# N‑Sulfated Heparan Sulfate Promotes Reelin Signaling as a Co-receptor

## TL;DR

N-sulfated heparan sulfate helps Reelin signaling in the brain, acting as a co-receptor and influencing diseases like Alzheimer's.

## Contribution

HS N-sulfation is identified as a key determinant for Reelin signaling, establishing HS as a co-receptor.

## Key findings

- Full-length Reelin binds HS with high affinity (K_D = 17 ± 5 nM), enhanced by the COLBOS variant.
- HS N-sulfation is critical for Reelin–HS binding, confirmed by SPR, glycan arrays, and Haddock modeling.
- Heparinase treatment or NDST1 knockout reduces Reelin binding and receptor activation, showing HS's coreceptor role.

## Abstract

Heparan sulfate (HS)
plays a central role in signal transduction,
while Reelin is an essential signaling protein in both the developing
and adult brain. A Reelin COLBOS variant was recently discovered with
enhanced HS binding and resilience against autosomal dominant Alzheimer’s
disease (ADAD), underscoring the importance of Reelin–HS interactions.
However, the glycan determinants of Reelin–HS interactions
have not been well-characterized, which we systematically investigated
here. Surface plasmon resonance (SPR) showed that full length Reelin
binds HS with high affinity (K
D = 17 ±
5 nM), which is enhanced by the COLBOS variant (K
D = 10 ± 2 nM). Competition SPR and glycan array
studies further revealed that HS N-sulfation is critical for Reelin–HS
binding, consistent with Haddock modeling. In cell surface binding
assays, heparinase treatment, which degrades HS, or the knockout of
a key HS N-sulfation enzyme (NDST1) significantly reduced Reelin attachment.
Functionally, a cellular split-luciferase assay showed that heparinase
treatment or adding heparin in culture medium reduces Reelin-induced
ApoER2 dimerization, demonstrating that HS is a coreceptor for Reelin
receptor activation. In contrast, N-desulfated heparin does not inhibit
Reelin receptor dimerization. Our work establishes HS as a coreceptor
for Reelin signaling and N-sulfation as a key glycan determinant of
Reelin–HS recognition. Our work provides mechanistic insights
into diverse neurodevelopmental and neurodegenerative diseases associated
with Reelin signaling and suggests novel therapeutic strategies targeting
HS sulfation.