Wang Y P, Giblin L, Boesten B, O'Gara F
Microbiology Department, University College, Cork, Ireland.
Mol Microbiol. 1993 Apr;8(2):253-9. doi: 10.1111/j.1365-2958.1993.tb01569.x.
The expression of the Rhizobium meliloti C4-dicarboxylic acid permease gene (dctA) is controlled by the sensor DctB and the transcriptional regulator, DctD. The R. meliloti Dct system has been reconstituted in Escherichia coli. Expression of the dctA promoter is DctBD dependent and is induced in the presence of C4-dicarboxylic acids (dCA). Other carbon sources also influence dctA expression. We demonstrate that the cAMP receptor protein (CRP) has a repressive effect on the dctA promoter. A mutated CRP molecule (CRP-H159L), unable to activate catabolic promoters (but still proficient in DNA binding), gives similar results. This suggests that the CRP-cAMP complex represses the dctA promoter activity by direct interaction with the DNA. Direct binding of the CRP-cAMP complex to the dctA promoter was confirmed in vitro by gel mobility-shift assays. Sequence analysis of the dctA promoter indicates that the most likely binding sites for CRP are the two confirmed DctD-binding sites. It is proposed that the CRP-cAMP complex competes with DctD for occupancy of these sites. Since in the presence of CRP-cAMP complex the uninduced levels of dctA expression are reduced, whereas induced levels are largely unaffected, such competition appears to be an essential regulatory feature of dctA expression.
苜蓿中华根瘤菌C4-二羧酸通透酶基因(dctA)的表达受传感蛋白DctB和转录调节因子DctD的控制。苜蓿中华根瘤菌的Dct系统已在大肠杆菌中重建。dctA启动子的表达依赖于DctBD,并在C4-二羧酸(dCA)存在时被诱导。其他碳源也会影响dctA的表达。我们证明,环腺苷酸受体蛋白(CRP)对dctA启动子有抑制作用。一种无法激活分解代谢启动子(但仍能有效结合DNA)的突变CRP分子(CRP-H159L)也给出了类似结果。这表明CRP-cAMP复合物通过与DNA直接相互作用来抑制dctA启动子活性。通过凝胶迁移率变动分析在体外证实了CRP-cAMP复合物与dctA启动子的直接结合。dctA启动子的序列分析表明,CRP最可能的结合位点是两个已确认的DctD结合位点。有人提出,CRP-cAMP复合物与DctD竞争占据这些位点。由于在存在CRP-cAMP复合物的情况下,dctA未诱导表达水平降低,而诱导表达水平基本不受影响,这种竞争似乎是dctA表达的一个重要调节特征。