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Open Access Publications from the University of California

Functional Characterization of Kansl2 and Its Role in Mitochondrial DNA Repair and Maintenance

  • Author(s): Liau, Wei-Siang
  • Advisor(s): Koehler, Carla M.
  • et al.

Mitochondrial DNA repair is a highly regulated, stress-responsive event that is crucial for maintaining mtDNA integrity. Oxidative damage and the mutation load of mtDNA are greater than nuclear DNA. The base excision repair (BER) machinery is important for repair of mtDNA as well as nuclear DNA; however, the pathway has not been as well characterized in mitochondria compared to the nucleus. Recent studies have shown that several repair factors are localized to mitochondria and form stable complex with mtDNA. In this study, I characterize a newly identified mtDNA protein, Kansl2, and its role in mtDNA repair. Kansl2 is conserved from flies to vertebrates, expressed ubiquitously in developing zebrafish and localizes to the mitochondrial matrix. Inactivation of Kansl2 in developing zebrafish caused abnormal heart and muscle development, with eventual death because Kansl2 is essential. Knockdown of Kansl2 enhanced apoptosis. Loss of Kansl2 resulted in aberrant mitochondrial cristae and compromised complex II and complex IV activity. Kansl2 binds to the mtDNA promiscuously via its Rad51-like domain. More importantly, I found that Kansl2 assembles in a 300 kDa molecular weight complex with uracil DNA glycosylase (Ung) in zebrafish mitochondria. I further characterized the molecular interaction of Kansl2 and Ung by determining the effect of Kansl2 on Ung activity in-vitro. These results affirm the hypothesis that Kansl2 serves as a scaffold for Ung and the interaction enhances the removal of uracil from the mtDNA by Ung. This study employs western blot analysis, animal studies, knockdown technology, native PAGE analysis and electron microscopy.

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