A Proximal Promoter Element Required for Positive Transcriptional Control by Guanosine Tetraphosphate and DksA Protein during the Stringent Response
A Proximal Promoter Element Required for Positive Transcriptional Control by Guanosine Tetraphosphate and DksA Protein during the Stringent Response
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DOI：
10.1074/jbc.m113.479998
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发表时间：
2013-07-19
影响因子：
4.8
通讯作者：
Nystrom, Thomas
中科院分区：
文献类型：
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作者：
Gummesson, Bertil;Lovmar, Martin;Nystrom, Thomas
在Dks A存在的情况下，丙氨酸鸟苷四磷酸(PpGpp)既是基因表达的正性调节因子，也是负性调节因子，但这种差异调控的潜在机制尚不清楚。在这里，使用USPA混合启动子，我们证明了富含AT的鉴别子区域对于ppGpp/DksA的正调控至关重要。富含AT的鉴别器使RNA聚合酶-启动子复合体非常稳定，因此很容易与RNA聚合酶饱和。当RNA聚合酶-启动子复合体被ppGpp/DksA破坏稳定时，可以实现更有效的转录。我们发现，将富含AT的USPA鉴别子替换为富含GC的rrnB-P1鉴别子，在体内和体外都能使USPA启动子受到ppGpp/DksA的负调控。此外，富含GC的鉴别器破坏了RNA聚合酶-启动子复合体的稳定，ppGpp/DksA对启动子动力学性质的影响被逆转。我们认为，与USPA启动子相比，带有富含GC的鉴别子的启动子的转录起始率不受开放复合体稳定性的限制。鉴于ppGpp/DksA对转录权衡的直接和间接影响的模型，对这些发现进行了讨论。
The alarmone guanosine tetraphosphate (ppGpp) acts as both a positive and a negative regulator of gene expression in the presence of DksA, but the underlying mechanisms of this differential control are unclear. Here, using uspA hybrid promoters, we show that an AT-rich discriminator region is crucial for positive control by ppGpp/DksA. The AT-rich discriminator makes the RNA polymerase-promoter complex extremely stable and therefore easily saturated with RNA polymerase. A more efficient transcription is achieved when the RNA polymerase-promoter complex is destabilized with ppGpp/DksA. We found that exchanging the AT-rich discriminator of uspA with the GC-rich rrnB-P1 discriminator made the uspA promoter negatively regulated by ppGpp/DksA both in vivo and in vitro. In addition, the GC-rich discriminator destabilized the RNA polymerase-promoter complex, and the effect of ppGpp/DksA on the kinetic properties of the promoter was reversed. We propose that the transcription initiation rate from promoters with GC-rich discriminators, in contrast to the uspA-promoter, is not limited by the stability of the open complex. The findings are discussed in view of models for both direct and indirect effects of ppGpp/DksA on transcriptional trade-offs.