ORGANIZATION AND EXPRESSION OF A GENE-CLUSTER INVOLVED IN THE BIOSYNTHESIS OF THE LANTIBIOTIC LACTOCIN-S

Some 8.8 kb of the Lactobacillus sake plasmid pCIMI was sequenced, rev ealing eight tightly clustered open reading frames (ORFs) downstream f rom lasA, which encodes pre-lactocin S. Transcription analyses demonst rated that the genes are expressed as an operon. with transcription in itiating upstream of lasA and terminating immediately 3' to the ninth ORF . lasA is also represented by two small RNAs (RNAI and RNAII) whic h differ in size by approximately 90 nucleotides, and primer extension experiments demonstrated a corresponding difference in the 5' termini . A palindromie sequence constitutes the 3' terminus of both RNAI and RNAII. and we propose that this sequence has a dual regulatory functio n in controlling the expression of las operon, acting both as a barrie r to 3'-5' exonuclease degradation of the lasA-specific transcript(s), and as a ''leaky'' transcriptional terminator which limits the expres sion of down-stream genes. Three of the genes in the las operon have i dentifiable counterparts in other lantibiotic systems: lasM is likely to be involved in peptide modification, lasT, which encodes an ATP-dep endent transport protein, is probably involved in the secretion of lac tocin S, while lasP specifies a subtilisin-type serine protease which may be the lactocin S leader peptidase. Insertional mutation of either lasT or lasM by the resident transposable element IS1163 abolishes la ctocin S production. The remaining five ORFs in the las operon are app arently unique. and their significance with respect to the lactocin S phenotype is presently not known.