NELL2-Robo3 complex structure reveals mechanisms of receptor activation for axon guidance

Joseph S Pak,Engin Özkan,Nischal Acharya,Zachary J Deloughery,Jing Wang,Alexander Jaworski,Yeonwoo Park

doi:10.1038/s41467-020-15211-1

Abstract

Axon pathfinding is critical for nervous system development, and it is orchestrated by molecular cues that activate receptors on the axonal growth cone. Robo family receptors bind Slit guidance cues to mediate axon repulsion. In mammals, the divergent family member Robo3 does not bind Slits, but instead signals axon repulsion from its own ligand, NELL2. Conversely, canonical Robos do not mediate NELL2 signaling. Here, we present the structures of NELL-Robo3 complexes, identifying a mode of ligand engagement for Robos that is orthogonal to Slit binding. We elucidate the structural basis for differential binding between NELL and Robo family members and show that NELL2 repulsive activity is a function of its Robo3 affinity and is enhanced by ligand trimerization. Our results reveal a mechanism of oligomerization-induced Robo activation for axon guidance and shed light on Robo family member ligand binding specificity, conformational variability, divergent modes of signaling, and evolution.

Highlights

Axon pathfinding is critical for nervous system development, and it is orchestrated by molecular cues that activate receptors on the axonal growth cone
We first examined the relationship of precrossing axons and the NELL2 protein expression domain in mouse embryonic day 9.75 (E9.75) spinal cord sections by immunohistochemistry and found that the first Robo3-expressing axons extend toward the midline by precisely circumnavigating a sharply delineated NELL2-positive region in the ventral horn (Fig. 1b)
This result indicates that NELL2 forms a very steep gradient in the neural tube and supports the idea that Robo3.1mediated NELL2 repulsion steers precrossing pioneer axons along the border formed by this gradient

Summary

Introduction

Axon pathfinding is critical for nervous system development, and it is orchestrated by molecular cues that activate receptors on the axonal growth cone. Robo family receptors bind Slit guidance cues to mediate axon repulsion. The divergent family member Robo[3] does not bind Slits, but instead signals axon repulsion from its own ligand, NELL2. Our results reveal a mechanism of oligomerization-induced Robo activation for axon guidance and shed light on Robo family member ligand binding specificity, conformational variability, divergent modes of signaling, and evolution. Netrin-1, acting through its DCC family receptors, promotes axon growth and mediates attraction[2]; Slit proteins, on the other hand, are prototypical repulsive cues that signal through receptors of the Robo family[3,4]. Slits are multidomain secreted proteins (Fig. 1a)[11] that interact with

Methods

Results

Conclusion