Abstract

Recombinational repair-dependent mutants identify ways to avoid chromosomal lesions. Starting with a recBC(Ts) strain of Escherichia coli, we looked for mutants unable to grow at 42 degrees C in conditions that inactivate the RecBCD(Ts) enzyme. We isolated insertions in ackA and pta, which comprise a two-gene operon responsible for the acetate<-->acetyl coenzyme A interconversion. Using precise deletions of either ackA or pta, we showed that either mutation makes E. coli cells dependent on RecA or RecBCD enzymes at high temperature, suggesting dependence on recombinational repair rather than on the RecBCD-catalyzed linear DNA degradation. Complete inhibition of growth of pta/ackA rec mutants was observed only in the presence of nearby growing cells, indicating cross-inhibition. pta rec mutants were sensitive to products of the mixed-acid fermentation of pyruvate, yet none of these substances inhibited growth of the double mutants in low-millimolar concentrations. pta, but not ackA, mutants also depend on late recombinational repair functions RuvABC or RecG. pta/ackA recF mutants are viable, suggesting, together with the inviability of pta/ackA recBC mutants, that chromosomal lesions due to the pta/ackA defect are of the double-strand-break type. We have isolated three insertional suppressors that allow slow growth of pta recBC(Ts) cells under nonpermissive conditions; all three are in or near genes with unknown functions. Although they do not form colonies, ackA rec and pta rec mutants are not killed under the nonpermissive conditions, exemplifying a case of synthetic inhibition rather than synthetic lethality.

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