-Catenin with Adenomatous Polyposis Coli Protein and Altered MDCK Cell Adhesion , 1997, The Journal of cell biology.

[38]  Yoshiakira Kanai,et al.  Depletion of definitive gut endoderm in Sox17-null mutant mice. , 2002, Development.

[39]  P. Lásló,et al.  Multilineage Transcriptional Priming and Determination of Alternate Hematopoietic Cell Fates , 2006, Cell.

[40]  D. Loebel,et al.  Gene function in mouse embryogenesis: get set for gastrulation , 2007, Nature Reviews Genetics.

[41]  C. Gilles,et al.  Epithelial-mesenchymal transition process in human embryonic stem cells cultured in feeder-free conditions. , 2007, Molecular human reproduction.

[42]  E. Fuchs,et al.  Loss of a quiescent niche but not follicle stem cells in the absence of bone morphogenetic protein signaling , 2007, Proceedings of the National Academy of Sciences.

[43]  N. Copeland,et al.  Cloning, expression analysis, and chromosomal localization of murine and human homologues of a Xenopus Mix gene , 2000, Developmental dynamics : an official publication of the American Association of Anatomists.

[44]  H Ueno,et al.  Fibroblast growth factor receptor is required for in vivo cardiac myocyte proliferation at early embryonic stages of heart development. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[45]  D. Solter,et al.  Stabilization of β-catenin in the mouse zygote leads to premature epithelial-mesenchymal transition in the epiblast , 2004, Development.

[46]  Michael Kyba,et al.  Canonical Wnt signaling is required for development of embryonic stem cell-derived mesoderm , 2006, Development.

[47]  M. Trotter,et al.  Derivation of pluripotent epiblast stem cells from mammalian embryos , 2007, Nature.

[48]  E. Kroon,et al.  Efficient differentiation of human embryonic stem cells to definitive endoderm , 2005, Nature Biotechnology.

[49]  Gordon Keller,et al.  BMP-4 is required for hepatic specification of mouse embryonic stem cell–derived definitive endoderm , 2006, Nature Biotechnology.

[50]  M. Okabe,et al.  Activation of Akt signaling is sufficient to maintain pluripotency in mouse and primate embryonic stem cells , 2006, Oncogene.

[51]  Ariel J. Levine,et al.  TGFβ/activin/nodal signaling is necessary for the maintenance of pluripotency in human embryonic stem cells , 2005 .

[52]  Ossama Tawfik,et al.  BMP signaling inhibits intestinal stem cell self-renewal through suppression of Wnt–β-catenin signaling , 2004, Nature Genetics.

[53]  Ajamete Kaykas,et al.  WNT and β-catenin signalling: diseases and therapies , 2004, Nature Reviews Genetics.