Deletion of the Dishevelled family of genes disrupts anterior-posterior axis specification and selectively prevents mesoderm differentiation.

The Dishevelled proteins transduce both canonical Wnt/ß-catenin and non-canonical Wnt/planar cell polarity (PCP) signaling pathways to regulate many key developmental processes during embryogenesis. Here, we disrupt both canonical and non-canonical Wnt pathways by targeting the entire Dishevelled family of genes (Dvl1, Dvl2, and Dvl3) to investigate their functional roles in the early embryo. We identified several defects in anterior-posterior axis specification and mesoderm patterning in Dvl1+/-; Dvl2-/-; Dvl3-/- embryos. Homozygous deletions in all three Dvl genes (Dvl TKO) resulted in defects in distal visceral endoderm migration and a complete failure to induce mesoderm formation. To identify potential mechanisms that lead to the defects in the developmental processes preceding gastrulation, we generated Dvl TKO mouse embryonic stem cells (mESCs) and compared the transcriptional profile of these cells with wild-type (WT) mESCs during germ lineage differentiation into 3D embryoid bodies (EBs). While the Dvl TKO mESCs displayed similar morphology, self-renewal properties, and minor transcriptional variation from WT mESCs, we identified major transcriptional dysregulation in the Dvl TKO EBs during differentiation in a number of genes involved in anterior-posterior pattern specification, gastrulation induction, mesenchyme morphogenesis, and mesoderm-derived tissue development. The absence of the Dvls leads to specific down-regulation of BMP signaling genes. Furthermore, exogenous activation of canonical Wnt, BMP, and Nodal signaling all fail to rescue the mesodermal defects in the Dvl TKO EBs. Moreover, endoderm differentiation was promoted in the absence of mesoderm in the Dvl TKO EBs, while the suppression of ectoderm differentiation was delayed. Overall, we demonstrate that the Dvls are dispensable for maintaining self-renewal in mESCs but are critical during differentiation to regulate key developmental signaling pathways to promote proper axis specification and mesoderm formation.

Dishevelled 1, a pivotal positive regulator of the Wnt signalling pathway, mediates 5-fluorouracil resistance in HepG2 cells.

Acquired resistance to 5-fluorouracil (5-FU) frequently occurs in patients with hepatocellular carcinoma (HCC), the underlying molecular mechanisms of which are poorly understood. The aim of this study was to identify candidate genes and associated signalling pathways that may play a role in developing drug resistance following repeated 5-FU treatments. In this work, we established 5-FU-resistant cells (HepG2/5-FU) using stepwise increasing concentrations of 5-FU in parental HepG2 cells. Using transcriptome sequencing, we found that the expressions of the Wnt signalling genes, including negative regulators (DKK1, DKK3, ZNRF3, RNF43 and APC2) and positive regulators (FZD10 and DVL1), were significantly downregulated and upregulated in HepG2/5-FU cells, respectively, resulting in increased Wnt signalling. Dishevelled-1 (DVL1) is an essential Wnt signalling pathway component that stabilizes ß-catenin and mediates the Wnt pathway. Silencing DVL1 using siDVL1 or other small molecular inhibitors in HepG2/5-FU cells could restore 5-FU responsiveness via reduced cell proliferation and migration, and increased apoptosis. Moreover, DVL1 was found to be upregulated in BEL-7402/5-FU cells when compared to the parental BEL-7402 cells. Collectively, our results provide the first clue towards understanding the contribution of DVL1-mediated acquired resistance to 5-FU in HepG2/5-FU cells, suggesting a promising therapeutic strategy for liver cancer resistant to 5-FU.

Fzd7 (Frizzled-7) Expressed by Endothelial Cells Controls Blood Vessel Formation Through Wnt/ß-Catenin Canonical Signaling.

OBJECTIVE: Vessel formation requires precise orchestration of a series of morphometric and molecular events controlled by a multitude of angiogenic factors and morphogens. Wnt/frizzled signaling is required for proper vascular formation. In this study, we investigated the role of the Fzd7 (frizzled-7) receptor in retinal vascular development and its relationship with the Wnt/ß-catenin canonical pathway and Notch signaling. APPROACH AND RESULTS: Using transgenic mice, we demonstrated that Fzd7 is required for postnatal vascular formation. Endothelial cell (EC) deletion of fzd7 (fzd7ECKO) delayed retinal plexus formation because of an impairment in tip cell phenotype and a decrease in stalk cell proliferation. Dvl (dishevelled) proteins are a main component of Wnt signaling and play a functionally redundant role. We found that Dvl3 depletion in dvl1-/- mice mimicked the fzd7ECKO vascular phenotype and demonstrated that Fzd7 acted via ß-catenin activation by showing that LiCl treatment rescued impairment in tip and stalk cell phenotypes induced in fzd7 mutants. Deletion of fzd7 or Dvl1/3 induced a strong decrease in Wnt canonical genes and Notch partners' expression. Genetic and pharmacological rescue strategies demonstrated that Fzd7 acted via ß-catenin activation, upstream of Notch signaling to control Dll4 and Jagged1 EC expression. CONCLUSIONS: Fzd7 expressed by EC drives postnatal angiogenesis via activation of Dvl/ß-catenin signaling and can control the integrative interaction of Wnt and Notch signaling during postnatal angiogenesis.