Functional analysis of promoters in the nisin gene cluster of Lactococcus lactis.
The promoters in the nisin gene cluster nisABTCIPRKFEG of Lactococcus lactis were characterized by primer extension and transcriptional fusions to the Escherichia coli promoterless beta-glucuronidase gene (gusA). Three promoters preceding the nisA, nisR, and nisF genes, which all give rise to gusA expression in the nisin-producing strain L. lactis NZ9700, were identified. The transcriptional autoregulation of nisA by signal transduction involving the sensor histidine kinase NisK and the response regulator NisR has been demonstrated previously (0. P. Kuipers, M. M. Beerthuyzen, P. G. G. A. de Ruyter, E. J. Luesink, and W. M. de Vos, J. Biol. Chem. 270: 27299-27304, 1995), and therefore the possible nisin-dependent expression of gusA under control of the nisR and nisF promoters was also investigated. The nisR promoter was shown to direct nisin-independent gusA expression in L. lactis MG 1363, which is a nisin-transposon- and plasmid-free strain. L. lactis NZ9800, which does not produce nisin because of a deletion in the nisA gene, containing the nisF-gusA fusion plasmid, gave rise to beta-glucuronidase production only after induction by nisin. A similar regulation was found in L. lactis NZ3900, which contains a single copy of the nisR and nisK genes but no other genes of the nisin gene cluster. In contrast, when the nisK gene was disrupted, no beta-glucuronidase activity directed by the nisF promoter could be detected even after induction with nisin. These results show that, like the nisA promoter, the nisF promoter is nisin inducible. The nisF and nisA promoter sequences have significant similarities and contain a conserved region that could be important for transcriptional control.
de Ruyter
P G
PG
Department of Biophysical Chemistry, Netherlands Institute for Dairy Research, Ede The Netherlands.
Kuipers
O P
OP
Beerthuyzen
M M
MM
van Alen-Boerrigter
I
I
de Vos
W M
WM
D001483:Base Sequence; D017931:DNA Primers / Q000737:chemistry; D004269:DNA, Bacterial / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D013294:Lactococcus lactis / Q000235:genetics*; D008969:Molecular Sequence Data; D009561:Nisin / Q000235:genetics*; D009876:Operon; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
D017931:DNA Primers; D004269:DNA, Bacterial; D012333:RNA, Messenger; D009561:Nisin