RESUMO
There are two modes of bacteriophage lambda DNA replication during its lytic development in Escherichia coli cells. The circle-to-circle (theta) replication predominates at early stages of the phage growth, whereas rolling-circle (sigma) replication occurs late after infection to produce long concatemers that serve as substrates for packaging of lambda DNA into phage proheads. The mechanism regulating the switch from theta to sigma replication remains unknown. Our previous genetic studies indicated that the bacteriophage lambda Pts1piA66 mutant cannot replicate at 43 degrees C in the wild-type E. coli host, but it can replicate in the dnaA46(ts) mutant. Density shift experiments suggested that the parental DNA molecules of the infecting phage enter sigma replication. Here, using electron microscopy, we demonstrate that as soon as 5 min after infection of the dnaA46(ts) mutant by the lambdaPts1piA66 phage at 43 degrees C, the sigma replication intermediates are highly predominant over theta replication intermediates, contrary to the wild-type conditions (wild-type bacteria infected with the lambdaP(+) phage). The initiation of replication of the lambdaPts1piA66 mutant at 43 degrees C was strongly inhibited in the dnaA(+) host, as demonstrated by electron microscopy and by pulse-labeling of the phage-derived plasmid replicon. Implications for the mechanism of the regulation of the switch from theta to sigma replication mode are discussed.