Summary
Thermal inactivation of the dnaA gene product leads to a considerable decrease in the rate of replication of ColE1-like plasmids. To test the possiblity that the dnaA protein may affect synthesis of RNA I, which is an inhibitor of primer formation, or synthesis of RNA II, which is the primer precursor for replication of ColE1 (Tomizawa and Itoh 1982), the effect of the dnaA46 mutation on the efficiency of the RNA I and the RNA II promoters was examined. It appears that thermal inactivation of the dnaA protein results in a considerable increase in the activity of the RNA I promoter. We suggest that overproduction of RNA I in dnaA mutants grown at the restrictive temperature is responsible for the reduced replication of ColE1-like plasmids.
It has been found that addition of rifampicin to cultures of the dnaA46 or the dna + strain grown at 42°C results in a dramatic increase in the rate of replication of ColE1-like plasmids. We show that the activity of the RNA II promoter at 42°C is exceptionally resistant to rifampicin. In the presence of the drug, this leads, to an altered ratio of RNA I to RNA II, in favor of the latter RNA species.
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References
Abe M (1980) Replication of ColE1 plasmid deoxyribonucleic acid in a thermosensitive dnaA mutant of Escherichia coli. J Bacteriol 141:1024–1030
Atlung T (1979) Function of dnaA product. In: Control of replication and partioning of bacterial chromosomes and plasmids, Abstracts. EMBO Workshop, Leicester, p 1
Bagdasarian MM, Izakowska M, Bagdasarian M (1977) Supression of the DnaA phenotype by mutations in the rpoB cistron of RNA polymerase in Salmonella typhimurium and Escherichia coli. J Bacteriol 130:577–582
Cesareni G, Muesing MA, Polisky B (1982) Control of ColE1 replication: The rop gene product negatively affects transcription from the replication primer promoter. Proc Natl Acad Sci USA 79:6313–6317
Chakraborty T, Yoshinaga K, Lother H, Messer W (1982) Purification of the Escherichia coli dnaA gene product. EMBO J 1:1545–1549
Conrad SE, Campbell JL (1979) Role of plasmid—coded RNA and ribonuclease III in plasmid DNA replication. Cell 18:61–71
Frey J, Chandler M, Caro L (1979) The effects of an Escherichia coli dnaAts mutation on the replication of plasmids ColE1, pSC101, R100.1 and RTF-TC. Mol Gen Genet 174:117–126
Goebel W (1973) The influence of dnaA and dnaC mutations on the initation of plasmid DNA replication. Biochem Biophys Res Commun 51:1000–1007
Hansen EB, Hansen FG, von Meyenburg K (1982) The nucleotide sequence of the dnaA gene and the first part of the dnaN gene of Escherichia coli K-12. Nucl Acids Res 10:7373–7385
Itoh T, Tomizawa J (1978) Initiation of replication of plasmid ColE1 DNA by RNA polymerase, ribonuclease H and DNA polymerase. I. Cold Spring Harbor Symp Quant Biol 43:409–418
Itoh T, Tomizawa J (1980) Formation of an RNA primer for initiation of replication of ColE1 DNA by ribonuclease H. Proc Natl Acad Sci USA 77:2450–2454
Kellenberger-Gujer G, Podhajska AJ, Caro L (1978) A cold sensitive dnaA mutant of E. coli which overinitiates chromosome replication at low temperature. Mol Gen Genet 162:9–16
McKenney K, Shimatake H, Court D, Schmeissner U, Brady C, Rosenberg M (1981) In: Chirikjian JG, Papas TS (eds) Gene amplification and analysis, vol II. Elsevier—North Holland, Amsterdam, pp 384–417
Meijer M, Beck E, Hansen FG, Bergmans HEN, Messer W, von Meyenburg K, Schaller H (1979) Nucleotide sequence of the origin of replication of the Escherichia coli K-12 chromosome. Proc Natl Acad Sci USA 76:580–584
Muesing M, Tamm J, Shepard M, Polisky B (1981) A single basepair alternation is responsible for the DNA overproduction phenotype of a plasmid copy-number mutant. Cell 24:235–242
Polaczek P, Cieśla Z (1983) Rifampicin induced replication of the plasmid pBR322 in Escherichia coli strains carrying dnaA mutations. Mol Gen Genet 190:326–330
Sargent MG (1968) Rapid fixed-time assay for penicillinase. J Bacteriol 95:1493–1494
Tippe-Schindler R, Zahn G, Messer W (1979) Control of the initiation of DNA replication in Escherichia coli I. Negative control of initiation. Mol Gen Genet 168:185–195
Tomizawa J, Itoh T (1982) The importance of RNA secondary structure in ColE1 primer formation. Cell 31:575–583
Tomizawa J, Itoh T, Selzer G, Som T (1981) Inhibition of ColE1 RNA primer formation by a plasmid-specified small RNA. Proc Natl Acad Sci USA 78:1421–1425
Uhlin BE, Nordström K (1977) R plasmid gene dosage effects in Escherichia coli K-12: Copy mutants of the R plasmid R1 drd-19. Plasmid 1:1–7
Wong EM, Muesing MA, Polisky B (1982) Temperature-sensitive copy number mutants of ColE1 are located in an untranslated region of the plasmid gene. Proc Natl Acad Sci USA 79:3570–3574
Wouters JTM, Driehuis FL, Polaczek PJ, van Oppenraay M-LHA, van Andel JG (1980) Persistence of the pBR322 plasmid in Escherichia coli K-12 grown in chemostat cultures. Antonie van Leeuwenhoek 46:353–362
Zyskind JW, Deen LT, Smith DW (1977) Temporal sequence of events during the initiation process in Escherichia coli DNA replication: Roles of the dnaA and dnaC gene products and RNA polymerase. J Bacteriol 129:1466–1475
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Polaczek, P., Cieśla, Z. Effect of altered efficiency of the RNA I and RNA II promoters on in vivo replication of ColE1-like plasmids in Escherichia coli . Molec. Gen. Genet. 194, 227–231 (1984). https://doi.org/10.1007/BF00383521
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DOI: https://doi.org/10.1007/BF00383521