Excess SeqA prolongs sequestration of oriC and delays nucleoid segregation and cell division.
Following initiation of chromosomal replication in Escherichia coli, newly initiated origins (oriCs) are prevented from further initiations by a mechanism termed sequestration. During the sequestration period (which lasts about one-third of a cell cycle), the origins remain hemimethylated. The SeqA protein binds hemimethylated oriC in vitro. In vivo, the absence of SeqA causes overinitiation and strongly reduces the duration of hemimethylation. The pattern of immunostained SeqA complexes in vivo suggests that SeqA has a role in organizing hemimethylated DNA at the replication forks. We have examined the effects of overexpressing SeqA under different cellular conditions. Our data demonstrate that excess SeqA significantly increases the time oriC is hemimethylated following initiation of replication. In some cells, sequestration continued for more than one generation and resulted in inhibition of primary initiation. SeqA overproduction also interfered with the segregation of sister nucleoids and caused a delay in cell division. These results suggest that SeqA's function in regulation of replication initiation is linked to chromosome segregation and possibly cell division.
Bach
Trond
T
Department of Cell Biology, Institute for Cancer Research, The Norwegian Radium Hospital, 0310 Oslo, Norway.
Krekling
Martin A
MA
Skarstad
Kirsten
K
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002455:Cell Division / Q000502:physiology*; D002876:Chromosomes, Bacterial / Q000235:genetics / Q000378:metabolism; D004261:DNA Replication; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D005434:Flow Cytometry; D015964:Gene Expression Regulation, Bacterial; D008745:Methylation; D008856:Microscopy, Fluorescence; D009690:Nucleic Acid Conformation; D051738:Origin Recognition Complex; D011485:Protein Binding; D018741:Replication Origin; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; 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