# Structure-function analysis of the volume-regulated anion channel VRAC using novel LRRC8 chimeras

> **NIH NIH R01** · VANDERBILT UNIVERSITY MEDICAL CENTER · 2024 · $474,220

## Abstract

PROJECT SUMMARY
 The volume-regulated anion channel (VRAC) is expressed ubiquitously in vertebrate cells where it
mediates the efflux of Cl- and organic solutes required for cell volume regulation, an essential physiological
process. VRACs are activated and inactivated by cell swelling and shrinkage, respectively. They also detect
changes in intracellular ionic strength, which modifies their sensitivity to cell volume changes. VRACs and the
genes that encode them are implicated in multiple diseases including diabetes, obesity, cancer and immunity.
 Whole genome RNA interference screening led to the demonstration in 2014 that VRACs are encoded
by five members of the Lrrc8 gene family, Lrrc8a–e. VRAC/LRRC8 channels are hexaheteromers and
require co-assembly of the essential subunit LRRC8A with one or more other LRRC8 proteins. Subunit
assembly order and stoichiometry are unknown.
 Cryo-electron microscopy (EM) structures of homomeric LRRC8A and LRRC8D channels were
recently determined. However, LRRC8A and LRRC8D homomers do not exist in Nature. Furthermore,
LRRC8A homomeric channels have non-native functional properties and LRRC8D channel properties are
undefinable because they are not trafficked to the plasma membrane. Existing cryo-EM structures thus have
limitations for understanding VRAC/LRRC8 structure-function relationships. Directly translating LRRC8A and
LRRC8D cryo-EM structural information into functional understanding is further constrained by the unknown
and likely variable stoichiometry and assembly of hexaheteromeric VRAC/LRRC8 channels.
 Our laboratory, funded by DK51610, has studied VRAC extensively and was the first to demonstrate
many of the channel's unique functional properties. Most recently, we described novel LRRC8 chimeric
channel constructs that allow detailed molecular study of homomeric channels with physiologically
relevant functional properties and defined stoichiometry and assembly. Our chimera studies uniquely
demonstrated that 1) the LRRC8A intracellular loop, IL1, has unique structural features, 2) it is required for cell
volume sensing, 3) the LRRC8 C-terminus is required for sensing changes in intracellular ionic strength and 4)
both the LRRC8A IL1 and C-terminus are required for correct cellular processing of VRAC/LRRC8 channels.
 The overarching goal of this R01 renewal application is to utilize these novel LRRC8 chimeras to better
elucidate VRAC/LRRC8 channel structure-function relationships. We will characterize the roles of the
LRRC8A C-terminus in VRAC/LRRC8 channel regulation and will test the hypothesis that the LRRC8 IL1
determines VRAC/LRRC8 channel pore properties and regulates channel gating. We will also determine the
cryo-EM structure of a unique LRRC8 chimera in multiple physiologically relevant conformations. Our studies
will provide novel insights into the regulation and function of VRAC/LRRC8 channels and will provide a higher
confidence foundation for detailed mutagenesis-based structure-functio...

## Key facts

- **NIH application ID:** 10873221
- **Project number:** 5R01DK051610-29
- **Recipient organization:** VANDERBILT UNIVERSITY MEDICAL CENTER
- **Principal Investigator:** Jerod S. Denton
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $474,220
- **Award type:** 5
- **Project period:** 1996-09-01 → 2026-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10873221

## Citation

> US National Institutes of Health, RePORTER application 10873221, Structure-function analysis of the volume-regulated anion channel VRAC using novel LRRC8 chimeras (5R01DK051610-29). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10873221. Licensed CC0.

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