Ouimet M C, Marczynski G T
Department of Microbiology and Immunology, McGill University, Montreal, Quebec, H3A 2B4, Canada.
J Mol Biol. 2000 Sep 29;302(4):761-75. doi: 10.1006/jmbi.2000.4500.
Caulobacter crescentus CtrA protein, an OmpR-type response regulator, receives cell-cycle signals and binds the proposed consensus TTAA-N7-TTAA present inside the chromosome replication origin and cell-cycle transcription promoters. We synthesized a 42 bp cell-cycle promoter based on this consensus and elements of the fliL promoter. Over 100 promoter mutations were assayed for transcription directed by CtrA. Although both CtrA binding half-sites cooperate and the N7 spacing is critical for transcription, the upstream half-site is relatively flexible. The downstream CtrA half-site is less flexible and more important for cell-cycle regulation. A CtrA binding site and a -10 promoter element are sufficient for cell-cycle transcription, and both sequences cooperate and compensate for respective defects. Mutations in the CtrA binding site, but not in the -10 promoter sequence, perturb cell-cycle transcription. A single base-pair change switches a cell-cycle promoter into a strong conventional promoter. We propose rules for CtrA binding and promoter interactions implying how CtrA evolved into a versatile regulator of cell-cycle functions including flagellar biogenesis, cell division, DNA methylation, and chromosome replication.
新月柄杆菌CtrA蛋白是一种OmpR型应答调节因子，它接收细胞周期信号，并结合位于染色体复制起点和细胞周期转录启动子内的假定共有序列TTAA-N7-TTAA。我们基于此共有序列和fliL启动子的元件合成了一个42 bp的细胞周期启动子。对超过100个启动子突变体进行了CtrA指导的转录检测。虽然CtrA结合的两个半位点协同作用，且N7间距对转录至关重要，但上游半位点相对灵活。下游的CtrA半位点灵活性较低，对细胞周期调控更为重要。一个CtrA结合位点和一个-10启动子元件足以进行细胞周期转录，且这两个序列协同作用并弥补各自的缺陷。CtrA结合位点的突变而非-10启动子序列的突变会扰乱细胞周期转录。单个碱基对的改变会将细胞周期启动子转变为一个强的常规启动子。我们提出了CtrA结合和启动子相互作用的规则，这暗示了CtrA是如何演变成一个多功能的细胞周期功能调节因子的，这些功能包括鞭毛生物合成、细胞分裂、DNA甲基化和染色体复制。
