Common elements regulating gene expression in temperate and lytic bacteriophages of Lactococcus species

Abstract

A phage-inducible middle promoter (P(15A10)) from the lytic, lactococcal bacteriophage Φ31, a member of the P335 species, is located in an 888-base pair fragment near the right cohesive end. Sequence analysis revealed extensive homology (>95%) to the right cohesive ends of two temperate phages of the P335 species, Φr1t and ΦLC3. Sequencing upstream and downstream of P(15A10) showed that the high degree of homology between Φ31 and Φr1t continued beyond the phage promoter. With the exception of one extra open reading frame in Φ31, the sequences were highly homologous (95 to 98%) between nucleotides 13448 and 16320 of the published Φr1t sequence. By use of a β-galactosidase (β-Gal) gene under the control of a smaller, more tightly regulated region within the P(15A10) promoter, P(566-888), it was established that mitomycin C induction of a lactococcal strain harboring the prophage Φr1t induced the P(566-888) promoter, as determined from an increase in β-Gal activity. Hybridization of nine other lactococcal strains with 32P-labeled P(566-888) showed that the Lactococcus lactis strains C10, ML8, and NCK203 harbored sequences homologous to that of the phage-inducible promoter. Mitomycin C induced the resident prophages in all these strains and concurrently induced the P(566-888) promoter, as determined from an increase in β-Gal activity. DNA restriction analysis revealed that the prophages in C10, ML8, and NCK203 had identical restriction patterns which were different from that of Φr1t. In addition, DNA sequencing showed that the promoter elements in the three phages were identical to each other and to P566-888 from the lytic phage Φ31. These results point to a conserved mechanism in the regulation of gene expression between the lyric phage Φ31 and at least two temperate bacteriophages and provide further evidence for a link in the evolution of certain temperate phages and lytic phages.