Error-prone Activity Research Articles

Mutagenic DNA repair in Escherichia coli. V. Mutation frequency decline and error-free post-replication repair in an excision-proficient strain

Mutation frequency decline (MFD) is an irreversible loss of newly-induced suppressor mutations occurring in excision-proficient Escherichia coli during a short period of incubation in minimal medium before plating on broth- or Casamino acids-enriched selective agar. It is known that MFD of UV-induced mutations may occur before DNA containing pre-mutagenic lesions is replicated, but we conclude that MFD can also occur after the damaged DNA has been replicated on the basis of the following evidence. (1) Mutation fixation in rich medium (i.e., loss of susceptibility to mutation frequency decline) with ethyl methanesulphonate mutagenesis begins immediately, whereas with UV it is delayed for 20--30 min. (2) The delay in mutation fixation after UV can be explained neither by inhibition of DNA replication nor by a delay in the appearance of error-prone repair activity in the irradiated population. (3) MFD at later times after UV irradiation is more rapid and is less strongly inhibited by caffeine than is MFD immediately after irradiation. (4) Excision is virtually complete 20 min after 3 J m-2 UV but at that time virtually all mutations are still susceptible to MFD. We have presented evidence elsewhere that in bacteria there is an alternative error-free excision-dependent type of post-replication repair of potentially mutagenic daughter strand gaps. We suggest that this process is inhibited at tRNA loci in the presence of nutrient broth or Casamino acids, possibly because of a broth-dependent change in the structure of the single-stranded region including the tRNA locus.

Persistence and decay of thermoinducible error-prone repair activity in nonfilamentous derivatives of tif-1, Escherichia coli B/r: the timing of some critical events in ultraviolet mutagenesis.

Ultraviolet (UV) mutagenesis in E. coli is associated with a UV-inducible type of error-prone postreplication repair ("SOS" repair) which, in tif-1 strains, is thermo-inducible in coordination with other recA+ lexA+-dependent inducible functions, including filamentous growth. Mutants of E. coli B/r tif-1 strains have been isolated which retain thermoinducibility of SOS repair activity, but lack the thermosensitivity caused by filamentous growth at 42 degrees C. These strains have been used to determine: the kinetics of decay at 30 degrees C of thermally induced ability to enhance UV mutagenesis; the kinetics of thermal enhancement of spontaneous and UV-induced mutability at 42 degrees C, and the kinetics of decay at 30 degrees C of susceptibility to thermal enhancement of spontaneous and UV-induced mutability. Mutations from tryptophane requirement to prototrophy (Trp- to Trp+) were scored. UV doses were 0.2 J/M2 for excision repair-deficient (Uvr-) and 2J/m2 for Uvr+ strains. The results support the following conclusions. 1) thermally induced SOS repair activity decays at 30 degrees C to about 25% of its maximum level in 45 min, and is no longer detectable after 90 min. 2) Thermal enhancement of UV mutability occurs at sites produced primarily (perhaps exclusively) before completion of the first post-irradiation cell division. 3) UV-induced sites susceptible to thermally induced SOS repair are stable at 30 degrees C in cells not containing the error-prone repair system, and are refractory to constitutive error-free repair for at least 2-3 hours. 4) UV produces a potentially mutagenic type of photoproduct in DNA which can, without interacting with another UV lesion, provide a site susceptible to SOS repair, but which is not a sufficient signal for SOS induction. 5) 50-70% of the SOS-mutable SOS-noninducing UV photoproducts are photoreversible pyrimidine dimers. The results are discussed in relation to current models of UV mutagenesis and induction of UV-inducible functions.

Thermal enhancement of ultraviolet mutability in a dnaB uvrA derivative of Escherichia coli B/r: evidence for inducible error-prone repair.

DNA damage triggers coordinate expression of a cluster of diverse functions in Escherichia coli, including prophage induction, filamentous growth, and "aberrant" reintiation of DNA replication at the chromosomal origin. The "SOS repair" hypothesis proposes that one of these coordinately inducible functions is an error-prone system of DNA repair ("SOS repair") which is responsible for ultraviolet mutagenesis. In dnaB strains, incubation of 42 degrees C stops DNA synthesis and induces lambda prophage and should, therefore, also induce the postulated error-prone repair activity. Thermal posttreatment of a dnaB urvA derivative of E. coli B/r is found to enhance the yield of ultraviolet-light-induced mutations as much as 50-fold, while having no such effect in the dnaB+ parent strain. The results support the SOS repair hypothesis. The possibility is discussed that the inducible repair system is a mutagenic DNA polymerase.