Mitochondrial DNA (
mtDNA) damaged by
reactive oxygen species (ROS) triggers so far poorly understood processes of
mtDNA maintenance that are coordinated by a complex interplay among DNA repair,
DNA degradation, and DNA replication. This study was designed to identify the
proteins involved in
mtDNA maintenance by applying a special long-range PCR, reflecting
mtDNA integrity in the minor
arc. A
siRNA screening of literature-based candidates was performed under conditions of enforced oxidative phosphorylation revealing the functional group of polymerases and therein polymerase ζ (POLZ) as top hits. Thus, POLZ knockdown caused
mtDNA accumulation, which required the activity of the base excision repair (BER) nuclease APE1, and was followed by compensatory
mtDNA replication determined by the single-cell mitochondrial in situ hybridization protocol (mTRIP). Quenching
reactive oxygen species (ROS) in mitochondria unveiled an additional, ROS-independent involvement of POLZ in the formation of a typical deletion in the minor
arc region. Together with data demonstrating the localization of POLZ in mitochondria, we suggest that POLZ plays a significant role in
mtDNA turnover, particularly under conditions of oxidative stress.