# Regulation of the orphan G-protein–coupled receptor GPRC5B by MLC1 and the cell adhesion molecule GlialCAM in megalencephalic leukoencephalopathy

## TL;DR

This study explores how the proteins MLC1 and GlialCAM regulate the GPRC5B receptor, offering new insights into the mechanisms behind a rare neurological disorder.

## Contribution

The study identifies GPRC5B as a novel interactor of MLC1 and GlialCAM, revealing their regulatory roles in receptor signaling.

## Key findings

- GPRC5B's constitutive activity is inhibited by MLC1, possibly through interference with its oligomerization.
- GlialCAM enhances β-arrestin 2 recruitment and mislocalizes from cell junctions.
- MLC-associated GPRC5B mutants show increased stability and affinity for GlialCAM without causing mislocalization.

## Abstract

Megalencephalic leukoencephalopathy with subcortical cyst (MLC) is a rare leukodystrophy primarily caused by mutations in two genes: MLC1, encoding a membrane protein of unknown function, and GlialCAM, a cell adhesion molecule. Although MLC1 has been implicated in downregulating signaling pathways, its molecular mechanisms remain elusive. Recently, the orphan G-protein–coupled receptor GPRC5B was identified as a novel interactor of both GlialCAM and MLC1, with dominant heterozygous mutations found in MLC patients, suggesting that GlialCAM and MLC1 may regulate cell signaling via GPRC5B. Here, we show that GPRC5B exhibits constitutive activity, which is inhibited by MLC1, likely through interference with GPRC5B oligomerization. Conversely, GlialCAM enhances β-arrestin 2 recruitment, leading to its own mislocalization from cell–cell junctions. MLC-associated GPRC5B mutants show enhanced maturation and increased stability at the plasma membrane, retain normal constitutive activity and responsiveness to MLC1 and GlialCAM but display increased affinity for GlialCAM and localize to cell–cell junctions in its presence. Notably, coexpression of GlialCAM with these mutants does not induce GlialCAM mislocalization. We propose a model in which finely tuned interactions among GPRC5B, GlialCAM, and MLC1 regulate receptor signaling. These findings provide the first biochemical evidence of GlialCAM and MLC1 modulating GPRC5B activity, suggesting a biochemical explanation for the gain-of-function phenotype observed in GPRC5B MLC mutants. Importantly, our work supports the potential of targeting GPRC5B as a therapeutic strategy in MLC.