UV Sensitivity and Mutagenesis of the Extremely Thermophilic Eubacterium Thermus thermophilus HB27

Abstract

Thermus thermophilus is an aerobic, extremely thermophilic eubacterium that grows optimally at 70-75°C. Phylogenetically there is a close relationship between T. thermophilus and Deinococcus radiodurans, an extraordinary radiation-resistant bacterium. In contrast to the D. radiodurans, DNA repair and mutagenesis in T. thermophilus have not been investigated intensively. DNA repair and related mutagenesis at high temperature are particularly interesting because the frequency of DNA damage, such as deamination, depurination, and single strand breaks, is expected to be greatly increased. We disrupted the uvrA gene for subunit A of excinuclease ABC and uvdE gene for probable UV endonuclease by inserting the thermostable kanamycin-resistant gene (HTK) or orotate phosphoribosyltransferase (pyrE) gene. A uvrA mutant showed moderate sensitivity to UV-irradiation, while a uvdE mutant was not UV sensitive. A uvdE uvrA double mutant was highly sensitive to UV-irradiation compared with a uvrA mutant, indicating that UV endonuclease, a uvdE gene product, plays an important role in repair of UV-induced DNA damages. On the other hand, no obvious UV-induced mutations were observed when it was assayed by His+ reversion. It has been reported that T. thermophilus HB27 does not have an SOS response system, because no genes homologous to umuD, umuC, and lexA were found. In the absence of error-prone translesion DNA polymerases in T. thermophilus, mutations would not occur at the site of pyrimidine dimers