LETHALITY OF REPRECB AND REPRECC DOUBLE MUTANTS OF ESCHERICHIA-COLI

A rep mutation in combination with a recB or a recC mutation renders Escherichia coli non-viable. This conclusion is based on the following lines of evidence: (i) double mutants cannot be constructed by P1 transduction; (ii) induction of the lambda Gam protein, which inactivates most of the RecBCD activities, is lethal in rep mutants; (iii) rep recBts recCts mutants are not viable at high temperature, The reasons for a requirement for the RecBCD enzyme in rep strains were investigated. Initiation of chromosome replication, elongation and chromosomal segregation do not seem impaired in the rep recBts recCts mutant at the non-permissive temperature. The viability of other rep derivatives was tested, rep recA recD triple mutants are not viable, whereas rep recD and rep recA double mutants are. Inactivation of both exoV activity and recBC-dependent homologous recombination is therefore responsible for the non-viability of rep recBC strains. However, sbcA and sbcB mutations, which render recBC mutants recombination proficient, do not restore viability of rep recBC mutants, indicating that recombination via the RecF or the RecE pathways cannot functionally replace RecBCD-mediated recombination. The specific requirement for RecBCD suggests the occurrence of double-strand DNA breaks in rep strains. Additional arguments in favour of the presence of DNA lesions in rep mutants are as follows: (i) expression of SOS repair functions delays lethality of rep derivatives after inactivation of RecBCD; (ii) sensitivity of rep strains to ultraviolet light is increased by partial inactivation of RecBCD. A model for the recovery of cells from double-strand breaks in rep mutants is discussed.