The genetic switch regulating activity of early promoters of the temperatelactococcal bacteriophage TP901-1

A functional analysis of open reading frame 4 (ORF4) and ORF5 from the temp erate lactococcal phage TP901-1 was performed by mutant and deletion analys is combined with transcriptional studies of the early phage promoters p(R) and p(L). ORF4 (180 amino acids) was identified as a phage repressor necess ary for repression of both promoters. Furthermore, the presence of ORF4 con fers immunity of the host strain to TP901-1, ORF5 (72 amino acids) was foun d to be able to inhibit repression of the lytic promoter p(L) by ORF4, Upon transformation with a plasmid containing both ORF4 and ORF5 and their cogn ate promoters, clonal variation is observed: in each transformant, either p (L) is open and p(R) is closed or vice versa. The repression is still depen dent on ORF4, and the presence of ORF5 is needed for the clonal variation. Induction of a repressed p(L) fusion containing orf4 and orf5 was obtained by addition of mitomycin C, and the induction was also shown to be dependen t on the presence of the RecA protein, even though ORF4 does not contain a recognizable autocleavage site. Our results suggest that the relative amoun ts of the two proteins ORF4 and ORF5 determine the decision between lytic o r lysogenic life cycle after phage infection and that a protein complex con sisting of ORF4 and ORF5 may constitute a new type of genetic switch in bac teriophages.