A Study of genetic instability and DNA repair in selected mutants of streptomyces cattleya

Abstract

Three mutants of Streptomyces cattleya NRRL 8057 with lesions in the DNA repair pathway were obtained by NTG mutagenesis. The mutants were selected on the basis of their resistance or sensitivity to Mitomycin c. Of the three mutants, two, R6 and Rl2, were selected as MMC resistant and one, S26, as MMC sensitive. They were also UVr and UVˢ, respectively. The mutants were subjected to UV irradiation in order to obtain UV kill curves. The curves generated for the two MMCʳ mutants showed induction or derepression of a second repair system. In instability studies performed on them, the two MMCʳ mutants showed increased instability compared to the wild type at low UV dosage. However, the instability dropped to below that of the wild type at high UV dosages. Both these mutants appeared to roughly mimic the plateau shown by the wild type as higher levels of UV irradiation were achieved. S26 in contrast showed a much higher initial incidence of instability, and no plateau was observed at high UV dosages. Single strand breaks were induced in the wild type and mutant strains by incorporation of P³² into their DNA. R6 showed a five-fold increase in instability after this treatment. Rl2 showed no significant change, and S26 showed a slight decrease, which may be due to the increased lethality of DNA damage sustained by this strain due to its relatively labile nature. The strains were further analysed using polyacrylamide gel electrophoresis of total cellular proteins. S26 showed a startlingly different protein profile. R6 was almost identical to the wild type, while Rl2 was extremely similar, showing only the loss of one major band and a significant lowering of the amount of detectable protein in the region below this band.