Induction of a large deletion in mitochondrial genome of mouse cells by X-ray irradiation

Abstract

Large deletions and point mutations of mitochondrial DNA (mtDNA) is causally associated with mitochondrial diseases, and accumulates with age in human tissues. In order to find out the role of oxidative stress in the generation of large mtDNA deletions, using normal Balb and severe combined immunodeficiency (SCID) mouse cells, we assessed whether X-ray irradiation induces large mtDNA deletions. Cultured cells were irradiated with X-rays and assayed for a large mtDNA deletion using a PCR technique with a specific pair of primers. X-ray doses as low as 1 Gy were effective for the induction of the large mtDNA deletion (5823 bp long), which corresponds to the human 4977 bp common deletion. The breakpoints were flanked by the 5-bp long tandem repeats, 5′-TACCC-3′. A dose-dependent induction of the large mtDNA deletion was observed, the yield being higher in normal cells than in SCID cells. The fraction of large mtDNA deletion increased in the normal cells but decreased in the SCID cells within 7 days after X-ray irradiation. As the SCID cells carry a mutation in the gene encoding DNA–PKcs, the key enzyme in DNA double-strand break repair, it can be concluded that repair of DNA strand breaks may be involved in the formation of X-ray induced large mtDNA deletions.