Chromosomal fragmentation is the major consequence of the rdgB defect in Escherichia coli

Abstract

The rdgB mutants depend on recombinational repair of double-strand breaks. To assess other consequences of rdgB inactivation in Escherichia coli, we isolated RdgB-dependent mutants. All transposon inserts making cells dependent on RdgB inactivate genes of double-strand break repair, indicating that chromosomal fragmentation is the major consequence of RdgB inactivation.

Figure 1.

The color screen for RdgB-dependent mutants. The screen looks for the inability to lose a temperature-sensitive plasmid carrying the rdgB⁺ gene. (A) The properties of the strain for the screen [LL12 = ΔlacZ ΔrdgB p(ori-Ts)-lacZ⁺-rdgB⁺] at three temperatures, plated on MacConkey + lactose agar. (B) The scheme of the experimental strain (LL12) and the expected colony phenotype of RdgB-independent (sectoring) mutants, as well as a single RdgB-dependent mutant (converging arrows). (C) The colony phenotype of the parental color screen strain [LL12 ΔrdgB prdgB + (Ts)] and its two derivatives, carrying either ΔrecA (LL13) or ΔrecBCD (LL14) mutations, on MacConkey–lactose at 34°. The parental strain forms sectoring colonies, whereas the double mutants form smaller nonsectoring colonies, serving as positive controls.

Figure 2.

The position and orientation of inserts, conferring RdgB dependence, in recombinational repair genes. The vicinity maps of the disrupted genes showing the chromosomal coordinates were generated with Colibri server (http://genolist.pasteur.fr/Colibri/genome.cgi). Open reading frames and their direction are shown by lightly shaded arrows, identified by gene names. The positions of pRL27 inserts are shown by darkly shaded arrows on sticks, identifying the orientation of the kan gene of the insert. The numbers near inserts identify individual mutants. Note that the three maps have different scales. (A) The 4-kbp recA region. (B) The 13-kbp recBCD region. (C) The 5-kbp ruvABC region.