Regulation of the switch from early to late bacteriophage lambda DNA replication.
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 orilambda proceeds both bidirectionally and unidirectionally, whereas in bacteria devoid of a functional DnaA protein, replication from orilambda is predominantly unidirectional. The regulation of directionality of replication from orilambda is mediated by positive control of lambda p(R) promoter activity by DnaA, since the mode of replication of an artificial lambda replicon bearing the p(tet) promoter instead of p(R) 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 orilambda, 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 orilambda resulting in unidirectional theta replication followed by sigma type replication.
Department of Molecular Biology, University of Gdańsk1 and Laboratory of Molecular Biology (affiliated with the University of Gdańsk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences2, Kładki 24, 80-822 Gdańsk, Poland.
Sylwia
S
Barańska
Department of Molecular Biology, University of Gdańsk1 and Laboratory of Molecular Biology (affiliated with the University of Gdańsk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences2, Kładki 24, 80-822 Gdańsk, Poland.
Magdalena
M
Gabig
Department of Molecular Biology, University of Gdańsk1 and Laboratory of Molecular Biology (affiliated with the University of Gdańsk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences2, Kładki 24, 80-822 Gdańsk, Poland.
Alicja
A
Węgrzyn
Department of Molecular Biology, University of Gdańsk1 and Laboratory of Molecular Biology (affiliated with the University of Gdańsk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences2, Kładki 24, 80-822 Gdańsk, Poland.
Grażyna
G
Konopa
Department of Molecular Biology, University of Gdańsk1 and Laboratory of Molecular Biology (affiliated with the University of Gdańsk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences2, Kładki 24, 80-822 Gdańsk, Poland.
Anna
A
Herman-Antosiewicz
Departamento de Biologı́a Celular y del Desarrollo, Centro de Investigaciones Biológicas (CSIC), Velázquez 144, 28006 Madrid, Spain3.
Pablo
P
Hernandez
Departamento de Biologı́a Celular y del Desarrollo, Centro de Investigaciones Biológicas (CSIC), Velázquez 144, 28006 Madrid, Spain3.
Jorge B
JB
Schvartzman
Center for Molecular Genetics and Department of Biology, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA4.
Donald R
DR
Helinski
Marine Biology Center, Polish Academy of Sciences, Św. Wojciecha 5, 81-347 Gdynia, Poland5.|Department of Molecular Biology, University of Gdańsk1 and Laboratory of Molecular Biology (affiliated with the University of Gdańsk), Institute of Biochemistry and Biophysics, Polish Academy of Sciences2, Kładki 24, 80-822 Gdańsk, Poland.
Grzegorz
G
Węgrzyn
D001426:Bacterial Proteins; D010582:Bacteriophage lambda / Q000235:genetics* / Q000378:metabolism; D004261:DNA Replication; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000821:virology*; D015967:Gene Expression Regulation, Viral; D008854:Microscopy, Electron; D018741:Replication Origin; D015533:Transcriptional Activation
D001426:Bacterial Proteins; D004279:DNA, Viral; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria