Canonical and noncanonical Wnt pathways share some common elements but differ in the responses they evoke. Similar to Wnt ligands acting through the canonical pathway, Wnts that activate the noncanonical signaling, such as Wnt5a, promote Disheveled (Dvl) phosphorylation and its binding to the Frizzled (Fz) Wnt receptor complex. The protein kinase CK1ε is required for Dvl/Fz association in both canonical and noncanonical signaling. Here we show that differently to its binding to canonical Wnt receptor complex, CK1ε does not require p120‐catenin for the association with the Wnt5a co‐receptor Ror2. Wnt5a promotes the formation of the Ror2–Fz complex and enables the activation of Ror2‐bound CK1ε by Fz‐associated protein phosphatase 2A. Moreover, CK1ε also regulates Ror2 protein levels; CK1ε association stabilizes Ror2, which undergoes lysosomal‐dependent degradation in the absence of this kinase. Although p120‐catenin is not required for CK1ε association with Ror2, it also participates in this signaling pathway as p120‐catenin binds and maintains Ror2 at the plasma membrane; in p120‐depleted cells, Ror2 is rapidly internalized through a clathrin‐dependent mechanism. Accordingly, downregulation of p120‐catenin or CK1ε affects late responses to Wnt5a that are also sensitive to Ror2, such as SIAH2 transcription, cell invasion, or cortical actin polarization. Our results explain how CK1ε is activated by noncanonical Wnt and identify p120‐catenin and CK1ε as two critical factors controlling Ror2 function.
Colon tumors of the mesenchymal subtype have the lowest overall survival. Snail1 is essential for the acquisition of this phenotype, characterized by increased tumor stemness and invasion, and high resistance to chemotherapy. Here, we find that Snail1 expression in colon tumor cells is dependent on an autocrine noncanonical Wnt pathway. Accordingly, depletion of Ror2, the co-receptor for noncanonical Wnts such as Wnt5a, potently decreases Snail1 expression. Wnt5a, Ror2, and Snail1 participate in a selfstimulatory feedback loop since Wnt5a increases its own synthesis in a Ror2-and Snail1-dependent fashion. This Wnt5a/Ror2/Snail1 axis controls tumor invasion, chemoresistance, and formation of tumor spheres. It also stimulates TGFb synthesis; consequently, tumor cells expressing Snail1 are more efficient in activating cancer-associated fibroblasts than the corresponding controls. Ror2 downmodulation or inhibition of the Wnt5a pathway decreases Snail1 expression in primary colon tumor cells and their ability to form tumors and liver metastases. Finally, the expression of SNAI1, ROR2, and WNT5A correlates in human colon and other tumors. These results identify inhibition of the noncanonical Wnt pathway as a putative colon tumor therapy.
International audienceOur project for iGEM 2006 consisted of designing a cellular vanillin biosensor. We used an EnvZ-E. coli strain as a chassis, and constructed two different devices: a sensor and an actuator, assembled using OmpR-P as a standardised mediator. The sensor device contained a computationally designed vanillin receptor and a synthetic two-component signal transduction protein (Trz). The receptor protein was based on a ribose-binding protein as scaffold. The Trz was built by fusion of the periplasmic and transmembrane domains of a Trg protein with an EnvZ kinase domain. When the receptor complex binds Trg, an allosteric motion is propagated to the cytoplasmic EnvZ kinase domain, resulting in autophosphorylation and subsequent phosphate transfer to the OmpR transcription factor, which finally induces transcription of the ompC promoter. As actuator, we used a synthetic transcriptional circuit, which implements an OmpR-P band detector having GFP and RFP as an output. We designed this circuit using a synthetic promoter working as an AND gate, which is synergistically activated by cI and CRP. Our constructed Trg-EnvZ fusion and AND promoter will be very useful to future synthetic biology projects. © 2007 The Institution of Engineering and Technology
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