Abstract
The nucleotide sequence was determined of the region upstream of the mukB gene of Escherichia coli. Two new genes were found, designated kicA and kicB (killing of cell); the gene order is kicB-kicA-mukB. Promoter activities were detected in the regions immediately upstream of kicB and kicA, but not in front of mukB. Gene disruption experiments revealed that the kicA disruptant was nonviable, but the kicB-disrupted mutant and the mutant lacking both the kicB and kicA genes were able to grow. When kicA disruptant cells bearing a temperature-sensitive replication plasmid carrying the kicA + gene were grown at 30° C and then transferred to 42° C, the mutant cells gradually lost colony-forming ability, even in the presence of a mukB + plasmid. Rates of protein synthesis, but not of RNA or DNA synthesis, fell dramatically during incubation at 42° C. These results suggested that the kicB gene encodes a killing factor and the kicA gene codes for a protein that suppresses the killing function of the kicB gene product. It was also demonstrated that KicA and KicB can function as a post-segregational killing system, when the genes are transferred from the E. coli chromosome onto a plasmid.
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References
Bernard P, Couturier M (1992) Cell killing by the F plasmid CcdB protein involves poisoning of DNA-topoisomerase II complexes. J Mol Biol 226:735–745
Chang ACY, Cohen SN (1978) Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the p15A cryptic miniplasmid. J Bacteriol 134:1141–1156
Gerdes K, Rasmussen P, Molin S (1986) Unique type of plasmid maintenance function: postsegregational killing of plasmid-free cells. Proc Natl Acad Sci USA 83:3116–3120
Gerdes K, Poulsen LK, Thisted T, Nielsen AK, Martinussen J, Andreasen PH (1990) The hok killer gene family in gram-negative bacteria. New Biologist 2:946–956
Hashimoto-Gotoh T, Franklin FCH, Nordheim A, Timmis KN (1981) Specific-purpose plasmid cloning vectors. I. Low copy number, temperature-sensitive, mobilization-defective pSC101-derived containment vectors. Gene 16:227–235
Hiraga S, Jaffè A, Ogura T, Mori H, Takahashi H (1986) F plasmid ccd mechanism in Escherichia coli. J Bacteriol 166:100–104
Hiraga S, Niki H, Ogura T, Ichinose C, Mori H, Ezaki B, Jaffé A (1989) Chromosome partitioning in Escherichia coli: novel mutants producing anucleate cells. J Bacteriol 171:1496–1505
Hirano M, Shigesada K, Imai M (1987) Construction and characterization of plasmid and lambda phage vector systems for study of transcriptional control in Escherichia coli. Gene 57:89–99
Jaffé A, Ogura T, Hiraga S (1985) Effects of the ccd function of the F plasmid on bacterial growth. J Bacteriol 163:841–849
Kohara Y, Akiyama K, Isono K (1987) The physical map of the whole E. coli chromosome: application of a new strategy for rapid analysis and sorting of a large genomic library. Cell 50:495–508
Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
Lehnherr H, Maguin E, Jafri S, Yarmolinsky M (1993) Plasmid addition genes of bacteriophage P1: doc, which causes cell death on curing of prophage, and phd, which prevents host death when prophage is retained. J Mol Biol 233:414–428
Maki S, Takiguchi S, Miki T, Horiuchi T (1992) Modulation of DNA supercoiling activity of Escherichia coli DNA gyrase by F plasmid proteins. J Biol Chem 267:12244–12251
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning, a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Marck C (1988) ‘DNA Strider’: A ‘C’ program for the fast analysis of DNA and protein sequences on the Apple Macintosh family of computers. Nucleic Acids Res 16:1829–1836
Masuda Y, Tsuchimoto S, Nishimura A, Ohtsubo E (1993) Isolation of temperature-sensitive aminoacyl-tRNA synthetase mutants from an Escherichia coli strain harboring the pemK plasmid. Mol Gen Genet 238:169–176
Miki T, Park JA, Nagao K, Murayama N, Horiuchi T (1992) Control of segregation of chromosomal DNA by sex factor F in Escherichia coli. J Mol Biol 225:39–52
Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Niki H, Ogura T, Hiraga S (1990) Linear multimer formation of plasmid DNA in Escherichia coli hopE (recD) mutants. Mol Gen Genet 224:1–9
Niki H, Jaffé A, Imamura R, Ogura T, Hiraga S (1991) The new gene mukB codes for a 177 kd protein with coiled-coil domains involved in chromosome partitioning of E. coli. EMBO J 10:183–193
Niki H, Imamura R, Kitaoka M, Yamanaka K, Ogura T, Hiraga S (1992) E. coli MukB protein involved in chromosome partition forms a homodimer with a rod-and-hinge structure having DNA binding and ATP/GTP binding activities. EMBO J 11:5101–5109
Ogura T, Hiraga S (1983) Partition machanism of F plasmid: two plasmid gene-encoded products and a cis-acting region are involved in partition. Cell 32:351–360
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Stahl FW, Kobayashi I, Thaler D, Stahl MM (1986) Direction of travel of RecBC recombinase through bacteriophage lambda DNA. Genetics 113:215–227
Takeshita S, Sato M, Toba M, Masahashi W, Hashimoto-Gotoh T (1987) High-copy-number plasmid vectors for lacZ α-complementation and chloramphenicol- or kanamycin-resistance selection. Gene 61:63–74
Tsuchimoto S, Ohtsubo E (1989) Effect of the pem system on stable maintenance of plasmid R100 in various Escherichia coli hosts. Mol Gen Genet 215:463–468
Tsuchimoto S, Ohtsubo H, Ohtsubo E (1988) Two genes, pemK and pemI, responsible for stable maintenance of resistance plasmid R100. J Bacteriol 170:1461–1466
Tsuchimoto S, Nishimura Y, Ohtsubo E (1992) The stable maintenance system pem of plasmid R100: degradation of PemI protein may allow PemK protein to inhibit cell growth. J Bacteriol 174:4205–4211
Vieira J, Messing J (1987) Production of single-stranded plasmid DNA. Methods Enzymol 153:3–11
Yanisch-Perron C, Vieira J, Messing J (1985) Improved M13 phage cloning vectors and host strains: nucleotide; sequence of the M13mp18 and pUC19 vectors. Gene 33:103–119
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Communicated by M. Sekiguchi
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Feng, J., Yamanaka, K., Niki, H. et al. New killing system controlled by two genes located immediately upstream of the mukB gene in Escherichia coli . Molec. Gen. Genet. 243, 136–147 (1994). https://doi.org/10.1007/BF00280310
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DOI: https://doi.org/10.1007/BF00280310