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Microbiology 147 (2001), 535-547
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Microbiology (2001), 147, 535-547.
© 2001 Society for General Microbiology

Genetics and Molecular Biology

Regulation of the switch from early to late bacteriophage {lambda} DNA replication

Sylwia Baraska1, Magdalena Gabig2, Alicja Wegrzyn2, Grayna Konopa1, Anna Herman-Antosiewicz1, Pablo Hernandez3, Jorge B. Schvartzman3, Donald R. Helinski4 and Grzegorz Wegrzyn1,5

Department of Molecular Biology, University of Gdask1 and Laboratory of Molecular Biology (affiliated with the University of Gdask), Institute of Biochemistry and Biophysics, Polish Academy of Sciences2, Kladki 24, 80-822 Gdask, Poland
Departamento de Biología Celular y del Desarrollo, Centro de Investigaciones Biológicas (CSIC), Velázquez 144, 28006 Madrid, Spain3
Center for Molecular Genetics and Department of Biology, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA4
Marine Biology Center, Polish Academy of Sciences, w. Wojciecha 5, 81-347 Gdynia, Poland5

Author for correspondence: Grzegorz Wegrzyn. Tel: +48 58 346 3014. Fax: +48 58 301 0072. e-mail: wegrzyn{at}biotech.univ.gda.pl

There are two modes of bacteriophage {lambda} DNA replication following infection of its host, Escherichia coli. Early after infection, replication occurs according to the theta ({theta} or circle-to-circle) mode, and is later switched to the sigma ({sigma} or rolling-circle) mode. It is not known how this switch, occurring at a specific time in the infection cycle, is regulated. Here it is demonstrated that in wild-type cells the replication starting from ori{lambda} proceeds both bidirectionally and unidirectionally, whereas in bacteria devoid of a functional DnaA protein, replication from ori{lambda} is predominantly unidirectional. The regulation of directionality of replication from ori{lambda} is mediated by positive control of {lambda} pR promoter activity by DnaA, since the mode of replication of an artificial {lambda} replicon bearing the ptet promoter instead of pR was found to be independent of DnaA function. These findings and results of density-shift experiments suggest that in dnaA mutants infected with {lambda}, phage DNA replication proceeds predominantly according to the unidirectional {theta} mechanism and is switched early after infection to the {sigma} mode. It is proposed that in wild-type E. coli cells infected with {lambda}, phage DNA replication proceeds according to a bidirectional {theta} mechanism early after infection due to efficient transcriptional activation of ori{lambda}, stimulated by the host DnaA protein. After a few rounds of this type of replication, the resulting increased copy number of {lambda} genomic DNA may cause a depletion of free DnaA protein because of its interaction with the multiple DnaA-binding sites in {lambda} DNA. It is proposed that this may lead to inefficient transcriptional activation of ori{lambda} resulting in unidirectional {theta} replication followed by {sigma} type replication.

Keywords: bacteriophage {lambda} development, Escherichia coli DnaA protein, rolling-circle DNA replication, theta DNA replication, transcriptional activation of origin




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