Insertion of inverted Ter sites into the terminus region of the Escherichia coli chromosome delays completion of DNA replication and disrupts the cell cycle.
To investigate the co-ordination between DNA replication and cell division, we have disrupted the DNA-replication cycle of Escherichia coli by inserting inverted Ter sites into the terminus region to delay completion of the chromosome. The inverted Ter sites (designated InvTer::spcr) were initially inserted into the chromosome of a delta tus strain to allow unrestrained chromosomal replication. We then introduced a functional tus gene by transforming the InvTer::spcr strain with a plasmid carrying the tus gene under control of an arabinose-inducible promoter. In the presence of 0.2% arabinose, the cells formed long filaments, suggesting that activation of the inverted Ter sites by Tus arrested DNA replication and delayed the onset of cell division. Induction of sfiA, a gene in the SOS regulon, was observed following arrest of DNA replication; however, when a sfiB114 allele was introduced into InvTer::spcr strain, long filaments were still formed, suggesting that the sfi-independent pathway also caused filamentation. Either recA::camr or lexA3 alleles suppressed filamentation when introduced in the InvTer strain. Interestingly, in both the recA::camr and lexA3 mutants, virtually all cells had a nucleoid, suggesting that cell division was proceeding even though DNA replication was not complete. These results suggest that DNA replication and cell division are uncoupled when recA is inactivated or when genes repressed by LexA cannot be induced.
Sharma
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Department of Bioscience and Biotechnology, Drexel University, Philadelphia, Pennsylvania 19104, USA.
Hill
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D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics; D001483:Base Sequence; D002453:Cell Cycle / Q000235:genetics*; D007446:Chromosome Inversion; D002876:Chromosomes, Bacterial; D004261:DNA Replication; D004269:DNA, Bacterial / Q000096:biosynthesis / Q000737:chemistry; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009690:Nucleic Acid Conformation; D011938:Rec A Recombinases / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D012697:Serine Endopeptidases; D013489:Suppression, Genetic; D013997:Time Factors; D000099160:LexA Repressor Protein
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D011938:Rec A Recombinases; D011994:Recombinant Proteins; D012697:Serine Endopeptidases; D000099160:LexA Repressor Protein; C053139:sulA protein, E coli; C061910:tus protein, Bacteria; C489236:tus protein, E coli