FLRT Structure: Balancing Repulsion and Cell Adhesion in Cortical and Vascular Development

Abstract

SummaryFLRTs are broadly expressed proteins with the unique property of acting as homophilic cell adhesion molecules and as heterophilic repulsive ligands of Unc5/Netrin receptors. How these functions direct cell behavior and the molecular mechanisms involved remain largely unclear. Here we use X-ray crystallography to reveal the distinct structural bases for FLRT-mediated cell adhesion and repulsion in neurons. We apply this knowledge to elucidate FLRT functions during cortical development. We show that FLRTs regulate both the radial migration of pyramidal neurons, as well as their tangential spread. Mechanistically, radial migration is controlled by repulsive FLRT2-Unc5D interactions, while spatial organization in the tangential axis involves adhesive FLRT-FLRT interactions. Further, we show that the fundamental mechanisms of FLRT adhesion and repulsion are conserved between neurons and vascular endothelial cells. Our results reveal FLRTs as powerful guidance factors with structurally encoded repulsive and adhesive surfaces.

Structural basis of adhesive binding by desmocollins and desmogleins

“…S1), yielding heterophilic K D s ranging between 3.6 and 43.9 μM (Fig. 1C), similar to the affinities of other tight-binding cell adhesion molecules (22)(23)(24)(25)(26)(27)(28). Interestingly, the strongest-binding pairs (Dsg1:Dsc1 and Dsg4:Dsc1) correspond to those expressed in the outermost layers of human skin, and the weakest (Dsg3:Dsc3) to those restricted to basal layers (1,10), consistent with a potential role for differential affinities in maintenance of cell arrangement in stratified epithelia (1).…”

Placental labyrinth formation in mice requires endothelial FLRT2–UNC5B signaling

“…Transmembrane FLRT2 proteins are proteolytically cleaved to release soluble ectodomains (Seiradake et al, 2014;Yamagishi et al, 2011) that diffuse into the extracellular spaces. Thus, it is likely that one FLRT2-expressing endothelial cell acts repulsively on multiple UNC5B-expressing cells, both in paracrine and autocrine manners.…”

“…Previously, X-ray crystallography (Seiradake et al, 2014) revealed that FLRT2 binds to FLRT2 (homophilic binding) as well as to UNC5B, UNC5C and UNC5D (heterophilic binding) via different extracellular regions of FLRT2 (Fig. 5A).…”

“…5B). To evaluate whether the repulsive mechanism of interaction between FLRT2 and UNC5B acts in endothelial cells in the same way as in neuronal cells (Yamagishi et al, 2011;Seiradake et al, 2014), a Transwell assay and a stripe assay were performed in HUVECs with recombinant FLRT2 and UNC5B proteins. Compared with vehicle only, FLRT2 significantly decreased migration of HUVECs in the Transwell assay and resulted in repulsion of the cells away from stripes containing the recombinant protein ( Fig.…”

“…Fibronectin leucine-rich transmembrane protein 2 (FLRT2) is a member of the fibronectin leucine-rich transmembrane (FLRT) family of proteins, which act as repulsive ligands of the UNC5 receptor family and as chemorepellents for neurons (Lacy et al, 1999;Yamagishi et al, 2011;Seiradake et al, 2014). Outside of the nervous system, FLRT2 is expressed in the epicardium, and its global knockout leads to reduced thickness of the ventricular myocardium, suggesting a role for FLRT2 in heart morphogenesis (Müller et al, 2011).…”