Reelin-Nrp1 Interaction Regulates Neocortical Dendrite Development in a Context-Specific Manner

Abstract

Reelin plays versatile roles in neocortical development. The C-terminal region (CTR) of Reelin is required for the correct formation of the superficial structure of the neocortex; however, the mechanisms by which this position-specific effect occurs remain largely unknown. In this study, we demonstrate that Reelin with an intact CTR binds to neuropilin-1 (Nrp1), a transmembrane protein.Both male and female mice were used. Nrp1 is localized with very-low-density lipoprotein receptor (VLDLR), a canonical Reelin receptor, in the superficial layers of the developing neocortex. It forms a complex with VLDLR, and this interaction is modulated by the alternative splicing of VLDLR. Reelin with an intact CTR binds more strongly to the VLDLR/Nrp1 complex than to VLDLR alone. Knockdown of Nrp1 in neurons leads to the accumulation of Dab1 protein. Since the degradation of Dab1 is induced by Reelin signaling, it is suggested that Nrp1 augments Reelin signaling. The interaction between Reelin and Nrp1 is required for normal dendritic development in superficial-layer neurons. All of these characteristics of Reelin are abrogated by proteolytic processing of the six C-terminal amino acid residues of Reelin (0.17% of the whole protein). Therefore, Nrp1 is a coreceptor molecule for Reelin and, together with the proteolytic processing of Reelin, can account for context-specific Reelin function in brain development.

Structure of Reelin repeat 8 and the adjacent C-terminal region

“…While the CTR is likely flexible due to its disproportionately basic composition, structural predictions suggest that the CTR is not completely disordered but does have secondary/tertiary structure. This fits with the current understanding of the CTR, as it has increasingly been shown to serve as a multifunctional domain that not only affects Reelin's secretion and signaling levels but also interacts with the recently reported Reelin binding partner, NRP-1 (26,28,37).…”

“…RR8 has been of increasing interest to those in the Reelin field, as new developments in its function and regulation have been reported, especially regarding the CTR (26,28,37). Despite not being able to represent the CTR in our crystallographic model of RR8, we confidently identify the region that the CTR occupies using SAXS.…”

“…A large amount of work has previously focused on the CF, as it contains both the lipoprotein receptor binding site and the Reelin dimerization site, enabling it to induce canonical Reelin signaling in dissociated neurons (21)(22)(23)(24). However, there is a growing repository of research being published on the peripheral Reelin fragments and domains, showing that they, and the proteolytic events surrounding them, play biologically relevant roles (17,(25)(26)(27)(28).…”

“…Near the C-terminus of the CTR, four consecutive arginine residues function as a Furin recognition site, leading to the cleavage of the final six amino acids from the FL protein (28). The inclusion of these final six residues has recently been shown to mediate an interaction with Neuropilin-1 (NRP-1), which has been reported to function as a co-receptor with VLDLR, and NRP-1 has previously been shown to interact with proteins bearing arginine-rich C-termini (26,38). Despite the functional importance of RR8, its structure has yet to be solved.…”

Functional characterization of RELN missense mutations involved in recessive and dominant forms of Neuronal Migration Disorders

“…The substitution of a negatively charged amino acid with a neutral one could have an impact on the folding of RELN, thus preventing the protein to acquire the proper conformation to be correctly cleaved or to bind to its receptors (VLDLR and ApoER2). A recent study ( 60 ) described a similar phenotype after overexpression of RELN in a Nrp1 knockdown (KD) mouse model. The authors demonstrated that Nrp1 forms a complex with VLDLR to which RELN strongly binds and its knocked-down results in a misorientation of electroporated cells in the aggregates.…”