Resident enhanced repair: novel repair process action on plasmid DNA transformed into Escherichia coli K-12

Abstract

The survival of UV-irradiated DNA of plasmid NTP16 was monitored after its transformation into recipient cells containing an essentially homologous undamaged plasmid, pLV9. The presence of pLV9 resulted in a substantial increase in the fraction of damaged NTP16 molecules which survived in the recipicnt cells. This enhanced survival requires the host uvrA+ and uvrB+ gene products, but not the host recA+ gene product. The requirement for both homologous DNA and the uvrA+ and uvrB+ gene products suggests that a novel repair process may act on plasmid DNA. Possible mechanisms for this process are considered.The repair of DNA damaged by UV irradiation has been well studied in Escherichia coli and involves a complex series of pathways (1,4). Two main groups of repair pathways have been identified which operate in the dark: namely, the excision repair pathways which characteristically require the uvrA+, uvrB+, and uvrC+ gene products; and the postreplicational repair pathways which require the recA+ gene product. The sensitivities to UV irradiation displayed by mutant strains indicate that excision repair and post-replicational repair are of roughly equal importance for the survival of cells.We previously studied the repair of plasmid DNA in E. coli (9, 13) by transforming UVdamaged DNA of the plasmid NTP16 into calcium-treated bacterial cells and showed that recA+-dependent repair contributes little to plasmid survival, whereas excision repair acts effectively on plasmid DNA. NTP16 is a nonconjugative plasmid (molecular weight, 5.6 x 106) which encodes resistance to ampicillin and kanamycin (5).To find out whether recA+-dependent repair of NTP16 was limited by the absence of homologous DNA, we measured the survival of UVirradiated NTP16 in cells already containing the closely related plasmid pLV9. This latter plasmid was derived from NTP16 by hydroxylamine mutagenesis and is thought to carry a point mutation in the ampicillin resistance gene (G. 0. Humphreys, personal communication). Thus, the survival of NTP16 may be gauged by following the inheritance of ampicillin resistance.The resident pLV9 did increase the survival of the incoming NTP16 in wild-type cells and in recA and uvrC mutant cells (Fig. la, b, and c), but not in uvrA or uvrB mutant cells (Fig. lb and c). Additional experiments with pLV9-carrying derivatives of the recombinationor repair-defi-cient mutants revealed no other strains which failed to give enhanced survival of NTP16. In contrast, cells containing the plasmid pBR313, which is unrelated to NTP16, did not give enhanced survival (selecting in this instance for the stable acquisition of kanamycin resistance [Fig. la]).Increased survival has been observed in similar conditions with phage (prophage reactivation, multiplicity reactivation, and marker rescue) (2) and with transducing phage-plasmid hybrids (6). In each of these systems, the increase in survival appears to be because of recombination between the damaged molecule and its undamaged homolog. However, since the enhancement of NTP16 surviva...