Search

Within the central nervous system, Wnt7a/b are unambiguously discriminated from other Wnt ligands by an endothelial receptor complex made of the glycosylphosphatidylinositol (GPI)-anchored Reck and the adhesion G protein-coupled receptor (GPCR) Gpr124. Reck is a Wnt7a/b-specific receptor, while Gpr124 facilitates the delivery of Reck-bound Wnt7a/b ligands to Frizzled, through partially characterized mechanisms. We report that, in zebrafish, the Gpr124-Frizzled interactions are dominated by intracellular scaffolds that exploit the striking molecular mimicry between Gpr124 and Frizzled intracellular domains (ICDs): an internal Dvl-binding motif and a C-terminal ETTV motif that recruits Dlg4 and Magi3. By contrast, mammalian Gpr124 receptors exhibit an ICD-independent interaction mechanism governed by species-specific attributes of their transmembrane and extracellular domains. This mechanism seemingly evolved to replace the Dvl-mediated mechanism. By contrasting zebrafish, mouse, and human Gpr124, this study provides insights into the evolution of Gpr124/Reck function across the vertebrate clade, a receptor complex uniquely implicated in Wnt ligand-specific cellular responses., Competing Interests: Declaration of interests The ULB (B.V.) has filed for patent protection for Gpr124/Reck-specific agonism. B.V. is a founder, shareholder, and consultant of NeuVasQ Biotechnologies., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

The blood-brain barrier (BBB) protects the central nervous system (CNS) from harmful blood-borne factors. Although BBB dysfunction is a hallmark of several neurological disorders, therapies to restore BBB function are lacking. An attractive strategy is to repurpose developmental BBB regulators, such as Wnt7a, into BBB-protective agents. However, safe therapeutic use of Wnt ligands is complicated by their pleiotropic Frizzled signaling activities. Taking advantage of the Wnt7a/b-specific Gpr124/Reck co-receptor complex, we genetically engineered Wnt7a ligands into BBB-specific Wnt activators. In a "hit-and-run" adeno-associated virus-assisted CNS gene delivery setting, these new Gpr124/Reck-specific agonists protected BBB function, thereby mitigating glioblastoma expansion and ischemic stroke infarction. This work reveals that the signaling specificity of Wnt ligands is adjustable and defines a modality to treat CNS disorders by normalizing the BBB.

Wnt signaling is key to many developmental, physiological, and disease processes in which cells seem able to discriminate between multiple Wnt ligands. This selective Wnt recognition or "decoding" capacity has remained enigmatic because Wnt/Frizzled interactions are largely incompatible with monospecific recognition. Gpr124 and Reck enable brain endothelial cells to selectively respond to Wnt7. We show that Reck binds with low micromolar affinity to the intrinsically disordered linker region of Wnt7. Availability of Reck-bound Wnt7 for Frizzled signaling relies on the interaction between Gpr124 and Dishevelled. Through polymerization, Dishevelled recruits Gpr124 and the associated Reck-bound Wnt7 into dynamic Wnt/Frizzled/Lrp5/6 signalosomes, resulting in increased local concentrations of Wnt7 available for Frizzled signaling. This work provides mechanistic insights into the Wnt decoding capacities of vertebrate cells and unravels structural determinants of the functional diversification of Wnt family members., (Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Adgra2, formerly known as Gpr124, is a key regulator of cerebrovascular development in vertebrates. Together with the GPI-anchored glycoprotein Reck, this adhesion GPCR (aGPCR) stimulates Wnt7-dependent Wnt/β-catenin signaling to promote brain vascular invasion in an endothelial cell-autonomous manner. Adgra2 and Reck have been proposed to assemble a receptor complex at the plasma membrane, but the molecular modalities of their functional synergy remain to be investigated. In particular, as typically found in aGPCRs, the ectodomain of Adgra2 is rich in protein-protein interaction motifs whose contributions to receptor function are unknown. In opposition to the severe ADGRA2 genetic lesions found in previously generated zebrafish and mouse models, the zebrafish ouchless allele encodes an aberrantly-spliced and inactive receptor lacking a single leucine-rich repeat (LRR) unit within its N-terminus. By characterizing this allele we uncover that, in contrast to all other extracellular domains, the precise composition of the LRR domain determines proper receptor trafficking to the plasma membrane. Using CRISPR/Cas9 engineered cells, we further show that Adgra2 trafficking occurs in a Reck-independent manner and that, similarly, Reck reaches the plasma membrane irrespective of Adgra2 expression or localization, suggesting that the partners meet at the plasma membrane after independent intracellular trafficking events., (© 2016. Published by The Company of Biologists Ltd.)

Despite the critical role of endothelial Wnt/β-catenin signaling during central nervous system (CNS) vascularization, how endothelial cells sense and respond to specific Wnt ligands and what aspects of the multistep process of intra-cerebral blood vessel morphogenesis are controlled by these angiogenic signals remain poorly understood. We addressed these questions at single-cell resolution in zebrafish embryos. We identify the GPI-anchored MMP inhibitor Reck and the adhesion GPCR Gpr124 as integral components of a Wnt7a/Wnt7b-specific signaling complex required for brain angiogenesis and dorsal root ganglia neurogenesis. We further show that this atypical Wnt/β-catenin signaling pathway selectively controls endothelial tip cell function and hence, that mosaic restoration of single wild-type tip cells in Wnt/β-catenin-deficient perineural vessels is sufficient to initiate the formation of CNS vessels. Our results identify molecular determinants of ligand specificity of Wnt/β-catenin signaling and provide evidence for organ-specific control of vascular invasion through tight modulation of tip cell function.