Polymerase zeta Is Involved in Mitochondrial DNA Maintenance Processes in Concert with APE1 Activity

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 zeta (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.