Genome-wide Screen Identifies Peroxisomal Role in APOL1 Podocytopathy.

Abstract

Risk variants G1 and G2 of APOL1 confer a markedly increased risk of chronic kidney disease (CKD) in individuals of African ancestry, yet disease expression requires secondary insults such as inflammation or hypoxia. How these stressors intersect with APOL1 risk variants to drive podocyte injury and cell death remains poorly defined. To identify pathways that modify APOL1 variant-induced cytotoxicity under hypoxic stress, we performed an unbiased, genome-wide RNA interference (RNAi) screen in cells expressing APOL1 G1 or G2. Candidate genes were validated using targeted genetic manipulation, pharmacologic interventions, and subcellular localization and structure-function analyses. The RNAi screen identified multiple peroxisomal biogenesis (PEX) genes as modifiers of APOL1 G1 or G2 cytotoxicity, with PEX gene silencing markedly exacerbating cell death during hypoxia. This implicates impaired peroxisomal homeostasis as a previously unappreciated vulnerability in APOL1-associated cellular injury. In contrast, genetic or pharmacologic enhancement of peroxisomal function significantly attenuated cytotoxicity induced by either APOL1 risk variant. Mechanistically, we identified a functional peroxisomal targeting signal at the C-terminus of APOL1 that mediates hypoxia-dependent trafficking to peroxisomes. Disruption of this targeting signal reduced peroxisomal localization of APOL1 G1/G2 variants and mitigated cytotoxicity, linking peroxisomal trafficking to variant-specific injury. Our findings identify peroxisomal dysfunction as an important determinant of APOL1 G1/G2- mediated cytotoxicity under hypoxic stress. By establishing a mechanistic connection between hypoxia, peroxisomal biology, and APOL1 risk variants, our work highlights peroxisomes as a therapeutically actionable pathway to limit podocyte injury and CKD progression in genetically susceptible population.