# Functional and mechanistic interrogation of alpha neurexin extracellular domains

> **NIH NIH R01** · UNIVERSITY OF COLORADO DENVER · 2020 · $379,759

## Abstract

Neurexins (Nrxns) are a family of essential but poorly understood presynaptic cell-adhesion molecules that are
frequently linked to neuropsychiatric and neurodevelopmental disorders such as autism spectrum disorders
(ASDs), schizophrenia and intellectual disability (ID). Three evolutionarily conserved neurexin genes produce
longer alpha and shorter beta neurexin mRNAs that undergo extensive alternative splicing. α- and β-Nrxns
share common transmembrane and cytoplasmic sequences but differ in the length and complexity of their
extracellular domains (ECDs; 9 α-Nrxn domains compared to 1 β-Nrxn domain). Individual α-Nrxns are
associated with distinct neuropsychiatric disorders and disease-relevant mutations are commonly located in
genomic regions that code for α-Nrxn-specific extracellular sequences, suggesting that individual alpha
neurexin ECDs may control distinct aspects of synapse function. Despite their discovery over twenty years
ago, the fundamental question regarding the essential role of individual α-Nrxn ECDs at the synapse remains
unresolved. As an important first step in understanding how α-Nrxn-specific extracellular sequences function at
the synapse, our laboratory has identified an Nrxn3α compound heterozygous patient with profound ID and
epilepsy. One allele produces a non-functional protein and the second harbors a missense mutation in an
extracellular sequence shared by all α-Nrxns. Intriguingly, there are multiple ASD associated mutations in the
equivalent region of Nrxn1α indicating that this region plays an important role at the synapse. Preliminary data
from primary neurons and ex vivo acute slices revealed that expression of the missense Nrxn3α mutant
produced striking morphological and functional phenotypes at excitatory and inhibitory synapses.
Biochemically, the Nrxn3α missense mutation unexpectedly differentially modulated binding to two excitatory
postsynaptic ligands. Based on our preliminary data, we hypothesize that extracellular sequences of individual
alpha neurexins control distinct aspects of excitatory and inhibitory synapse function. Here, we will test our
central hypothesis in three specific aims: 1. Determine the impact of Nrxn3α extracellular sequences on
synaptic morphology and function in in vitro neuron cultures; 2. Biochemically assess how the Nrxn3α
missense mutation affects transsynaptic binding; and 3. Manipulate Nrxn3α ECD in vivo and assess its impact
on basal excitatory and inhibitory synaptic transmission and activity-dependent plasticity in ex vivo slices. To
accomplish aims 1 and 3, we will use molecular replacement, shRNA-mediated knockdown of endogenous
Nrxn3α and replacement with wild-type or mutant Nrxn3α to faithfully recapitulate the disease state, combined
with immunocytochemistry, electrophysiology and electron microscopy. Aim 2 will use in vitro biochemical and
structure/function approaches to measure binding affinities to known Nrxn ligands. These aims will provide first
insight into t...

## Key facts

- **NIH application ID:** 9901552
- **Project number:** 5R01MH116901-03
- **Recipient organization:** UNIVERSITY OF COLORADO DENVER
- **Principal Investigator:** Jason Aoto
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $379,759
- **Award type:** 5
- **Project period:** 2018-06-01 → 2023-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9901552

## Citation

> US National Institutes of Health, RePORTER application 9901552, Functional and mechanistic interrogation of alpha neurexin extracellular domains (5R01MH116901-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9901552. Licensed CC0.

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