Abstract
Defects in the tricarboxylic acid cycle (TCA) are associated with a spectrum of neurological phenotypes that are often difficult to diagnose and manage. Whole-exome sequencing (WES) led to a rapid expansion of diagnostic capabilities in such disorders and facilitated a better understanding of disease pathogenesis, although functional characterization remains a bottleneck to the interpretation of potential pathological variants. We report a 2-year-old boy of Afro-Caribbean ancestry, who presented with neuromuscular symptoms without significant abnormalities on routine diagnostic evaluation. WES revealed compound heterozygous missense variants of uncertain significance in mitochondrial aconitase (ACO2), which encodes the TCA enzyme ACO2. Pathogenic variants in ACO2 have been described in a handful of families as the cause of infantile cerebellar-retinal degeneration syndrome. Using biochemical and cellular assays in patient fibroblasts, we found that ACO2 expression was quantitatively normal, but ACO2 enzyme activity was <20% of that observed in control cells. We also observed a deficiency in cellular respiration and, for the first time, demonstrate evidence of mitochondrial DNA depletion and altered expression of some TCA components and electron transport chain subunits. The observed cellular defects were completely restored with ACO2 gene rescue. Our findings demonstrate the pathogenicity of two VUS in ACO2, provide novel mechanistic insights to TCA disturbances in ACO2 deficiency, and implicate mitochondrial DNA depletion in the pathogenesis of this recently described disorder.