Differing Effects of T4 DNA Ligase in the Modulation of the Damage Induced in Mammalian Cells by either X-Rays or Restriction Endonucleases

Abstract

Background: Of the different lesions induced by X-rays, DNA double-strand breaks (DSBs) are considered the main cause of chromosomal aberrations and cell death. Restriction endonucleases (REs) induce only DNA DSBs and have frequently been used to mimic the effects of ionizing radiation in the study of DNA damage and repair. Methods: The present work makes use of clonogenic and cytogenetic assays to study the effect of T4 DNA ligase on modulating the damage induced by either X-rays or an RE (MspI) that produces breaks with cohesive ends. A CHO cell line defective in ligase III activity (EM9) and its corresponding parental line (AA8) were used. Results: Our results show that T4 DNA ligase increased cell survival and decreased chromosomal aberrations in cells treated with MspI, suggesting that most RE-induced DSBs can be repaired by a simple ligation. This enzyme was, however, unable to promote repair of the DNA damage induced by X-rays. Analysis of the ratios of exchange-type aberrations to chromatid break-type aberrations indicated that T4 ligase increased misrejoining of the DNA damage induced by X-rays. The results were similar in EM9 and AA8 cells, although the effect was greater in the cells deficient in DNA strand break rejoining. In addition, depending on whether the end strand break structure is 3′-hydroxyl and 5′-phosphoryl (REs) or more complex (X-rays), T4 DNA ligase could either promote the correct repair or, conversely, increase misrejoining. Conclusion: The present results confirm the idea that DNA DSBs induced by cohesive cutting RE are repaired by different mechanisms than those induced by X-rays causing cell lethality.