engleski

Genetic Requirements for High Constitutive SOS Expression in recA730 Mutants of Escherichia coli

The RecA protein in its functional state is in complex with single-stranded DNA, i.e., in the form of a RecA-filament. In SOS induction, the RecA filament functions as a co-protease enabling the auto-digestion of LexA repressor. The RecA filament can be formed by different mechanisms, but all of them require three enzymatic activities essential for the processing of DNA double stranded ends. These are helicase, 5'-3' exonuclease and RecA loading onto ssDNA. In some mutants, the SOS response can be expressed constitutively during the process of normal DNA metabolism. The RecA730 mutant protein is able to form RecA filament without the help of RecBCD and RecFOR mediators since it better competes with SSB protein for ssDNA. As a consequence, the recA730 mutants show high constitutive SOS expression. In this paper we studied the genetic requirements for constitutive SOS expression in recA730 mutants. Using a β-galactosidase assay, we showed that constitutive SOS response in recA730 mutants exhibits different requirements in different backgrounds. In a wild type background constitutive SOS is partially dependent on RecBCD function. In a recB1080 background (the recB1080 mutation retains only helicase), constitutive SOS is partially dependent on RecBCD helicase function, and is strongly dependent on RecJ nuclease. Finally, in a recB null background, constitutive SOS expression of recA730 mutant is dependent on the RecJ nuclease. Our results emphasize the importance of the 5'-3' exonuclease for high constitutive SOS expression in recA730 mutants, and show that RecBCD function can further enhance the excellent intrinsic abilities of the RecA730 protein in vivo.

recA730; constitutive SOS expression; RecA filament; E. coli

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