Wnt/beta-catenin signaling controls Mespo expression to regulate segmentation during Xenopus somitogenesis

The vertebral column is derived from somites, which are transient segments of the paraxial mesoderm that are present in developing vertebrates. The strict spatial and temporal regulation of somitogenesis is of crucial developmental importance. Signals such as Wnt and FGF play roles in somitogenesis, but details regarding how Wnt signaling functions in this process remain unclear. In this study, we report that Wnt/beta-catenin signaling regulates the expression of Mespo, a basic-helix-loop-helix (bHLH) gene critical for segmental patterning in Xenopus somitogenesis. Transgenic analysis of the Mespo promoter identifies Mespo as a direct downstream target of Wnt/beta-catenin signaling pathway. We also demonstrate that activity of Wnt/beta-catenin signaling in somitogenesis can be enhanced by the PI3-K/AKT pathway. Our results illustrate that Wnt/beta-catenin signaling in conjunction with PI3-K/AKT pathway plays a key role in controlling development of the paraxial mesoderm.     

Non-canonical Wnt signaling in Xenopus: regulation of axis formation and gastrulation

Wnt proteins form a family of secreted glycoproteins that are involved in different developmental processes such as differentiation, proliferation, cell migration and cell polarity. To exert its function, Wnt proteins activate different intracellular signaling cascades. Whereas the canonical, Wnt/beta-catenin pathway is well characterized, less is known about the function of non-canonical Wnt pathways in vertebrates. I here summarize recent findings implicating important roles for Wnt/Ca(2+) and Wnt/JNK signaling during different aspects of early Xenopus laevis development, namely axis formation and gastrulation movements.     

Dissecting signaling pathways that govern self-renewal of rabbit embryonic stem cells

The pluripotency and self-renewal of embryonic stem cells (ESC) are regulated by a variety of cytokines/growth factors with some species differences. We reported previously that rabbit ESC (rESC) are more similar to primate ESC than to mouse ESC. However, the signaling pathways that regulate rESC self-renewal had not been identified. Here we show that inhibition of the transforming growth factor beta (TGFbeta), fibroblast growth factor (FGF), and canonical Wnt/beta-catenin (Wnt) pathways results in enhanced differentiation of rESC accompanied by down-regulation of Smad2/3 phosphorylation and beta-catenin expression and up-regulation of phosphorylation of Smad1 and beta-catenin. These results imply that the TGFbeta, FGF, and Wnt pathways are required for rESC self-renewal. Inhibition of the MAPK/ERK and PI3K/AKT pathways, which lie downstream of the FGF pathway, led to differentiation of rESC accompanied by down-regulation of phosphorylation of ERK1/2 or AKT, respectively. Long-term self-renewal of rESC could be achieved by adding a mixture of TGFbeta ligands (activin A, Nodal, or TGFbeta1) plus basic FGF (bFGF) and Noggin in the absence of serum and feeder cells. Our findings also suggest that there is a regulatory network consisting of the FGF, Wnt, and TGFbeta pathways that controls rESC pluripotency and self-renewal. We conclude that bFGF controls the stem cell properties of rESC both directly and indirectly through TGFbeta or other pathways, whereas the effect of Wnt on rESC might be mediated by the TGFbeta pathway.     

Wnt, activin, and BMP signaling regulate distinct stages in the developmental pathway from embryonic stem cells to blood

The embryonic stem cell differentiation system was used to define the roles of the Activin/Nodal, BMP, and canonical Wnt signaling pathways at three distinct developmental stages during hematopoietic ontogeny: induction of a primitive streak-like population, formation of Flk1(+) mesoderm, and induction of hematopoietic progenitors. Activin/Nodal and Wnt, but not BMP, signaling are required for the induction of the primitive streak. Although BMP is not required for primitive streak induction, it displays a strong posteriorizing effect on this population. All three signaling pathways regulate induction of Flk1(+) mesoderm. The specification of Flk1(+) mesoderm to the hematopoietic lineages requires VEGF and Wnt, but not BMP or Activin/Nodal signaling. Specifically, Wnt signaling is essential for commitment of the primitive erythroid, but not the definitive lineages. These findings highlight dynamic changes in signaling requirements during blood cell development and identify a role for Wnt signaling in the establishment of the primitive erythroid lineage.     

