Bacteriophage and host mutants causing the rolling-circle lambda DNA replication early after infection

There are two modes of bacteriophage lambda DNA replication during its lyti c development in Escherichia coli cells, The circle-to-circle (theta) repli cation predominates at early stages of the phage growth, whereas rolling-ci rcle (sigma) replication occurs late after infection to produce long concat emers that serve as substrates for packaging of lambda DNA into phage prohe ads, The mechanism regulating the sw itch from theta to sigma replication r emains unknown, Our previous generic studies Indicated that the bacteriopha ge lambda Pts1 pi A66 mutant cannot replicate at 43 degrees C in the wild-t ype E. coli host, but it can replicate in the dnaA46(ts) mutant, Density sh ift experiments suggested that the parental DNA molecules of the infecting phage enter sigma replication. Here, using electron microscopy, we demonstr ate that as soon as 5 min after infection of the dnaA46(ts) mutant by the l ambda Pts1 pi A66 phage at 43 degrees C, the sigma replication intermediate s are highly predominant over theta replication intermediates, contrary to the wild-type conditions (wild-type bacteria infected with the lambda P+ ph age), The initiation of replication of the lambda Pts1 pi A66 mutant at 43 degrees C was strongly inhibited in the dnaA(+) host, as demonstrated by el ectron microscopy and by pulse-labeling of the phage-derived plasmid replic on, Implications for the mechanism of the regulation of the switch from the ta to sigma replication mode are discussed. (C) 2000 Federation of European Biochemical Societies.