Characterization of long patch excision repair of DNA in ultraviolet-irradiated Escherichia coli: An inducible function under rec-lex control

Abstract

Excision repair in ultraviolet-irradiated wild-type Escherichia coli produces a bimodal distribution of repair patch sizes in the DNA. Approximately 99% of the repair events result in short patches of 20-30 nucleotides produced by a constitutive repair system. The remaining 1% result in patches which are at least 1,500 nucleotides in length. This long patch repair is shown to be a damage-inducible process under control of the rec-lex regulatory circuit. The kinetics of the two processes differ; short patch synthesis begins immediately after irradiation and is virtually completed prior to synthesis of the majority of the long patches. Long patch repair synthesis is a linear function of UV dose up to a plateau at 60 J/m2, and hence each long patch event is the consequence of a single UV-induced lesion. Long patch repair does not appear to be necessarily error-prone, since no alteration in repair synthesis occurs as a result of a mutation umuC- which renders cells nonmutable by UV. Evidence is presented suggesting that DNA polymerase I is responsible for both long and short patch synthesis in wild type cells under inducing conditions. In the absence of polymerase I the constitutive patch size averages 80-90 nucleotides, and this distribution is unchanged by induction.

Elimination of multicopy plasmid R6K by bleomycin

“…R6K and multicopy plasmids in general show a requirement for polymerase I during replication (21,25,26), whereas low-copy plasmids replicate independently of this enzyme, utilizing polymerase III (17,21). Polymerase I is essential for the repair of DNA strand gaps (3,9), and its availability for replicative synthesis is therefore likely to be reduced in cells that have been exposed to the DNA strand-breaking drug BLM. This could well lead to a fall in R6K copy number and eventual plasmid elimination.…”

Translesion synthesis is the main component of SOS repair in bacteriophage lambda DNA

“…We could not verify this requirement for UmuDC proteins to replicate lambda under our conditions, since we were unable to isolate a dnaA uvrA umuC triple mutant, which appears not to be viable. Moreover, the higher efficiency of Weigle reactivation of double-stranded DNA phage compared with that of the single-stranded phage is probably due to inducible excision repair, which does not occur on single-stranded DNA (9,10).…”

“…We present evidence that replication proceeds despite the presence of persisting lesions in the phage DNA, which suggests that translesion synthesis is the primary component of Weigle reactivation of lambda in the absence of excision repair. The greater efficiency of Weigle reactivation in double-stranded DNA phage may then be due to inducible excision repair which cannot repair singlestranded DNA (9,10). In addition, after irradiation, the mode of lambda replication seems to switch prematurely from the theta type to the rolling circle type.…”

Recombinational DNA repair: the ignored repair systems

“…(54) The early repair seems to be short-patch excision repair, which occurs immediately after UV irradiation and is controlled by DNA polymerase I, (48) while the induced repair appears to be the long-patch system that is controlled by recA. (50) Additional copies of the UvrA protein (55) and the UvrB protein (56) are synthesized after UV irradiation, and may be relevant to the inducible long-patch excision-repair process. The excision repair that occurs in cells that contain completely replicated chromosomes, i.e.…”

“…macromolecular synthesis is required) and it produces long repair patches (1500-9000 nucleotides long). (48)(49)(50) Long-patch excision repair also requires the recF gene, (51) but does NOT require the recBC genes. (52) When wild-type cells are allowed to repair their DNA after UV irradiation in the presence of chloramphenicol to inhibit the synthesis of induced proteins, only about 80% of the dimers are excised.…”