THE ROLE OF ENDONUCLEASE III, ENDONUCLEASE IV, AND EXONUCLEASE III IN THE PROTECTION AGAINST NEAR-UV MEDIATED DNA DAMAGE IN ESCHERICHIA COLI

Abstract

The largest single risk factor for the development of skin cancer is exposure to ultraviolet light. Speculation that increasing levels of ultraviolet (UV) radiation might be reaching the Earth’s surface began following observations of ozone depletion in the Earth’s stratosphere. This concern has been substantiated by studies showing that increases in UVB irradiation (290 nm < λ < 320 nm) are correlated with ozone depletion. The cause and effect relationship between UVB induced dipyrimidine photoproducts and tumor formation is well established. However, less is known about the effects of longer wavelengths on DNA repair. The effects of near-UV (UVA: 320 nm < λ < 400 nm) are complicated by the fact that these wavelengths are absorbed by photosensitizers, which can indirectly damage DNA through the production of reactive oxygen species (ROS). When studying complex biological phenomena, it is sometimes advantageous to use a relatively simple organism. It is for this reason that much of our present knowledge on the subject of DNA repair has come from studies using the enteric bacterium Escherichia coli as a model. E. coli enzymes known to be important in the repair of oxidative DNA damage may play a role in protecting the cell from the lethal effects of near-UV. Isogenic strains deficient in one or more of exonuclease III (xthA), endonuclease IV (nfo), and endonuclease ΠΙ (nth) were exposed to increasing fluences of far-UV and near-UV to evaluate the contribution of these enzymes to the repair of near-UV mediated DNA damage. A far-UV dose of 50 J/m2 and a near-UV dose of 400 kJ/m2 were found to be equally lethal, reducing survival of the wildtype strain to similar levels (approximately 10%). All strains, with the exception of the nth single mutant, were hypersensitive to the lethal effects of near-UV. Double mutants were no more sensitive to near-UV mediated lethality than single mutants. However, a triple mutant strain (nth nfo xthA) exhibited the greatest sensitivity to the lethal effects of near-UV compared to wildtype. DNA repair mutants lacking both exonuclease ΙΠ and endonuclease IV (nfo xthA and nth nfo xthA) were hypersensitive to the mutagenic and lethal effects of far-UV. Endonuclease IV and exonuclease ΙΠ were required for the protection against near-UV and far-UV. Endonuclease III appears to be specific for the protection against near-UV mediated DNA damage, and we therefore propose a previously unrecognized role for this enzyme