On the mechanism of post-replication repair in Escherichia coli cells: The role of DNA polymerase III

Abstract

The newly synthesized DNA after UV-irradiation of uvr+ or uvrE. coli cells has a low single-stranded molecular weight equal to the average spacing between pyrimidine dimers in parental strands of DNA [ 1,2] . The decrease of DNA molecular weight in alkaline sucrose gradients is caused by the gaps (1000 nucleotides long) in daughter strands, opposite to the pyrimidine dimers, arising at the replication fork [ 1,3]. In the course of subsequent incubation in growth medium these gaps are repaired. The finding of DNA strands with intermediate density in alkaline CsCl in UV-irradiated E. coli uvrA6 cells allowed the authors to suggest that post-replication gaps are repaired by a recombination mechanism involving actual breakage and reunion of DNA molecules [4]. However, many facts do not accord with this hypothesis [5,6]. For example, E. coli AB 2487 recAI3 cells show no deficiency in post-replication repair after 15 erg/mm? of 254 nm W/-irradiation although the relative recombination frequency in these cells is about 10T4 compared to a value of 1 for ret+ cells [7] ; on the other hand, there is no breakage of parental DNA strands in UV-irradiated uvrA6 or uvrB5 cells at all [7,8]. It is possible therefore that UV-stimulated recombination between sister DNA duplexes, if it occurs, does not causally relate to the filling of gaps and the latter are repaired by de novo DNA synthesis [5 ] . In this paper data are presented indicating that inactivation at the non-permissive temperature (45’CO of thermolabile DNA polymerase III in a double polAI p0lC26~~ mutant of E. coli results in the failure to repair post-replication gaps after W-irradiation.