Functional analysis of a novel POLγA mutation associated with a severe perinatal mitochondrial encephalomyopathy
Niklas Darin1, Triinu Siibak2, Bradley Peter2
1Department of Pediatrics, Institute of Clinical Sciences, University of Gothenburg, Gothenburg, Sweden.
Abstract:
Mutations in the mitochondrial DNA polymerase gamma catalytic subunit (POLγA) compromise the stability of mitochondrial DNA (mtDNA) by leading to mutations, deletions and depletions in mtDNA. Patients with mutations in POLγA often differ remarkably in disease severity and age of onset. In this work we have studied the functional consequence of POLγA mutations in a patient with an uncommon and a very severe disease phenotype characterized by prenatal onset with intrauterine growth restriction, lactic acidosis from birth, encephalopathy, hepatopathy, myopathy, and early death. Muscle biopsy identified scattered COX-deficient muscle fibers, respiratory chain dysfunction and mtDNA depletion. We identified a novel POLγA mutation (p.His1134Tyr) in trans with the previously identified p.Thr251Ile/Pro587Leu double mutant. Biochemical characterization of the purified recombinant POLγA variants showed that the p.His1134Tyr mutation caused severe polymerase dysfunction. The p.Thr251Ile/Pro587Leu mutation caused reduced polymerase function in conditions of low dNTP concentration that mimic postmitotic tissues. Critically, when p.His1134Tyr and p.Thr251Ile/Pro587Leu were combined under these conditions, mtDNA replication was severely diminished and featured prominent stalling. Our data provide a molecular explanation for the patient´s mtDNA depletion and clinical features, particularly in tissues such as brain and muscle that have low dNTP concentration.
Insights
Mutations in mitochondrial DNA polymerase gamma (POLγA) cause severe disease by impairing mtDNA replication. A novel mutation, p.His1134Tyr, combined with existing mutations, explains severe mtDNA depletion in patients.
Area of Science:
- Genetics
- Molecular Biology
- Biochemistry
Background:
- Mutations in the mitochondrial DNA polymerase gamma catalytic subunit (POLγA) destabilize mitochondrial DNA (mtDNA), causing various diseases with differing severity and onset.
- POLγA mutations lead to mtDNA mutations, deletions, and depletions, impacting cellular respiration and energy production.
Purpose of the Study:
- To investigate the functional consequences of POLγA mutations in a patient with a severe, prenatal-onset phenotype.
- To elucidate the molecular mechanisms underlying mtDNA depletion and the patient's clinical manifestations.
Main Methods:
- Identified POLγA mutations in a patient presenting with intrauterine growth restriction, lactic acidosis, encephalopathy, and early death.
- Performed biochemical characterization of purified recombinant POLγA variants.
- Assessed mtDNA replication fidelity and efficiency under varying conditions, including low deoxynucleotide triphosphate (dNTP) concentrations.
Main Results:
- A novel POLγA mutation (p.His1134Tyr) was identified in trans with a double mutant (p.Thr251Ile/Pro587Leu).
- The p.His1134Tyr mutation severely impaired polymerase function, while the p.Thr251Ile/Pro587Leu mutation reduced function at low dNTP levels.
- Combined mutations drastically diminished mtDNA replication and caused significant stalling, particularly under low dNTP conditions mimicking postmitotic tissues.
Conclusions:
- The study provides a molecular explanation for the severe mtDNA depletion and clinical phenotype observed in the patient.
- The findings highlight the critical role of POLγA function, especially in tissues with low dNTP concentrations like brain and muscle.
- This research deepens the understanding of POLγA-related mitochondrial disorders and their pathomechanisms.
