Convergent evolution of an oxidative damage tolerance mechanism preserves replication fidelity in plant organellar DNA polymerases

Plant mitochondria and chloroplasts are sources of reactive oxygen species (ROS) that trigger oxidative DNA lesions. Among these, 8-oxo-7,8-dihydroguanine (8-oxodG) is particularly mutagenic because it exhibits ambiguous base pairing, as DNA polymerases efficiently incorporate adenine opposite 8-oxodG, leading to G·C→T·A transversions. Despite exposure to oxidative stress, plant organellar genomes exhibit some of the lowest nucleotide substitution rates in nature, implying that their DNA replication machinery may have evolved mechanisms to cope with oxidative lesions such as 8-oxodG. In family-A DNA polymerases, including yeast and animal mitochondrial DNA polymerases, the balance between faithful dCMP incorporation and mutagenic dAMP incorporation opposite 8-oxodG is governed by a bulky or positively charged residue. Surprisingly, although the equivalent position in the Arabidopsis thaliana organellar DNA polymerases AtPolIA and AtPolIB is occupied by glycine—a residue predicted to promote mutagenic dAMP incorporation—both enzymes nevertheless preferentially incorporate accurate dCMP opposite 8-oxodG. Our structure–function analyses demonstrate that AtPolI´s fidelity opposite 8-oxodG is conferred by a lysine located near the templating base, which generates steric and electrostatic constraints that favor the 8-oxodG(anti):dCTP pairing while disfavoring the mutagenic 8-oxodG(syn):dATP pair. This mechanism represents an example of convergent evolution, as the fidelity-determining lysine in plant organellar polymerases is in a different position than the analogous residues in yeast-animal mitochondrial polymerases but achieves the same function. Together, these findings suggest that plant organellar DNA polymerases have evolved specialized properties that promote accurate bypass of oxidative DNA lesions, which may contribute to the maintenance of the exceptionally low mutation rates observed in plant organellar genomes.