Dishevelled-DEP domain interacting protein (DDIP) inhibits Wnt signaling by promoting TCF4 degradation and disrupting the TCF4/β-catenin complex

The TCF4/β-catenin complex, the executor of canonical Wnt/β-catenin signaling, is regulated by a variety of factors. Among these, Dishevelled (Dvl) is a critical regulator that releases β-catenin from degradation and stabilizes TCF4/β-catenin complex. Here, we report that DDIP (Dishevelled-DEP domain Interacting Protein, also named as Spats1, spermatogenesis associated, serine-rich 1), a novel protein that interacts with Dvl, regulates Wnt signaling. We provide evidence that DDIP suppresses Lef-1 luciferase reporter activity stimulated by Wnt1, Dvl2 or β-catenin, interacts with the TCF4/β-catenin complex, and disrupts the interaction of TCF4 and β-catenin by promoting TCF4 degradation through the proteasome pathway. Our results indicate that DDIP is a negative regulator of the canonical Wnt signaling.     

Hhex Is Necessary for the Hepatic Differentiation of Mouse ES Cells and Acts via Vegf Signaling

Elucidating the molecular mechanisms involved in the differentiation of stem cells to hepatic cells is critical for both understanding normal developmental processes as well as for optimizing the generation of functional hepatic cells for therapy. We performed in vitro differentiation of mouse embryonic stem cells (mESCs) with a null mutation in the homeobox gene Hhex and show that Hhex^-/- mESCs fail to differentiate from definitive endoderm (Sox17⁺/Foxa2⁺) to hepatic endoderm (Alb⁺/Dlk⁺). In addition, hepatic culture elicited a >7-fold increase in Vegfa mRNA expression in Hhex^-/- cells compared to Hhex^+/+ cells. Furthermore, we identified VEGFR2⁺/ALB⁺/CD34^- in early Hhex^+/+ hepatic cultures. These cells were absent in Hhex^-/- cultures. Finally, through manipulation of Hhex and Vegfa expression, gain and loss of expression experiments revealed that Hhex shares an inverse relationship with the activity of the Vegf signaling pathway in supporting hepatic differentiation. In summary, our results suggest that Hhex represses Vegf signaling during hepatic differentiation of mouse ESCs allowing for cell-type autonomous regulation of Vegfr2 activity independent of endothelial cells.

Highlights

• Hhex^-/- ESCs fail to differentiate from definitive endoderm to hepatic endoderm
• This defect involves perturbation of VEGF signaling pathway
• Differentiation involving this pathway produces VEGFR2+ hepatic progenitor cells
• VEGF regulation of hepatic specification is independent of endothelial cells