NIH Reporter #1ZIAAG000699-08

G-quadruplexes (G4) are potential DNA roadblocks that perturb mitochondrial replication machinery and pose a clinically relevant source of mitochondrial DNA mutation and instability

This award supports research investigating how G-quadruplex DNA structures in mitochondrial DNA contribute to mutations linked to cancer and aging by stalling replication machinery and promoting error-prone DNA synthesis.

Description

Mutations in the mitochondrial (mt) genome are correlated with cancer and aging. G-quadruplexes (G4), dynamic DNA structures that arise in guanine (G)-rich templates, potentially stall the mt replisome and thereby promote mutagenesis. Computational analyses of genome sequence data from two Italian cohorts demonstrated an association between G4s and mt variation. Using the software G4Hunter to predict G4-forming regions in mtDNA, statistically significant enrichment of mutations in stable G4 regions was found. In vitro biochemical data demonstrated that G4s potently block the mt replicative polymerase gamma (Pol ). Addition of mt replisome-associated factors including TWINKLE helicase and mt single-strand binding protein were unable to stimulate pol synthesis through the G4 block. Other mt polymerases further catalyze error-prone nucleotide incorporation into G4 structures, suggesting involvement in G4 bypass with increased risk of mutation. The computational and biochemical approaches indicate that mt point mutations are enriched at stable G4 structures, consistent with replisome stalling at G-quadruplexes and reliance on error-prone DNA synthesis.

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