The egl-9 family hypoxia-inducible factor (EGLN) enzymes, also known as prolyl hydroxylase domain (PHD) enzymes, are metabolic sensors regulating inflammatory responses and cellular metabolism. Both neuroinflammation and dysregulated energy metabolism are linked to motor neuron degeneration leading to amyotrophic lateral sclerosis (ALS) and correlate with disease progression. Therefore, the aim of this study was to explore the role of EGLN2 in ALS. Using a genetic and an oligonucleotide-based approach, we showed that downregulation of EGLN2 protects motor neurons and mitigates the ALS-like phenotypes in a mouse and a zebrafish model of ALS. Single-nuclei RNA sequencing of murine spinal cord revealed that loss of EGLN2 induced an astrocyte-specific downregulation of pro-inflammatory interferon-stimulated genes. In addition, we found that genetic deletion of EGLN2 restored the disturbed interferon response in iPSC-derived astrocytes from an ALS patient, confirming the novel link between EGLN2 and astrocytic interferon signaling. In conclusion, we identified EGLN2 as a potential novel ALS therapeutic target normalizing the astrocytic interferon-dependent inflammatory axis in vivo as well as in patient-derived cells. Overall design: To study the effect of EGLN2-KO in amyotrophic lateral sclerosis (ALS) patient-derived astrocytes, we performed RNA-seq analysis on three differentiations (DIFF) of iPSC-derived astrocytes from an ALS patient (SOD1-A4V), the isogenic control (SOD1-A4A) and a CRISPR-Cas9 generated EGLN2-KO (SOD1-A4V/EGLN2-/-) that were vehicle (VEH) treated or treated with TNFα, IL-1α and C1q (TIC) to provoke A1-like astrogliosis.