Changes in mitochondrial morphology and bioenergetics in human lymphoblastoid cells with four novel opa1 mutations

Purpose. Mutations in the optic atrophy 1 gene (OPA1) have been reported in patients with autosomal dominant optic atrophy (ADOA). OPA1 plays important roles in mitochondrial dynamics and cell apoptosis. The link between OPA1 mutations and changes in bioenergetics is still not fully resolved. The aim of this study was to investigate the effects of OP41 mutations on the mitochondrial tubular network and bioenergetics. Methods. We established lymphoblastoid cell lines from four ADOA families harboring different OP41 mutations, unaffected relatives (internal control cell lines), and unrelated normal controls (normal control cell lines). OP41 splice variants and mRNA were analyzed by reverse transcription-PCR and quantitative real-time PCR. Protein isoforms were examined by Western blotting. The mitochondrial network was visualized by confocal microscopy. Mitochondrial bioenergetics were assessed using a Seahorse XF24 flux analyzer. Mitochondrial membrane potential and oxidative damage were analyzed by flow cytometry. Results. OP41 mutant cell lines showed significant decreases in OP41 mRNA and protein expression, mitochondrial membrane potential, and ATP synthesis. A marked deficiency of the long isoform of OPA1 was observed in cells with OP41 mutations in the middle domain and GTPase effector domain. Confocal microscopy revealed increased mitochondrial fragmentation in OP41 mutant cells. OP41 mutant cells also displayed reduced oxygen consumption and underwent glycolysis to produce ATP. Moreover, OP41 mutations caused the accumulation of oxidative damage. Conclusions. Our experiments demonstrated that OP41 mutations induced mitochondrial fragmentation, uncoupled mitochondrial respiration, and elicited dysfunctional bioenergetics. However, there were no significant differences among the various OP41 mutations.