ndufs2-/- zebrafish model Leigh syndrome with shortened lifespan, developmental anomalies, altered transcriptome and disrupted metabolism

Mitochondrial complex I (CI) deficiency, one genetic etiology of Leigh syndrome spectrum (LSS), manifests with complex multi-system dysfunction with lactic acidemia and early mortality. To facilitate screening of therapeutic candidates for CI diseases, we used CRISPR/Cas9 to generate a ndufs2-/- zebrafish line. Ndufs2 deficient larvae displayed reduced survival, neuromuscular dysfunction, and morphological malformations. We performed transcriptomic sequencing of ndufs2-/- larvae at 6 vs 7 dpf, across and within clutches, and relative to sibling or WT controls. These results were compared to transcriptome data of ndufs2-/- missense mutant C. elegans (gas-1(fc21)) and CI disease patient fibroblasts. Transcriptome profiling revealed consistently dysregulated pathways in ndufs2-/- larvae, gas-1(fc21) worms, and human CI deficient cell lines with increased expression of the electron transport chain, TCA cycle, fatty acid beta oxidation, and one-carbon metabolism. Targeted biochemical analyses revealed a reduction in CI activity and unbiased metabolomic studies identified increased lactate, TCA cycle intermediates, and carnitine species. These changes suggest central metabolism has shunted glucose utilization towards lactate production. Additionally, we determined alterations in one-carbon metabolism associated pathways likely contribute to pathology. Our established, validated ndufs2-/- zebrafish model of CI deficiency recapitulates LSS phenotypes and will be useful for future studies of CI disease and therapeutics. Overall design: RNA-seq profiling of human-derived fibroblast cells grown in galactose media. Patients sequenced are either healthy (n=7) or have confirmed Complex I Primary Mitochondrial Disease (n=2).