For plasmids derived from bacteriophage lambda, the initiation of bidirectional DNA replication from orilambda depends on the stimulation of transcription from the p(R) promoter by the host replication initiator protein DnaA. Certain Escherichia coli dnaA(ts) mutants cannot be transformed by wild-type lambda plasmids even at the temperature permissive to cell growth. This plasmid-host incompatibility appeared to be due to inefficient stimulation of transcription from the p(R) promoter by the mutant DnaA protein. This paper shows that there is a second mechanism for the incompatibility between lambda plasmids and dnaA(ts) hosts, exemplified in this study by the dnaA46 mutant. This is based on the competition between the lambda P protein and the host DnaA and DnaC proteins for DnaB helicase. Both mechanisms must be operative for the incompatibility.