Regulation of reductive dehalogenase gene transcription in Dehalococcoides mccartyi
Regulation of reductive dehalogenase gene transcription in Dehalococcoides mccartyi
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DOI：
10.1098/rstb.2012.0317
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发表时间：
2013-04
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影响因子：
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通讯作者：
Anke Wagner;Lydia Segler;S. Kleinsteuber;G. Sawers;H. Smidt;U. Lechner
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作者：
Anke Wagner;Lydia Segler;S. Kleinsteuber;G. Sawers;H. Smidt;U. Lechner
Dehalococcoides属的显着能力，脱氯的许多不同的氯化有机化合物反映的数量和多样性的基因编码还原脱卤酶同系物（RDH）的Dehalococcoides物种的基因组。这些基因中的大多数位于编码多抗生素抗性调节子（马尔R）型或双组分系统调节子的基因附近。在这里，rdhA基因（编码的催化亚基）的转录反应，以2，3-和1，3-二氯二苯并-p-二恶英（DCDD）在Dehalococcoides mccartyi菌株CBDB 1进行了研究。几乎所有的rdhA基因在2，3-DCDD存在下转录，尽管在不同的水平，如cbrA、cbdbA 1453、cbdbA 1624和cbdbA 1588的转录物所示。相反，1，3-DCDD不诱导rdhA转录。推定的马尔CbdbA 1625是异源产生的，并证明其在体外结合到基因cbdbA 1624和cbdbA 1625的重叠启动子区的能力。为了分析体内调控，构建不同rdhA基因和cbdbA 1625的单拷贝转录启动子-lacZ融合体，并将其引入异源宿主大肠杆菌中，并测量融合体的表达水平。将cbdbA 1625基因克隆到允许通过阿拉伯糖调节表达的载体中，并将其转化到含有rdh-启动子-lacZ融合衍生物的菌株中。CbdbA 1625被证明从其自身的启动子下调转录，导致β-半乳糖苷酶活性降低40-50%，这首次提示它充当阻遏物。
The remarkable capacity of the genus Dehalococcoides to dechlorinate a multitude of different chlorinated organic compounds reflects the number and diversity of genes in the genomes of Dehalococcoides species encoding reductive dehalogenase homologues (rdh). Most of these genes are located in the vicinity of genes encoding multiple antibiotic resistance regulator (MarR)-type or two-component system regulators. Here, the transcriptional response of rdhA genes (coding for the catalytic subunit) to 2,3- and 1,3-dichlorodibenzo-p-dioxin (DCDD) was studied in Dehalococcoides mccartyi strain CBDB1. Almost all rdhA genes were transcribed in the presence of 2,3-DCDD, albeit at different levels as shown for the transcripts of cbrA, cbdbA1453, cbdbA1624 and cbdbA1588. By contrast, 1,3-DCDD did not induce rdhA transcription. The putative MarR CbdbA1625 was heterologously produced and its ability to bind in vitro to the overlapping promoter regions of the genes cbdbA1624 and cbdbA1625 was demonstrated. To analyse regulation in vivo, single-copy transcriptional promoter–lacZ fusions of different rdhA genes and of cbdbA1625 were constructed and introduced into the heterologous host Escherichia coli, and expression levels of the fusions were measured. The cbdbA1625 gene was cloned into a vector allowing a regulation of expression by arabinose and it was transformed into the strains containing the rdh-promoter–lacZ fusion derivatives. CbdbA1625 was shown to downregulate transcription from its own promoter resulting in a 40–50% reduction in the β-galactosidase activity, giving the first hint that it acts as a repressor.