Temperature-Sensitive Mutants for the Replication of Plasmids in ESCHERICHIA COLI II. Properties of Host and Plasmid Mutations

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Host mutations in Escherichia coli K12 selected for the temperature-sensitive replication of the bacterial plasmid colicinogenic factor E1 (ColE1) exhibit a pleiotropic effect with respect to the effect of the mutation on other extra-chromosomal elements. The mutations also vary with respect to the time of incubation of the cells at 43°C required for complete cessation of ColE1 DNA synthesis. While the synthesis of the bacterial chromosome appears unaffected, supercoiled ColE1 DNA replication stops immediately in some mutants and gradually decreases during several generations of cell growth before stopping in others. Mutations isolated in the ColE1 plasmid resulted in only a gradual cessation of ColE1 DNA synthesis over several generations of cell growth at 43°C. Conjugal transfer of the ColE1 and ColV factors occurs normally in the host mutants when the transfer is carried out at the permissive temperature; however, the presence of a group I mutation in the donor cell prohibited conjugal transfer of either plasmid DNA at 43°C to a normal recipient cell. Similarly, the presence of this mutation in the recipient prevented the establishment of ColE1 or ColV in the mutant recipient cell upon conjugation with a normal donor at 43°C. Various host ColE1 replication mutants carrying either ColE1 or ColE2 were also defective in the mitomycin C-induced production of colicin E1 or colicin E2 at 43°C. The majority of the host mutations examined exhibited a temperature sensitivity to growth in deoxycholate in addition to the inhibition of plasmid DNA replication, suggesting a membrane alteration in these mutants when grown at the restrictive temperature.

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