A Novel Role for the BMP Antagonist Noggin in Sensitizing Cells to Non-canonical Wnt-5a/Ror2/Disheveled Pathway Activation.

Mammalian limb development is driven by the integrative input from several signaling pathways; a failure to receive or a misinterpretation of these signals results in skeletal defects. The brachydactylies, a group of overlapping inherited human hand malformation syndromes, are mainly caused by mutations in BMP signaling pathway components. Two closely related forms, Brachydactyly type B2 (BDB2) and BDB1 are caused by mutations in the BMP antagonist Noggin (NOG) and the atypical receptor tyrosine kinase ROR2 that acts as a receptor in the non-canonical Wnt pathway. Genetic analysis of Nog and Ror2 functional interaction via crossing Noggin and Ror2 mutant mice revealed a widening of skeletal elements in compound but not in any of the single mutants, thus indicating genetic interaction. Since ROR2 is a non-canonical Wnt co-receptor specific for Wnt-5a we speculated that this phenotype might be a result of deregulated Wnt-5a signaling activation, which is known to be essential for limb skeletal elements growth and patterning. We show that Noggin potentiates activation of the Wnt-5a-Ror2-Disheveled (Dvl) pathway in mouse embryonic fibroblast (MEF) cells in a Ror2-dependent fashion. Rat chondrosarcoma chondrocytes (RCS), however, are not able to respond to Noggin in this fashion unless growth arrest is induced by FGF2. In summary, our data demonstrate genetic interaction between Noggin and Ror2 and show that Noggin can sensitize cells to Wnt-5a/Ror2-mediated non-canonical Wnt signaling, a feature that in cartilage may depend on the presence of active FGF signaling. These findings indicate an unappreciated function of Noggin that will help to understand BMP and Wnt/PCP signaling pathway interactions.

Negative regulation of Wnt signaling mediated by CK1-phosphorylated Dishevelled via Ror2.

Dishevelled (Dvl) is a multifunctional effector of different Wnt cascades. Both canonical Wnt3a and noncanonical Wnt5a stimulate casein-kinase-1 (CK1) -mediated phosphorylation of Dvl, visualized as electrophoretic mobility shift [phosphorylated and shifted Dvl (ps-Dvl)]. However, the role of this phosphorylation remains obscure. Here we report the functional interaction of ps-Dvl with the receptor tyrosine kinase Ror2, which is an alternative Wnt receptor and is able to inhibit canonical Wnt signaling. We demonstrate interaction between Ror2 and ps-Dvl at the cell membrane after Wnt3a or Wnt5a stimulus dependent on CK1. Ps-Dvl interacts with the C-terminal proline-serine-threonine-rich domain of Ror2, which is required for efficient inhibition of canonical Wnt signaling. We further show that the Dvl C terminus, which seems to be exposed in ps-Dvl and efficiently binds Ror2, is an intrinsic negative regulator of the canonical Wnt pathway downstream of beta-catenin. The Dvl C terminus is necessary and sufficient to inhibit canonical Wnt/beta-catenin signaling, which is dependent on the presence of Ror2. Furthermore, both the Dvl C terminus and CK1epsilon can inhibit the Wnt5a/Ror2/ATF2 pathway in mammalian cells and Xenopus explant cultures. This suggests that phosphorylation of Dvl triggers negative feedback regulation for different branches of Wnt signaling in a Ror2-dependent manner.