Rawlinson Elizabeth L Andersen, Nes Ingolf F, Skaugen Morten
Laboratory of Microbial Gene Technology, P O Box 5051, Agricultural University of Norway, N-1432 As, Norway.
Biochimie. 2002 May-Jun;84(5-6):559-67. doi: 10.1016/s0300-9084(02)01420-7.
The 11 kb las locus, present on the 50 kb plasmid pCIM1, specifies the production of the lantibiotic lactocin S in Lactobacillus sakei L45. The gene cluster is organized into two oppositely orientated operons, lasAMNTUVPJW (lasA-W) and lasXY, the former of which contains the biosynthetic, immunity and transport genes. We have previously shown that inactivation of lasX abolishes lactocin S production and causes a drastic reduction in lasA-specific transcripts (encoding pre-lactocin S). The aim of this study was to determine whether or not the product of lasX, which is significantly similar to Rgg-like regulators, was directly involved in transcriptional regulation of the lactocin S biosynthetic genes. The divergently orientated and overlapping promoters, P(lasA)(-W) and P(lasXY), were transcriptionally fused to the Escherichia coli gusA gene, and the activity of the fusions was assayed in the presence and absence of lasX, which was expressed on a separate plasmid. A significant stimulation of expression (5-6-fold) of the P(lasA-W)-gusA fusion was observed in the presence of lasX, whereas expression of the P(lasXY)-gusA construct was reduced 1.5-2-fold. Our results strongly suggest that LasX is a bifunctional regulatory protein, acting both as an activator of lasA-W transcription and as a repressor of lasXY transcription. While a transcription stimulation activity has been described for several of the Rgg-like proteins, the present study is the first to report an autorepressor function for a member of this protein group.
位于50 kb质粒pCIM1上的11 kb las基因座,决定了清酒乳杆菌L45中羊毛硫抗生素乳链菌素S的产生。该基因簇被组织成两个方向相反的操纵子,lasAMNTUVPJW(lasA-W)和lasXY,前者包含生物合成、免疫和转运基因。我们之前已经表明,lasX的失活消除了乳链菌素S的产生,并导致lasA特异性转录本(编码前乳链菌素S)大幅减少。本研究的目的是确定与Rgg样调节因子显著相似的lasX产物是否直接参与乳链菌素S生物合成基因的转录调控。将方向相反且重叠的启动子P(lasA)(-W)和P(lasXY)与大肠杆菌gusA基因进行转录融合,并在存在和不存在lasX(在单独的质粒上表达)的情况下检测融合体的活性。在存在lasX的情况下,观察到P(lasA-W)-gusA融合体的表达受到显著刺激(5-6倍),而P(lasXY)-gusA构建体的表达降低了1.5-2倍。我们的结果强烈表明,LasX是一种双功能调节蛋白,既是lasA-W转录的激活剂,又是lasXY转录的抑制剂。虽然已经描述了几种Rgg样蛋白的转录刺激活性,但本研究是首次报道该蛋白家族成员具有自动抑制功能。