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

Area of Science:

• Mitochondrial biology
• Molecular genetics
• DNA repair mechanisms

Background:

• Mitochondrial DNA (mtDNA) damage from reactive oxygen species (ROS) initiates complex maintenance processes involving DNA repair, degradation, and replication.
• The precise molecular players coordinating mtDNA maintenance under oxidative stress remain incompletely understood.

Purpose of the Study:

• To identify key proteins involved in mitochondrial DNA maintenance.
• To elucidate the role of polymerase ζ (POLZ) in mtDNA integrity and turnover.

Main Methods:

• Utilized long-range PCR to assess mtDNA integrity.
• Performed siRNA screening targeting candidate proteins under enforced oxidative phosphorylation.
• Employed single-cell mitochondrial in situ hybridization protocol (mTRIP) to analyze mtDNA replication.
• Investigated the effect of ROS quenching on POLZ function.

Main Results:

• siRNA screening identified polymerases, specifically polymerase ζ (POLZ), as critical for mtDNA maintenance.
• POLZ knockdown resulted in mtDNA accumulation, dependent on APE1 activity, and triggered compensatory mtDNA replication.
• ROS quenching revealed a ROS-independent role for POLZ in forming specific mtDNA deletions.
• POLZ was localized to mitochondria, confirming its direct involvement in mitochondrial processes.

Conclusions:

• Polymerase ζ (POLZ) plays a significant role in mitochondrial DNA turnover.
• POLZ is essential for maintaining mtDNA integrity, particularly under conditions of oxidative stress.
• The study implicates POLZ in a complex interplay of mtDNA repair, replication, and degradation pathways.