Neural tissue in ascidian embryos is induced by FGF9/16/20, acting via a combination of maternal GATA and Ets transcription factors

In chordates, formation of neural tissue from ectodermal cells requires an induction. The molecular nature of the inducer remains controversial in vertebrates. Here, using the early neural marker Otx as an entry point, we dissected the neural induction pathway in the simple embryos of Ciona intestinalis. We first isolated the regulatory element driving Otx expression in the prospective neural tissue, showed that this element directly responds to FGF signaling and that FGF9/16/20 acts as an endogenous neural inducer. Binding site analysis and gene loss of function established that FGF9/16/20 induces neural tissue in the ectoderm via a synergy between two maternal response factors. Ets1/2 mediates general FGF responsiveness, while the restricted activity of GATAa targets the neural program to the ectoderm. Thus, our study identifies an endogenous FGF neural inducer and its early downstream gene cascade. It also reveals a role for GATA factors in FGF signaling.     

Novel Daple-like protein positively regulates both the Wnt/beta-catenin pathway and the Wnt/JNK pathway in Xenopus

Wnt signaling pathways are essential in various developmental processes including differentiation, proliferation, cell migration, and cell polarity. Wnt proteins execute their multiple functions by activating distinct intracellular signaling cascades, although the mechanisms underlying this activation are not fully understood. We identified a novel Daple-like protein in Xenopus and named it xDal (Xenopus Daple-like). As with Daple, xDal contains several leucine zipper-like regions (LZLs) and a putative PDZ domain-binding motif, and can interact directly with the dishevelled protein. In contrast to mDaple, injection of xDal mRNA into the dorso-vegetal blastomere does not induce ventralization and acted synergistically with xdsh in secondary axis induction. XDal also induced expression of siamois and xnr-3, suggesting that XDal functions as a positive regulator of the Wnt/beta-catenin pathway. Injection of xDal mRNA into the dorso-animal blastomere, however, induced gastrulation-defective phenotypes in a dose-dependent manner. In addition, xDal inhibited activin-induced elongation of animal caps and enhanced c-jun phosphorylation. Based on these findings, xDal is also thought to function in the Wnt/JNK pathway. Moreover, functional domain analysis with several deletion mutants indicated that xDal requires both a putative PDZ domain-binding motif and at least one LZL for its activity. These findings with xDal will provide new information on the Wnt signaling pathways.     

Antagonistic crosstalk of Wnt/β-catenin/Bmp signaling within the Apical Ectodermal Ridge (AER) regulates interdigit formation

Digit and interdigit (D/ID) development is one of the important research fields in molecular developmental biology. Interdigital cell death (ICD) is a morphogenetic event which has been considered as an essential process for D/ID formation. Although some growth factors including Bmp and Fgf signaling can modulate ICD, growth factor crosstalk regulating ICD is poorly understood. Wnt canonical pathway and Bmp signal crosstalk has been considered as the essential growth factor crosstalk in organogenesis. To elucidate the crosstalk to regulate the D/ID formation, we analyzed conditional mutant mice with limb bud ectoderm expressing constitutively activated β-catenin signaling. We showed that modulation of Wnt/β-catenin signal in the limb ectoderm including the AER regulates ID apoptosis. We also demonstrated that Wnt/β-catenin signaling in the ectoderm can positively regulate Fgf8 possibly antagonizing the epithelial derived Bmp signaling. Human birth defects for digit abnormalities have been known to be affected by multiple parameters. Elucidation of the potential mechanisms underlying such D/ID development is an urgent medical issue to be solved. This work would be one of the first studies showing essential growth factor cascades in the D/ID formation.     

诱导心脏发生的早期信号通路

心脏是胚胎发生过程中最早形成的器官 .心脏前体的特化是组织间及细胞与细胞之间相互作用的结果 ,这一过程包含了诱导信号作用的时间和空间完整程序 .以脊椎动物和无脊椎动物作为模式动物 ,总结了在早期心脏发生中发挥重要作用的诱导信号通路 :BMP Dpp ,Wnt Wingless ,FGF及Notch信号通路 ,并阐述了信号通路之间的通讯 (crosstalk)以及信号通路与心脏发生相关的关键转录调节因子之间的协同诱导作用 .     

Activation of non-canonical Wnt/JNK pathway by Wnt3a is associated with differentiation fate determination of human bone marrow stromal (mesenchymal) stem cells

The canonical Wnt signaling pathway can determine human bone marrow stromal (mesenchymal) stem cell (hMSC) differentiation fate into osteoblast or adipocyte lineages. However, its downstream targets in MSC are not well characterized. Thus, using DNA microarrays, we compared global gene expression patterns induced by Wnt3a treatment in two hMSC lines: hMSC-LRP5^T253 and hMSC-LRP5^T244 cells carrying known mutations of Wnt co-receptor LRP5 (T253I or T244M) that either enhances or represses canonical Wnt signaling, respectively. Wnt3a treatment of hMSC activated not only canonical Wnt signaling, but also the non-canonical Wnt/JNK pathway through upregulation of several non-canonical Wnt components e.g. naked cuticle 1 homolog (NKD1) and WNT11. Activation of the non-canonical Wnt/JNK pathway by anisomycin enhanced osteoblast differentiation whereas its inhibition by SP600125 enhanced adipocyte differentiation of hMSC. In conclusion, canonical and non-canonical Wnt signaling cooperate in determining MSC differentiation fate.     

