The Y951N patient mutation inactivates the intramolecular switch in human mitochondrial DNA POLγ

Abstract

AbstractMitochondrial DNA (mtDNA) stability, essential for cellular energy production, relies on DNA polymerase gamma (POLγ). Here, we show that the POLγ Y951N disease causing mutation induces replication stalling and severe mtDNA depletion. However, unlike other POLγ disease causing mutations, Y951N does not directly impair exonuclease activity and only mildly affects polymerase activity. Instead, we found that Y951N compromises the enzyme’s ability to efficiently toggle between DNA synthesis and degradation, and is thus the first patient-derived mutation with impaired polymerase-exonuclease switching. These findings provide new insights into the intramolecular switch when POLγ proofreads the newly-synthesized DNA strand, and reveal a new mechanism for causing mitochondrial DNA instability.Significance StatementDNA polymerase gamma (POLγ) is essential for copying mitochondrial DNA (mtDNA), which is crucial for our energy production. POLγ must accurately switch between making new DNA (polymerase activity) and correcting errors (exonuclease activity). While it is known that mutations in POLγ can cause mitochondrial diseases by directly impairing these enzymatic functions, this study reveals a new mechanism. The Y951N mutation disrupts POLγ’s ability to switch between these activities, leading to severe blockages in DNA replication and a loss of mtDNA in human cells, even without significant direct impairment of polymerase or exonuclease activities. These findings provide new insights into the origins of mitochondrial diseases.