# Hypoxic stress is an early pathogenic event in human VCP-mutant ALS astrocytes

## TL;DR

This study shows that astrocytes with VCP mutations linked to ALS experience hypoxic stress even under normal conditions, leading to mitochondrial and metabolic issues that impair their support of neurons.

## Contribution

The study identifies hypoxic stress as an early and functionally significant event in VCP-mutant ALS astrocytes, with HIF-1ɑ signaling as a potential therapeutic target.

## Key findings

- VCP-mutant astrocytes show increased HIF-1ɑ activation, mitochondrial depolarization, and lipid droplet accumulation under normal conditions.
- HIF-1ɑ stabilization with DMOG in control astrocytes recapitulates the VCP-mutant astrocyte phenotypes.
- Hypoxic astrocytes fail to rescue RNA-binding protein mislocalization in motor neurons, unlike healthy astrocytes.

## Abstract

Astrocytes are essential regulators of neuronal health, and their dysfunction contributes to neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS). Using human induced pluripotent stem cell (iPSC)-derived astrocytes carrying ALS-associated VCP mutations, we uncover cell-autonomous activation of the hypoxia response under basal conditions. VCP-mutant astrocytes exhibit increased nuclear hypoxia-inducible factor (HIF)-1ɑ, mitochondrial depolarization, and lipid droplet accumulation. Mimicking hypoxia in control astrocytes by HIF-1ɑ stabilization with dimethyloxalylglycine recapitulates these phenotypes. Transcriptomic and CUT&RUN profiling reveal direct HIF-1ɑ binding to canonical hypoxia-responsive genes in VCP-mutant astrocytes and a transcriptional signature of metabolic reprogramming and mitochondrial dysfunction under normoxia. Furthermore, conditioned medium from hypoxia-exposed astrocytes fails to rescue RNA-binding protein mislocalization in motor neurons, unlike medium from healthy counterparts. Together, these findings demonstrate that aberrant HIF-1ɑ activation drives astrocytic dysfunction and compromises neuronal support, identifying hypoxic stress as an early and functionally consequential event in VCP-mutant ALS, with therapeutic implications for targeting HIF-1ɑ signaling.

•VCP-ALS astrocytes show cell-autonomous HIF-1α activation at baseline•Hypoxia or DMOG alone recapitulate mitochondrial and lipid droplet phenotypes•CUT&RUN confirms HIF-1α binding to canonical targets in VCP-ALS astrocytes•Hypoxic astrocytes fail to correct VCP-ALS neuronal RBP mislocalization

VCP-ALS astrocytes show cell-autonomous HIF-1α activation at baseline

Hypoxia or DMOG alone recapitulate mitochondrial and lipid droplet phenotypes

CUT&RUN confirms HIF-1α binding to canonical targets in VCP-ALS astrocytes

Hypoxic astrocytes fail to correct VCP-ALS neuronal RBP mislocalization

In this article, Franklin and colleagues show that human VCP-ALS astrocytes exhibit hypoxic stress through basal HIF-1α activation and binding to canonical targets, mitochondrial depolarization, and lipid droplet accumulation. Hypoxia or DMOG exposure phenocopies these defects in control astrocytes, which then fail to correct VCP-ALS motor neuron RNA-binding protein mislocalization. HIF-1α signaling is a potential therapeutic target in ALS.