Intermediate filament dysregulation and astrocytopathy in the human disease model ofKLHL16mutation in giant axonal neuropathy (GAN)

Abstract

AbstractGiant Axonal Neuropathy (GAN) is a pediatric neurodegenerative disease caused byKLHL16mutations.KLHL16encodes gigaxonin, a regulator of intermediate filament (IF) protein turnover. Previous neuropathological studies and our own examination of postmortem GAN brain tissue in the current study revealed astrocyte involvement in GAN. To study the underlying mechanisms, we reprogrammed skin fibroblasts from seven GAN patients carrying differentKLHL16mutations to iPSCs. Isogenic controls with restored IF phenotypes were derived via CRISPR/Cas9 editing of one patient carrying a homozygous missense mutation (G332R). Neural progenitor cells (NPCs), astrocytes, and brain organoids were generated through directed differentiation. All GAN iPSC lines were deficient for gigaxonin, which was restored in the isogenic control. GAN iPSCs displayed patient-specific increased vimentin expression, while GAN NPCs had decreased nestin expression compared to isogenic control. The most striking phenotypes were observed in GAN iPSC-astrocytes and brain organoids, which exhibited dense perinuclear IF accumulations and abnormal nuclear morphology. GAN patient cells with large perinuclear vimentin aggregates accumulated nuclearKLHL16mRNA. In over-expression studies, GFAP oligomerization and perinuclear aggregation were potentiated in the presence of vimentin. As an early effector ofKLHL16mutations, vimentin may serve as a potential therapeutic target in GAN.