Department of Molecular Biology, Umeå University, Umeå SE 901 87, Sweden.
Nucleic Acids Res. 2011 Aug;39(14):5853-65. doi: 10.1093/nar/gkr167. Epub 2011 Mar 29.
The activities of promoters can be temporally and conditionally regulated by mechanisms other than classical DNA-binding repressors and activators. One example is the inherently weak σ(70)-dependent Pr promoter that ultimately controls catabolism of phenolic compounds. The activity of Pr is up-regulated through the joint action of ppGpp and DksA that enhance the performance of RNA polymerase at this promoter. Here, we report a mutagenesis analysis that revealed substantial differences between Pr and other ppGpp/DksA co-stimulated promoters. In vitro transcription and RNA polymerase binding assays show that it is the T at the -11 position of the extremely suboptimal -10 element of Pr that underlies both poor binding of σ(70)-RNAP and a slow rate of open complex formation--the process that is accelerated by ppGpp and DksA. Our findings support the idea that collaborative action of ppGpp and DksA lowers the rate-limiting transition energy required for conversion between intermediates on the road to open complex formation.
启动子的活性可以通过除经典 DNA 结合抑制剂和激活剂以外的机制进行时间和条件调节。一个例子是固有较弱的 σ(70)依赖型 Pr 启动子，它最终控制酚类化合物的分解代谢。Pr 的活性通过 ppGpp 和 DksA 的联合作用上调，该作用增强了 RNA 聚合酶在该启动子上的性能。在这里，我们报告了一项诱变分析，该分析揭示了 Pr 和其他 ppGpp/DksA 共同刺激的启动子之间存在显著差异。体外转录和 RNA 聚合酶结合实验表明，正是 Pr 极弱的-10 元件中-11 位的 T 碱基，导致 σ(70)-RNAP 的结合不良以及开放复合物形成的速度较慢，而 ppGpp 和 DksA 则加速了这一过程。我们的发现支持了这样一种观点，即 ppGpp 和 DksA 的协同作用降低了开放复合物形成过程中转换中间物所需的限速过渡能。