A protein kinase A and Wnt-dependent network regulating an intermediate stage in epithelial tubulogenesis during kidney development

Genetic interactions regulating intermediate stages of tubulogenesis in the developing kidney have been difficult to define. A systems biology strategy using microarray was combined with in vitro/ex vivo and genetic approaches to identify pathways regulating specific stages of tubulogenesis. Analysis of the progression of the metanephric mesenchyme (MM) through four stages of tubule induction and differentiation (i.e., epithelialization, tubular organization and elongation and early differentiation) revealed signaling pathways potentially involved at each stage and suggested key roles for a number of signaling molecules. A screen of the signaling pathways on in vitro/ex vivo nephron formation implicated a unique regulatory role for protein kinase A (PKA), through PKA-2, in a specific post-epithelialization morphogenetic step (conversion of the renal vesicle to the S-shaped body). Microarray analysis not only confirmed this stage-specificity, but also highlighted the upregulation of Wnt genes. Addition of PKA agonists to LIF-induced nephrons (previously shown to be a Wnt/beta-catenin dependent pathway) disrupted normal tubulogenesis in a manner similar to PKA-agonist treated MM/spinal-cord assays, suggesting that PKA regulates a Wnt-dependent tubulogenesis step. PKA induction of canonical Wnt signaling during tubulogenesis was confirmed genetically using MM from Batgal-reporter mice. Addition of a Wnt synthesis inhibitor to activated PKA cultures rescued tubulogenesis. By re-analysis of existing microarray data from the FGF8, Lim1 and Wnt4 knockouts, which arrest in early tubulogenesis, a network of genes involving PKA, Wnt, Lhx1, FGF8, and hyaluronic acid signaling regulating the transition of nascent epithelial cells to tubular epithelium was derived, helping to reconcile in vivo and in vitro/ex vivo data.     

Combined activin A/LiCl/Noggin treatment improves production of mouse embryonic stem cell-derived definitive endoderm cells

Induction of definitive endoderm (DE) cells is a prerequisite for the whole process of embryonic stem (ES) cells differentiating into hepatic or pancreatic progenitor cells. We have established an efficient method to induce mouse ES cell-derived DE cells in suspension embryonic body (EB) culture. Similar to previous studies, mouse ES cell-derived DE cells, which were defined as Cxcr4(+) c-Kit(+) , Cxcr4(+) E-cadherin(+) cells or Cxcr4(+) PDGFRa(-) cells, could be induced in the serum-free EBs at Day 4 of induction. The activations of Wnt, Nodal, and FGF signaling pathways in differentiating EBs promoted DE cell differentiation, while activation of BMP4 signaling inhibited the process. In the present study, we found that chemical activation of canonical Wnt signaling pathway by LiCl could synergize with Activin A-mediated Nodal signaling pathway to promote induction of DE cells, and inhibition of Bmp4 signaling by Noggin along with Activin A/LiCl further improved the efficiency of DE cell differentiation. The derived DE cells were proved for their capacities to become hepatic progenitor cells or pancreatic progenitor cells. In conclusion, we significantly improved the efficiency of generating mouse ES cell-derived DE cells by combined Activin A/LiCl/Noggin treatment. Our work will be greatly helpful to generate ES cell-derived hepatic cells and ES cell-derived pancreatic cells for future regenerative medicine.     

Increasingly complex: new players enter the Wnt signaling network

Wnt proteins can activate different intracellular signaling cascades in various organisms by interacting with receptors of the Frizzled family. The first identified Wnt signaling pathway, the Wnt/beta-catenin pathway, has been studied in much detail and is highly conserved among species. As to non-canonical Wnt pathways, the current situation is more nebulous partly because the intracellular mediators of this pathway are not yet fully understood and, in some cases, even identified. However, there are increasing data that prove the existence of non-canonical Wnt signaling and demonstrate its involvement in different developmental processes. In vertebrates, Wnt-11 and Wnt-5A can activate the Wnt/JNK pathway, which resembles the planar cell polarity pathway in Drosophila. The Wnt/Ca(2+)-pathway has only been described in Xenopus and zebrafish so far and it is unclear whether it also exists in other organisms. Two recent papers provide us with new insight into non-canonical Wnt signaling by (1) presenting a new intracellular mediator of non-canonical signaling in Xenopus1 and (2) implicating the existence of an additional non-canonical Wnt signaling pathway in flies.