D'Orazio S E, Thomas V, Collins C M
Department of Microbiology and Immunology, University of Miami School of Miami School of Medicine, Florida 33101, USA.
Mol Microbiol. 1996 Aug;21(3):643-55. doi: 10.1111/j.1365-2958.1996.tb02572.x.
The Proteus mirabilis and plasmid-encoded urease loci contain seven contiguous structural and accessory genes (ureDABCEFG) and the divergently transcribed ureR, which codes for an AraC-like transcriptional activator. Previously, it was shown that the plasmid-encoded ureR to ureD intergenic region contained divergent promoters (ureRp and ureDp). Transcription from these promoters required both the effector molecule urea and the activator protein UreR. In this report, we demonstrate that the P. mirabilis urease gene cluster contains similar divergent urea- and UreR-dependent promoters. The ureR gene products from either urease locus were able to activate transcription at both the plasmid-encoded and P. mirabilis promoters. The minimal concentration of urea required to activate transcription at ureRp or ureDp from either gene cluster was approximately 4 mM. The transcriptional start sites for the plasmid-encoded and P. mirabilis divergent promoters were similar in an Escherichia coli DH5 alpha background, as determined by primer-extension analysis. However, in P. mirabilis HI4320, transcription of ureR initiated predominately at an alternative site. Physical mapping and inhibition studies were used to localize the UreR-binding sites within the plasmid-encoded ureRp and ureDp intergenic sequences to regions of 68 bp and 86 bp, respectively. Gel shift analysis demonstrated that UreR bound to a 135 bp fragment in the approximate centre of the plasmid-encoded ureR to ureD intergenic region. The results presented here suggest that the P. mirabilis and plasmid-encoded urease gene clusters utilize similar mechanisms of transcriptional activation in response to urea.
奇异变形杆菌和质粒编码的脲酶基因座包含七个相邻的结构和辅助基因（ureDABCEFG）以及反向转录的ureR，其编码一种AraC样转录激活因子。先前的研究表明，质粒编码的ureR至ureD基因间区域包含反向启动子（ureRp和ureDp）。这些启动子的转录需要效应分子尿素和激活蛋白UreR。在本报告中，我们证明奇异变形杆菌脲酶基因簇包含类似的依赖尿素和UreR的反向启动子。来自任一脲酶基因座的ureR基因产物都能够激活质粒编码的启动子和奇异变形杆菌启动子的转录。激活任一基因簇中ureRp或ureDp转录所需的尿素最低浓度约为4 mM。通过引物延伸分析确定，在大肠杆菌DH5α背景下，质粒编码的和奇异变形杆菌反向启动子的转录起始位点相似。然而，在奇异变形杆菌HI4320中，ureR的转录主要在另一个位点起始。通过物理图谱分析和抑制研究，将质粒编码的ureRp和ureDp基因间序列中的UreR结合位点分别定位到68 bp和86 bp的区域。凝胶迁移分析表明，UreR与质粒编码的ureR至ureD基因间区域大致中心的一个135 bp片段结合。本文给出的结果表明，奇异变形杆菌和质粒编码的脲酶基因簇利用类似的机制响应尿素进行转录激活。
