Repair of ultraviolet-damaged DNA in Micrococcus lysodeikticus

Abstract 1. 1. An investigation was carried out to clarify the in vivo role of an endonuclease found in extracts of Micrococcus lysodeikticus, which induces single-strand breaks in ultraviolet-irradiated DNA but not in non-irradiated DNA (referred to as UV-endonuclease). For this purpose, the occurrence of single-strand break, pyrimidine dimer excision and DNA breakdown after ultraviolet irradiation were examined in mutants of this organism. 2. 2. An ultraviolet-sensitive mutant, G7, did not show single-strand breaks and pyrimidine dimer excision after ultraviolet irradiation. This mutant was also defective in UV-endonuclease activity in its extract. However, transformants to ultraviolet resistance of G7, which were obtained by treatment with wild-type DNA and subsequent selection on nutrient agar containing mitomycin C, did not display UV-endonuclease activity in their extracts, but they could nevertheless produce single-strand breaks in their DNA and rejoin them in vivo. Thus, a discrepancy was observed between in vivo and in vitro endonuclease activity. The same relation was also obtained with Strain 1312 which displayed little UV-endonuclease activity in extracts but was ultraviolet-resistant. It seems that the in vivo activity of endonuclease specific for ultraviolet-irradiated DNA is essential for dark repair. 3. 3. Slower rates of pyrimidine dimer excision and subsequent rejoining of the gaps were observed in the transformant and Strain 1312. UV-endonuclease might participate in the release of dimer from ultraviolet-irradiated DNA, but the mechanism is unknown. 4. 4. An ultraviolet-sensitive, hcr− mutant, UVsN1, showed DNA degradation. In alkaline sucrose gradient centrifugation, however, single-strand breaks and rejoining were observed in this type of mutant. 5. 5. The relationship between the in vivo activity and the in vitro activity (UV-endonuclease) inducing single-strand breaks in ultraviolet-irradiated DNA is considered and the natures of the mutants are discussed from this point of view.