canonical wnt signaling is required for development of embryonic stem cell-derived mesoderm lindsley...
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Canonical Wnt signaling is required for development of embryonic st
em cell-derived mesoderm
Lindsley R.C. et al, Development (2006)
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Keller G., Genes Dev.(2005) Tam P.P.L. and behringer R.R., curr. Opin. Genet. Dev.(2003)
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• Generation of the primitive streak is regulated by multiple pathways, including Nodal, Wnt and Bmp
• Canonical Wnt signaling is absolutely required for primitive streak formation (Wnt3, LRP5/6)
• Wnt signaling can induce expression of at least one primitive streak gene (brachyury) and can antagonize neuronal differentiation
Rossant J. and Tam P. P. L., Dev. Cell (2006)
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ES cells differentiation
• Without discernible axes and lack tissue components
• Can study the mechanisms of signaling pathways during discrete developmental transitions in the absence of asymmetric embryonic structures and signaling centers
• Characterization of Wnt signaling in regulating features of in vitro ES cell differentiation that may be analogous to primitive streak formation in vivo.
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Activation of canonical Wnt signaling during early ES cell differentiation
In the pre-streak and early gastrula-stage
Kemp C. et al, Dev. Dyn. (2005)
Keller G., Genes Dev.(2005)
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• independent of extrinsic serum-derived factors• initiated by either cell autonomous
mechanisms or by intercellular signals present
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Wnt signaling is required for generation of Flk1+ mesoderm
Flk1 - VEGF receptor expressed by multipotent mesoderm cellsNOGGIN - Bmp2/4/7 inhibitor
Keller G., Genes Dev.(2005)
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Wnt-dependent expression of genes associated with primitive streak, EMT, endoderm and mesoderm
early primitive streak-stage embryo
epithelial-mesenchymal transition mes/endoderm mesoderm ES cell
associated gene
Dkk1 treatment causes a global block at the ES cell equivalent of primitive streak formation
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Early inhibition of canonical Wnt signaling enhances longterm Neuronal differentiation
Neuroectoderm associated genes
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Canonical Wnt signaling is required forgeneration of ES cell-derived mesoderm
Transcriptional factors for hematopoietic development
Cadio-specific genes
methylcellulose colonyformingassays
- 12day- 16day
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E-cadherin -> epithelial markerFibronectin -> mesenchymal marker
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Stabilized -catenin is insufficient for induction ofprimitive streak-associated gene expression
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Soluble factors can restore mesoderm generation
in the presence of Dkk1
CRIPTO -> EGF-CFC factor, cardiomyocyte developmentFST -> Follistatin
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Wnt and Bmp signaling act cooperatively to regulate brachyury expression
A2lox.scat cells
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• We demonstrate a requirement for Wnt signaling during the earliest steps of mesendodermal differentiation of ES cells
• How Wnt signaling regulates gene expression– Wnt pathway activation could act autonomously and directly to in
duce target gene expression through the -catenin-dependent conversion
– Wnt signaling regulate the threshold or stability of gene transcription induced by other pathways
• ES cells may provide a useful and experimentally tractable model system to define complex signaling interactions at the cellular and molecular levels– Evx1 -> b-catenin deficient embryo(3.2 fold reduced) ES cell (40-fold reduced)