Jaspers M C, Schmid A, Sturme M H, Goslings D A, Kohler H P, Roelof Van Der Meer J
Swiss Federal Institute for Environmental Science and Technology and Swiss Federal Institute of Technology, CH-8600 Dübendorf, Switzerland.
J Bacteriol. 2001 Jan;183(1):270-9. doi: 10.1128/JB.183-1.270-279.2001.
Pseudomonas azelaica HBP1 degrades the toxic substance 2-hydroxybiphenyl (2-HBP) by means of three enzymes that are encoded by structural genes hbpC, hbpA, and hbpD. These three genes form a small noncontiguous cluster. Their expression is activated by the product of regulatory gene hbpR, which is located directly upstream of the hbpCAD genes. The HbpR protein is a transcription activator and belongs to the so-called XylR/DmpR subclass within the NtrC family of transcriptional activators. Transcriptional fusions between the different hbp intergenic regions and the luxAB genes of Vibrio harveyi in P. azelaica and in Escherichia coli revealed the existence of two HbpR-regulated promoters; one is located in front of hbpC, and the other one is located in front of hbpD. Northern analysis confirmed that the hbpC and hbpA genes are cotranscribed, whereas the hbpD gene is transcribed separately. No transcripts comprising the entire hbpCAD cluster were detected, indicating that transcription from P(hbpC) is terminated after the hbpA gene. E. coli mutant strains lacking the structural genes for the RNA polymerase sigma(54) subunit or for the integration host factor failed to express bioluminescence from P(hbpC)- and P(hbpD)-luxAB fusions when a functional hbpR gene was provided in trans. This pointed to the active role of sigma(54) and integration host factor in transcriptional activation from these promoters. Primer extension analysis revealed that both P(hbpC) and P(hbpD) contain the typical motifs at position -24 (GG) and -12 (GC) found in sigma(54)-dependent promoters. Analysis of changes in the synthesis of the hbp mRNAs, in activities of the 2-HBP pathway enzymes, and in concentrations of 2-HBP intermediates during the first 4 h after induction of continuously grown P. azelaica cells with 2-HBP demonstrated that the specific transcriptional organization of the hbp genes ensured smooth pathway expression.
壬二酸假单胞菌HBP1通过由结构基因hbpC、hbpA和hbpD编码的三种酶降解有毒物质2-羟基联苯（2-HBP）。这三个基因形成一个小的不连续簇。它们的表达由调控基因hbpR的产物激活，hbpR位于hbpCAD基因的直接上游。HbpR蛋白是一种转录激活因子，属于转录激活因子NtrC家族中所谓的XylR/DmpR亚类。在壬二酸假单胞菌和大肠杆菌中，不同hbp基因间区域与哈维弧菌luxAB基因之间的转录融合揭示了存在两个受HbpR调控的启动子；一个位于hbpC之前，另一个位于hbpD之前。Northern分析证实hbpC和hbpA基因是共转录的，而hbpD基因是单独转录的。未检测到包含整个hbpCAD簇的转录本，这表明从P（hbpC）的转录在hbpA基因之后终止。当通过反式提供功能性hbpR基因时，缺乏RNA聚合酶σ54亚基或整合宿主因子结构基因的大肠杆菌突变株无法从P（hbpC）-和P（hbpD）-luxAB融合体中表达生物发光。这表明σ54和整合宿主因子在这些启动子的转录激活中起积极作用。引物延伸分析表明，P（hbpC）和P（hbpD）在-24位（GG）和-12位（GC）都含有在依赖σ54的启动子中发现的典型基序。在用2-HBP诱导连续生长的壬二酸假单胞菌细胞后的前4小时内，对hbp mRNA的合成变化、2-HBP途径酶的活性以及2-HBP中间体的浓度进行分析，结果表明hbp基因的特定转录组织确保了途径的顺利表达。
