AMPD2 deficiency causes variable-onset Leigh syndrome and implicates cytosolic purine balance in the structural preservation of respiratory complex V

Importance Cellular mechanisms underlying mitochondrial dysfunction, a hallmark feature of many neurodegenerative conditions, remain incompletely understood, and their true diversity is unknown. Objective To identify and functionally validate novel genetic variants causative of Leigh syndrome. Design We performed whole genome sequencing (WGS) and first-degree relative genotyping on two unrelated adult subjects with brain MRI abnormalities evoking Leigh syndrome. Blue native polyacrylamide gel electrophoresis (BN-PAGE) and respiratory chain enzymatic activity assays were performed to screen for respiratory complex assembly and/or oxidative phosphorylation impairments. Cells obtained from patient dermal and muscular biopsies were immortalized and later genetically corrected to evaluate cellular response to metabolic stress. Setting Subjects were recruited from The Neuro (McGill University), Rizk Hospital (Lebanese American University), and Centre Hospitalier Universitaire Sainte-Justine (University of Montreal). Research connections were established through the White Matter Rounds Network and GeneMatcher. Participants Four subjects representing three families with undiagnosed Leigh syndrome (age range 10-40 years) were ultimately recruited. Main outcome(s) and Measure(s) DNA sequencing uncovered a new autosomal recessive Leigh syndrome-associated gene that was functionally validated. Results Bi-allelic pathogenic variants in AMPD2 were detected in all subjects. BN-PAGE of patient skeletal muscle mitochondria captured an isolated complex V assembly defect in the context of mTOR activation, while the accompanying enzymological assays reported decreased activities of complexes I and IV. Opposite to controls, patient-derived cell lines lacked AMPD2 protein, attributing null status to the variants detected. During metabolic challenge, mutant cells suffered from mitochondrial hyperfusion and high-order cytosolic IMPDH2 oligomerization, implying simultaneous ATP accumulation and GTP deficiency. However, both complex V assembly and mTOR status were impervious to these conditions. All cellular phenotypes observed collectively reverted upon exogenous introduction of wild-type AMPD2. Conclusions and Relevance The recognition of AMPD2-related Leigh syndrome (AMPD2-LS) as a novel entity provides strong evidence for classifying AMPD2 deficiency as a mitochondrial disease. Our data suggest that respiratory capacity is significantly modulated by AMPD2, a cytosolic enzyme selectively regulating complex V assembly through an elusive process.