Signaling Switch of the Axon Guidance Receptor Robo3 during Vertebrate Evolution

Abstract

Development of neuronal circuits is controlled by evolutionarily conserved axon guidance molecules, including Slits, the repulsive ligands for roundabout (Robo) receptors, and Netrin-1, which mediates attraction through the DCC receptor. We discovered that the Robo3 receptor fundamentally changed its mechanism of action during mammalian evolution. Unlike other Robo receptors, mammalian Robo3 is not a high-affinity receptor for Slits because of specific substitutions in the first immunoglobulin domain. Instead, Netrin-1 selectively triggers phosphorylation of mammalian Robo3 via Src kinases. Robo3 does not bind Netrin-1 directly but interacts with DCC. Netrin-1 fails to attract pontine neurons lacking Robo3, and attraction can be restored in Robo3(-/-) mice by expression of mammalian, but not nonmammalian, Robo3. We propose that Robo3 evolution was key to sculpting the mammalian brain by converting a receptor for Slit repulsion into one that both silences Slit repulsion and potentiates Netrin attraction.

Commissural axonal corridors instruct neuronal migration in the mouse spinal cord

“…It is likely to be that all signalling pathways influencing commissural guidance integrate either directly or indirectly at some molecular level 45 . However, as we and others have demonstrated that Ntn-1, Dcc, Robo3 and Boc mutant embryos have different neuronal phenotypes, this strongly supports partial non-overlapping functions of different genes in commissural guidance.…”

Slit-Robo signaling

“…Heparan sulfate proteoglycans can also bind to both Slit and Robo, forming a ternary complex and stabilizing their interaction at the membrane (Hohenester, 2008;Morlot et al, 2007;Zelina et al, 2014;Zhang et al, 2013). Slits can also homodimerize; the LRR4 domain is required for this dimerization (Seiradake et al, 2009).…”

“…More recently, it was shown that, due to a few amino acid changes in its Ig1 domain, mammalian Robo3 has lost the ability to bind Slits (Zelina et al, 2014). Concomitantly, Robo3 has acquired new signaling properties, exhibiting interactions with the netrin 1/Dcc chemoattractive signaling pathway.…”

“…The Drosophila CNS midline has since served as a model for understanding Slit-Robo signaling. Nonetheless, it has become increasingly clear that axon guidance mechanisms, including those mediated by Slits and Robos, are not as evolutionarily conserved as previously assumed; evolution has led to the molecular diversification of axon guidance genes, thereby increasing crosstalk between different ligands and receptors (Dascenco et al, 2015;Delloye-Bourgeois et al, 2014;Wang et al, 2015;Zelina et al, 2014). Furthermore, it has emerged that the outcome of Slit-Robo interactions is highly context dependent, creating a multifunctional platform for cell-cell or cell-matrix interactions impacting multiple physiological and pathological processes.…”

Making Connections: Guidance Cues and Receptors at Nonneural Cell–Cell Junctions

“…Function blocking antibodies against P-cadherin reduced the secretion of Slit2 by oral epithelia, as well as reduced P-cadherin/Robo3 complex formation (Bauer et al 2011). Complicating matters, a recent study investigating the divergent roles of Robo3 compared to other Robo family members, revealed that Robo3 does not bind tightly to Slit proteins (Zelina et al 2014). Instead, the secreted factor NELL2 is the high-affinity ligand for Robo3 (Jaworski et al 2015).…”