Identification and characterization of cis ‐ and trans ‐acting elements involved in prophage induction in S treptococcus thermophilus J 34

Summary The genetic switch region of temperate S treptococcus thermophilus phage TP ‐ J 34 contains two divergently oriented promoters and several predicted operator sites. It separates lytic cycle‐promoting genes from those promoting lysogeny. A polycistronic transcript comprises the genes coding for repressor C rh, metalloproteinase‐motif protein R ir and superinfection exclusion lipoprotein Ltp. Weak promoters effecting monocistronic transcripts were localized for ltp and int (encoding integrase) by N orthern blot and 5′‐ RACE‐PCR . These transcripts appeared in lysogenic as well as lytic state. A polycistronic transcript comprising genes coh (encoding C ro homolog), ant (encoding putative antirepressor), orf7 , orf8 and orf9 was only detected in the lytic state. Four operator sites, of which three were located in the intergenic regions between crh and coh , and one between coh and ant , were identified by competition electromobility shift assays. Cooperative binding of C rh to two operator sites immediately upstream of coh could be demonstrated. Coh was shown to bind to the operator closest to crh only. Oligomerization was proven by cross‐linking C rh by glutaraldehyde. Knock‐out of rir revealed a key role in prophage induction. Rir and C rh were shown to form a complex in solution and Rir prevented binding of C rh to its operator sites.