Identification of operator sites of the CI repressor of phage TP901-1: evolutionary link to other phages

The repressor encoded by the cI gene of the temperate Lactococcus lactis subsp. cremoris bacteriophage TP901-1 has been purified. Gel-retardation and footprinting analyses identified three palindromic operator sites (O(R), O(L), and O(D)). The operator site O(R) is located between the two divergent early promoters P(R) and P(L), O(L) overlaps the transcriptional start of the lytic P(L) promoter, and O(D) is located downstream of the mor gene, the first gene in the lytic gene cluster. The function of O(L) was verified by mutational analysis. Binding was found to be specific and cooperative. Multimeric forms of the repressor were observed, thus indicating that the repressor may bind simultaneously to all three operator sites. Inverted repeats with homology to the operator sites of TP901-1 were identified in phage genomes encoding repressors homologous to CI of TP901-1. Interestingly, the locations of these repeats on the phage genomes correspond to those found in TP901-1, indicating that the same system of cooperative repression of early phage promoters has been inherited by modular evolution.