# Control of subsynaptic domain organization and nanocolumn alignment by neurexin-3

> **NIH NIH R21** · UNIVERSITY OF COLORADO DENVER · 2023 · $184,385

## Abstract

Project Summary/Abstract
Efficient synaptic communication between neurons requires precise transcellular alignment of the presynaptic
terminal with the postsynaptic specialization. The sites of neurotransmitter release and detection are not
random; instead the active zone release machinery is locally enriched and precisely aligned transsynaptically
with locally enriched postsynaptic scaffolds and receptors. Recently, super-resolution microscopy approaches
have revealed that many proteins critical for synaptic transmission are not uniformly distributed but highly
enriched in subsynaptic domains (SSDs). These SSDs are aligned to form transsynaptic nanocolumns that are
commonly thought to regulate the efficacy of synaptic transmission. An outstanding question is: how are SSDs
regulated, localized and aligned into transsynaptic nanocolumns? Transsynaptic cell adhesion molecules have
been proposed to control the nanoscale organization of synapses because they span the synaptic cleft and are
capable of participating in bidirectional signaling via intracellular and extracellular sequences. Underscoring the
importance of adhesion molecules, mutations in genes that encode these proteins are commonly linked to
mental health and substance use disorders. While adhesion molecules studied thus far cluster in SSDs in or
around the synapse, the endogenous manipulation of presynaptic adhesion proteins has yet to reveal an
instructive role for these molecules in controlling transsynaptic SSD properties. Using 3D dSTORM
superresolution imaging, we have identified neurexin-3 (Nrxn3) as the first neurexin and first presynaptic
adhesion molecule necessary for the nanoscale organization of excitatory synapses. Conditional ablation of
neurexin-3 in primary hippocampal cultures significantly reduced the synaptic density, volume and
transsynaptic alignment of active zone and postsynaptic SSDs. We hypothesize that distinct Nrxn3 signaling
sequences govern SSD properties and nanocolumn alignment and that neurexin-3 forms presynaptic SSDs
directly aligned with postsynaptic density SSDs. This proposal builds on our preliminary and published data
that neurexin-3 is necessary for subsynaptic organization and critical for AMPAR-mediated synaptic
transmission. We will utilize 3D dSTORM imaging, molecular structure/function approaches and mouse
genetics to test our hypothesis in two specific aims. First, we will directly examine the role of neurexin-3 on
AMPA-receptor SSDs at excitatory synapses and systematically perform structure/function experiments to
determine the intracellular and extracellular signaling sequences of neurexin-3 required for nanoscale
organization and SSD alignment. Second, we will investigate the subsynaptic localization and transsynaptic
alignment of endogenous neurexin-3. Together, our findings will provide critical insight into how neurexin-3
signaling governs subsynaptic architecture and how dysfunction of neurexin-3 can contribute to the etiologies
that...

## Key facts

- **NIH application ID:** 10584530
- **Project number:** 5R21MH129620-02
- **Recipient organization:** UNIVERSITY OF COLORADO DENVER
- **Principal Investigator:** Jason Aoto
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2023
- **Award amount:** $184,385
- **Award type:** 5
- **Project period:** 2022-03-04 → 2024-02-29

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10584530

## Citation

> US National Institutes of Health, RePORTER application 10584530, Control of subsynaptic domain organization and nanocolumn alignment by neurexin-3 (5R21MH129620-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10584530. Licensed CC0.

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