Open AccessDiscontinuity in DNA replication during expression of accumulated initiation potential in dnaA mutants of Escherichia coli
Author(s) -
Charles E. Helmstetter,
C. A. Krajewski,
Alan C. Leonard,
Martin Weinberger
Publication year - 1986
Publication title -
journal of bacteriology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.652
H-Index - 246
eISSN - 1067-8832
pISSN - 0021-9193
DOI - 10.1128/jb.165.2.631-637.1986
Subject(s) - dnaa , biology , dna replication , novobiocin , pre replication complex , microbiology and biotechnology , chloramphenicol , origin of replication , replication factor c , dna synthesis , ter protein , control of chromosome duplication , prokaryotic dna replication , escherichia coli , dna , genetics , antibiotics , gene
Potential for initiation of chromosome replication present in temperature-sensitive, initiation-defective dnaA5 mutants of Escherichia coli B/r incubated at nonpermissive temperature was expressed by shifting to a more permissive temperature (25 degrees C). Upon expression of initiation potential, the rate of [3H]thymidine incorporation varied in a bimodal fashion, i.e., there was an initial burst of incorporation, which lasted 10 to 20 min, then a sudden decrease in incorporation, and finally a second rapid increase in incorporation. Analyses of this incorporation pattern indicated that a round of replication initiated upon expression of initiation potential, but DNA polymerization stopped after replication of 5 to 10% of the chromosome. This round of replication appeared to resume about 30 min later coincident with initiation of a second round of replication. The second initiation was unusually sensitive to low concentrations of novobiocin (ca. 1 microgram/ml) when this inhibitor was added in the presence of chloramphenicol. In the absence of chloramphenicol, novobiocin at this concentration had no detectable effect on DNA replication. It is suggested that cis-acting inhibition, attributable to an attempted second initiation immediately after the first, caused the first round to stall until both it and the second round could resume simultaneously. This DNA replication inhibition, probably caused by overinitiation, could be a consequence of restraints on replication in the vicinity of oriC, possibly topological in nature, which limit the minimum interinitiation interval in E. coli.