The Crystal Structure of Netrin-1 in Complex with DCC Reveals the Bifunctionality of Netrin-1 As a Guidance Cue

Abstract

Netrin-1 is a guidance cue that can trigger either attraction or repulsion effects on migrating neurons, depending on the repertoire of receptors available on the growth cone. How a single chemotropic molecule can act in such contradictory ways has long been a puzzle at the molecular level. Here we present the crystal structure of netrin-1 in complex with the Deleted in Colorectal Cancer (DCC) receptor. We show that one netrin-1 molecule can simultaneously bind to two DCC molecules through a DCC-specific site and through a unique generic receptor binding site, where sulfate ions staple together positively charged patches on both DCC and netrin-1. Furthermore, we demonstrate that UNC5A can replace DCC on the generic receptor binding site to switch the response from attraction to repulsion. We propose that the modularity of binding allows for the association of other netrin receptors at the generic binding site, eliciting alternative turning responses.

Structural Characterization of the Extracellular Domain of CASPR2 and Insights into Its Association with the Novel Ligand Contactin1

“…Using several deletion constructs of both CASPR2 and CNTN1 enabled us to identify CNTN1 Ig-5/Fn1 and CASPR2 D1-6 as minimal binding cassette. However, the requirement for multiple domains is not unusual, especially for proteins containing Ig and FN3 domains (49). Our data indicate that the recombinant, purified extracellular domains of CASPR2, CNTN1, and TAG-1 are correctly folded because they appear non-aggregating (monodisperse) and without degradation in SEC experiments and SDS-PAGE, and that their expression levels were comparable with well folded proteins with which we have worked in the past (18,43).…”

CD146: the unveiling of a pro-angiogenic netrin receptor

“…Through deletion analysis they show that the V domain of netrin-1, which contains three laminin-like EGF domains, plays an important role in binding. By analogy with data from the crystal structure of netrin-1 in complex with DCC [5], regions in both the V domain and the N-terminal VI domain of netrin-1 may bind CD146, potentially explaining the residual binding the authors observe with their V domain mutant. Netrin-1 treatment of endothelial cells resulted in a biphasic response with low doses (50-200 ng/mL) inducing proliferation, migration and tube formation and high doses (1 000-2 000 ng/mL) inhibiting these effects.…”

“…A related DCC family member, neogenin, also promotes axon guidance, whereas members of the uncoordinated 5 family (UNC5A-D) predominantly inhibit neurogenesis. Recent structural studies demonstrate that netrin-1 is able to crosslink its receptors, with DCC/ DCC homodimers promoting chemoattraction and DCC/UNC5 heterodimers promoting chemorepulsion [5]. In the vascular system UNC5B is highly expressed in endothelial cells during embryonic development and pathological angiogenesis and, like the role of UNC5s in repression of neurogenesis, may mediate netrin-1 antiangiogenic activity [3].…”

Mutations in the netrin-1 gene cause congenital mirror movements

“…The 3 reported variants are 2 missense mutations and 1 in-frame deletion located in a single domain, the NTR domain, at the C-terminal part of the protein. The NTR domain is not necessary for secretion (53) and not required for DCC binding (54)(55)(56). The 3 mutations are predicted to be pathogenic by different algorithms (Table 1), whereas very few NTN1 variants in the ExAC database are predicted to be damaging.…”