# Optical interrogation of acid-sensing ion channel activation and desensitization through genetic code expansion

> **NIH NIH F31** · UNIVERSITY OF ROCHESTER · 2021 · $45,918

## Abstract

Project Summary This proposal will paint a clear picture of the molecular function of a unique family of proton-
sensitive ion channels, known as Acid-sensing ion channels (ASICs). Located primarily in the central and
peripheral nervous system, ASICs play a wide range of roles from synaptic plasticity to pain. In acidosis-related
CNS pathologies like cerebral ischemia, ASICs have been implicated in causing neurodegeneration. These
channels are activated by acidic shifts in extracellular pH below 7, where excess protons in solution bind the
large extracellular domain and allow for the pore to open. ASICs primarily reside in three states: 1. At
physiological or alkaline pH, the channel resides in an inactive resting state. 2. During mild acidification, the
channel reverts to a proton-bound, inactive conformation known as the desensitized state. 3. Upon initial heavy
acidification, the channel is in an open state, but over prolonged exposures, the channel desensitizes. While the
crystal structures of these states help illuminate potential changes in the channel between these state transitions,
experimental testing is required to functionally assess whether they are pertinent to channel gating. To elucidate
this, the two following specific aims will be employed. Aim 1 will determine conformational changes that are
necessary and sufficient for the mechanisms of activation using noncanonical amino acid incorporation to induce
optogenetic crosslinking and sidechain isomerization at regions that undergo large conformational changes and
test the functional consequences. Specifically, this aim will determine the contributions of the acidic pocket and
palm domain to activation. Aim 2 will evaluate the mechanism of β11-12 linker regulation of desensitization by
first understanding the role of Asp415, a residue within the linker, and its conformational consequences in
desensitization through the use site-directed mutagenesis and photocrosslinking. Further, we will investigate the
surrounding environment of the β11-12 linker by a phylogenetic comparison of ASIC isoforms and how this
environment can control the movement of the linker, which we have previously shown to control desensitization.
The outcomes of these two aims will fill the gaps in our knowledge for ASIC gating processes to allow for the
development of ASIC-targeted therapeutics. Furthermore, this study will lay the groundwork for optogenetic
modulation of ASICs through genetic code expansion. These tools will provide more precise control over the
channels activity in in vitro and in vivo model systems. The fellowship training plan that accompanies this
proposal will support my growth in reading, writing and communication skills to become a successful scientist.
Technically, the training plan acts as a guide to further enhance my electrophysiological skills, i.e. patch clamp,
and bolster my knowledge and skill in optogenetics, genetic code expansion and molecular modelling. The
University of Roches...

## Key facts

- **NIH application ID:** 10312523
- **Project number:** 1F31NS120445-01A1
- **Recipient organization:** UNIVERSITY OF ROCHESTER
- **Principal Investigator:** Matthew L Rook
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $45,918
- **Award type:** 1
- **Project period:** 2021-08-01 → 2022-07-15

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10312523

## Citation

> US National Institutes of Health, RePORTER application 10312523, Optical interrogation of acid-sensing ion channel activation and desensitization through genetic code expansion (1F31NS120445-01A1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10312523. Licensed CC0.

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