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Temperature-Dependent Hypermutational Phenotype in recA Mutants of Thermus thermophilus HB27

Centro de Biología Molecular Severo Ochoa, Consejo Superior de Investigaciones Científicas-Universidad Autónoma de Madrid, Campus de Cantoblanco, 28049 Madrid,¹ Departamento de Genética y Microbiología, Universidad Autónoma de Barcelona, 08290-Barcelona, Spain²

Received 29 January 2003/ Accepted 11 May 2003

The recA gene from Thermus thermophilus HB27 was cloned and engineered to obtain insertion (recA::kat) and deletion (recA) derivatives. Transcription of recA in this extreme thermophile was induced by mitomycin C, leading to the synthesis of a monocistronic mRNA. This DNA damage-mediated induction was dependent on the integrity of recA. In addition to UV sensitivity, the recA mutants of T. thermophilus showed severe pleiotropic defects, ranging from irregular nucleoid condensation and segregation to a dramatic reduction in viability during culture. An increase in the frequency of both carotenoidless and auxotrophic mutants within surviving cells of the recA strain indicated a high mutation rate. As RecA is not required for plasmid transformation, we have used the -lacZ gene fragment and the ampicillin resistance gene from Escherichia coli as passenger reporters to confirm such high mutation rates. Our data support the idea that the absence of RecA results in a hypermutational phenotype in T. thermophilus. Furthermore, a direct relationship is deduced between the growth temperature and mutation rate, which finally has a deleterious effect on cell survival in the absence of RecA.

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