Functional characterization of two variants in the mitochondrial topoisomerase gene TOP1MT that impact regulation of the mitochondrial genome

TOP1MT encodes a mitochondrial topoisomerase that is important for mtDNA regulation, and is involved in mitochondrial replication, transcription and translation. Two variants predicted to affect TOP1MT function (R199C and V338L) were identified by exome sequencing of a newborn with hypertrophic cardiomyopathy. As no pathogenic TOP1MT variants have been confirmed previously, we characterized these variants for their ability to rescue several TOP1MT functions in knockout cells. Consistent with a role for these TOP1MT variants contributing to the patient phenotype, comprehensive characterization suggests that both variants had impaired topoisomerase activity and demonstrates that neither variant was able to restore steady state levels of mitochondrial encoded proteins, nor reduced oxidative phosphorylation. However, the two variants behaved differently in some respects. While the R199C variant was better at restoring transcript levels, the V338L variant was able to restore mtDNA copy number and replication. These findings suggest that the different TOP1MT variants affect distinct TOP1MT functions. Altogether, these findings begin to provide insight into the many roles that TOP1MT plays in the maintenance and expression of the mitochondrial genome, and how impairments in this important protein may lead to human pathology.