Identification of a nisI Promoter within the nisABCTIP Operon That May Enable Establishment of Nisin Immunity Prior to Induction of the Operon via Signal Transduction

Abstract

ABSTRACT Certain strains of Lactococcus lactis produce the broad-spectrum bacteriocin nisin, which belongs to the lantibiotic class of antimicrobial peptides. The genes encoding nisin are organized in three contiguous operons: nisABTCIP , encoding production and immunity ( nisI ); nisRK , encoding regulation; and nisFEG , also involved in immunity. Transcription of nisABTCIP and nisFEG requires autoinduction by external nisin via signal transducing by NisRK. This organization poses the intriguing question of how sufficient immunity (NisI) can be expressed when the nisin cluster enters a new cell, before it encounters external nisin. In this study, Northern analysis in both Lactococcus and Enterococcus backgrounds revealed that nisI mRNA was present under conditions when no nisA transcription was occurring, suggesting an internal promoter within the operon. The nisA transcript was significantly more stable than nisI , further substantiating this. Reverse transcriptase PCR analysis revealed that the transcription initiated just upstream from nisI . Fusing this region to a lacZ gene in a promoter probe vector demonstrated that a promoter was present. The transcription start site (TSS) of the nisI promoter was mapped at bp 123 upstream of the nisI translation start codon. Ordered 5′ deletions revealed that transcription activation depended on sequences located up to bp −234 from the TSS. The presence of poly(A) tracts and computerized predictions for this region suggested that a high degree of curvature may be required for transcription initiation. The existence of this nisI promoter is likely an evolutionary adaptation of the nisin gene cluster to enable its successful establishment in other cells following horizontal transfer.