Minicells produced by abnormal cell division in a strain of Escherichia coli (K-12) have been employed here to investigate the phenomenon of "entry exclusion." When purified minicells from strains containing F' or R factors, or both, are mated with radioactive thymidine-labeled Hfr or R+ donors, the recipient minicells can be conveniently separated from normal-sized donors following mating, and the products of conjugation can be analyzed in the absence of donors and of further growth of the recipients. Transmissible plasmids or episomes are transferred less efficiently to purified minicells derived from strains carrying similar or related elements than to strains without them. Measurement of deoxyribonucleic acid (DNA) degradation and determination of weight-average molecular weights following transfer indicate that degradation of transferred DNA or transfer of smaller pieces cannot account for the comparative reduction in transfer to entry-excluding recipients. Therefore, we conclude that entry exclusion operates to prevent the physical entry of DNA into recipients expressing the exclusion phenotype. The R-produced repressor (product of the drd+ gene), which represses fertility (i.e., ability to act as donor), reduces exclusion mediated by R or F factor, or both, in matings between strains carrying homologous elements. Furthermore, the data suggest that the presence of the F pilus or F-like R pilus on recipient cells ensures maximum expression of the exclusion phenotype but is not essential for its expression. In contrast to previous suggestions, we found no evidence for a reduction of entry exclusion attributable to the DNA temperature-sensitive chromosomal mutation dnaB(TS).In 1958 Lederberg observed that conjugal matings between two F + strains were less efficient than matings between an F+ strain and a strain that did not harbor an F fertility factor. Currently it is accepted that transmissible plasmids or episomes usually are transferred less efficiently to strains that carry similar or related elements, suggesting that a determinant linked to the sex factor confers on the cell immunity against conjugal infection by an isogenic or closely related sex factor. Novick (23) has suggested that this immunity is the result of two distinct effects, "entry exclusion" and "plasmid incompatibility."Incompatibility is considered to be operative after deoxyribonucleic acid (DNA) enters the recipient cell and is evidenced by the segregation of the newly transferred plasmid out of the population. It has been postulated that the failure of the incoming plasmid to replicate normally and its subsequent loss reflect a lack of available attachment sites (16). The occupa-tion of a maintenance site(s) by the resident plasmid presumably blocks the attachment of a similar or isogenic plasmid. However, the maintenance site model does not explain all cases of incompatibility (see Novick [23] for detailed discussion).On the other hand, it has been suggested that entry exclusion imposes a barrier to the physical transfer of DNA betwee...
The main objective of these experiments was the further examination of whether the induction of alkaline phosphatase activity in HeLa cells by 5-iodo-2'-deoxyuridine (IdUrd) depends on the incorporation of IdUrd into DNA. Thymidine (dThd), deoxycytidine (dCyd), cytidine, and beta-cytosine arabinoside (Ara-C) inhibited a dose-dependent manner the induction of alkaline phosphatase activity by IdUrd in HeLa cells, and 5-iodo-2'-deoxycytidine induced activity in a dose-dependent manner at concentrations similar to those of IdUrd. Three of these compounds, dThd, dCyd, and Ara-C, were studied with regard to degree of inhibition of induction and IdUrd incorporation into DNA. Although the various doses of these three compounds decreased the incorporation of IdUrd into DNA, there was no apparent linear correlation between the extent of inhibition of IdUrd incorporation and the degree of inhibition of the induction of alkaline phosphatase activity. dCyd also inhibited a dose-dependent manner the induction of alkaline phosphatase by hydrocortisone, sodium butyrate, and choline chloridee. These results, although not unequivocal, support the idea that IdUrd induction of alkaline phosphatase activity in HeLa cells does not require IdUrd incorporation into DNA. The dCyd altered the thermostability for alkaline phosphatase activity from control or IdUrd-treated cells, and for controls cells the change in thermostability occurred without a change in the enzyme specific activity.
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