Wang P, Yang J, Pittard A J
Department of Microbiology, University of Melbourne, Parkville, Victoria, Australia.
J Bacteriol. 1997 Jul;179(13):4206-12. doi: 10.1128/jb.179.13.4206-4212.1997.
Analysis of in vitro transcriptional events initiating within the region immediately upstream of the aroP coding region has revealed the presence of three promoters, P1, P2, and P3. Both P1 and P2 give rise to mRNA encoding the AroP protein, whereas P3 initiates transcription in the opposite direction. Both P1 and P3 contain UP elements which contribute to promoter strength. Regulation of expression from these three promoters has been examined in vitro by using supercoiled DNA templates and in vivo by using lacZ transcriptional fusions and specific promoter mutants. Expression from P2 is partially repressed by TyrR alone both in vitro and in vivo. Addition of the aromatic amino acid tyrosine, phenylalanine, or tryptophan further increases this repression. P1 is not repressed by TyrR alone but is repressed in vivo in the presence of phenylalanine, tyrosine, or tryptophan. This also occurs in vitro but requires Ca2+ ions in the reaction mixture for its demonstration. Under these conditions, transcription from P3 is enhanced by TyrR protein with phenylalanine, tyrosine, or tryptophan. However, we were unable to demonstrate P3 expression in vivo. Under repressing conditions, there is no production of truncated RNA molecules (from P1), which would be expected if repression involved a roadblock mechanism.
对位于aroP编码区上游紧邻区域内起始的体外转录事件的分析揭示了存在三个启动子,即P1、P2和P3。P1和P2都产生编码AroP蛋白的mRNA,而P3则以相反方向起始转录。P1和P3都含有有助于启动子强度的上游元件(UP元件)。已通过使用超螺旋DNA模板在体外以及通过使用lacZ转录融合和特定启动子突变体在体内研究了这三个启动子的表达调控。P2的表达在体外和体内都被TyrR单独部分抑制。添加芳香族氨基酸酪氨酸、苯丙氨酸或色氨酸会进一步增强这种抑制作用。P1不会被TyrR单独抑制,但在苯丙氨酸、酪氨酸或色氨酸存在的情况下在体内会被抑制。这在体外也会发生,但反应混合物中需要Ca2+离子才能证明。在这些条件下,TyrR蛋白与苯丙氨酸、酪氨酸或色氨酸一起会增强P3的转录。然而,我们无法在体内证明P3的表达。在抑制条件下,不会产生截短的RNA分子(来自P1),如果抑制涉及一种阻碍机制,预计会产生这种分子。