title: Molecular insights into Adgra2/Gpr124 and Reck intracellular trafficking.
abstract: 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.
authors.affiliation: Laboratory of Neurovascular Signaling, Department of Molecular Biology, ULB Neuroscience Institute, Université libre de Bruxelles (ULB), Gosselies B-6041, Belgium.
authors.forename: Naguissa
authors.initials: N
authors.lastname: Bostaille
authors.affiliation: Laboratory of Neurovascular Signaling, Department of Molecular Biology, ULB Neuroscience Institute, Université libre de Bruxelles (ULB), Gosselies B-6041, Belgium.
authors.forename: Anne
authors.initials: A
authors.lastname: Gauquier
authors.affiliation: Center for Microscopy and Molecular Imaging, Université libre de Bruxelles (ULB), Gosselies B-6041, Belgium.
authors.forename: Laure
authors.initials: L
authors.lastname: Twyffels
authors.affiliation: Laboratory of Neurovascular Signaling, Department of Molecular Biology, ULB Neuroscience Institute, Université libre de Bruxelles (ULB), Gosselies B-6041, Belgium Benoit.Vanhollebeke@ulb.ac.be.|Center for Microscopy and Molecular Imaging, Université libre de Bruxelles (ULB), Gosselies B-6041, Belgium.
authors.forename: Benoit
authors.identifier: http://orcid.org/0000-0002-0353-365X
authors.initials: B
authors.lastname: Vanhollebeke
keywords: Adgra2/Gpr124; Blood-brain barrier; Leucine-rich repeat; Reck; Wnt7; Zebrafish