Eclipse period without sequestration in Escherichia coli.
The classical Meselson-Stahl density shift experiment was used to determine the length of the eclipse period in Escherichia coli, the minimum time period during which no new initiation is allowed from a newly replicated origin of chromosome replication, oriC. Populations of bacteria growing exponentially in heavy ((15)NH(4)+ and (13)C(6)-glucose) medium were shifted to light ((14)NH(4)+ and (12)C(6)-glucose) medium. The HH-, HL- and LL-DNA were separated by CsCl density gradient centrifugation, and their relative amounts were determined using radioactive gene-specific probes. The eclipse period, estimated from the kinetics of conversion of HH-DNA to HL- and LL-DNA, turned out to be 0.60 generation times for the wild-type strain. This was invariable for widely varying doubling times (35, 68 and 112 min) and was independent of the chromosome locus at which the eclipse period was measured. For strains with seqA, dam and damseqA mutants, the length of the eclipse period was 0.16, 0.40 and 0.32 generation times respectively. Thus, initiations from oriC were repressed for a considerable proportion of the generation time even when the sequestration function seemed to be severely compromised. The causal relationship between the length of the eclipse period and the synchrony of initiations from oriC is discussed.
Olsson
Jan
J
Department of Cell and Molecular Biology, Biomedical Centre, Uppsala University, Box 596, SE-751 24 Uppsala, Sweden.
Dasgupta
Santanu
S
Berg
Otto G
OG
Nordström
Kurt
K
D001425:Bacterial Outer Membrane Proteins; D002453:Cell Cycle; D002455:Cell Division; D002876:Chromosomes, Bacterial / Q000502:physiology*; D015342:DNA Probes; D004261:DNA Replication; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D029968:Escherichia coli Proteins; D020868:Gene Silencing; D007700:Kinetics; D008433:Mathematics; D008957:Models, Genetic; D051738:Origin Recognition Complex; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics; D014764:Viral Proteins
D001425:Bacterial Outer Membrane Proteins; D015342:DNA Probes; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C114650:OriC chromosomal replication origin; D051738:Origin Recognition Complex; C089746:SeqA protein, E coli; D014157:Transcription Factors; D014764:Viral Proteins; C058693:Dam methyltransferase; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); C499294:dam protein, E coli