Karlinsey J E, Tanaka S, Bettenworth V, Yamaguchi S, Boos W, Aizawa S I, Hughes K T
Department of Microbiology, Box 357242, University of Washington, Seattle, WA 98195, USA.
Mol Microbiol. 2000 Sep;37(5):1220-31. doi: 10.1046/j.1365-2958.2000.02081.x.
The flhDC operon of Salmonella typhimurium is the master control operon required for the expression of the entire flagellar regulon. The flagellar master operon was placed under the tetracycline-inducible promoter PtetA using the T-POP transposon. Cells containing this construct are motile in the presence of tetracycline and non-motile without inducer present. No flagella were visible under the electron microscope when cells were grown without inducer. The class 1, class 2 and class 3 promoters of the flagellar regulon are temporally regulated. After addition of tetracycline, the class 1 flhDC operon was transcribed immediately. Transcription of flgM (which is transcribed from both class 2 and class 3 promoters) began 15 min after induction. At 20 min after induction, the class 2 fliA promoter became active and intracellular FliA protein levels increased; at 30 min after induction, the class 3 fliC promoter was activated. Induction of fliC gene expression coincides with the appearance of FlgM anti-sigma factor in the growth medium. This also coincides with the completion of hook-basal body structures. Rolling cells first appeared 35 min after induction, and excess hook protein (FlgE) was also found in the growth medium at this time. At 45 min after induction, nascent flagellar filaments became visible in electron micrographs and over 40% of the cells exhibited some swimming behaviour. Multiple flagella assemble and grow on individual cells after induction of the master operon. These results confirm that the flagellar regulatory hierarchy of S. typhimurium is temporally regulated after induction. Both FlgM secretion and class 3 gene expression occur upon completion of the hook-basal body structure.
鼠伤寒沙门氏菌的flhDC操纵子是表达整个鞭毛调节子所需的主控操纵子。使用T - POP转座子将鞭毛主控操纵子置于四环素诱导型启动子PtetA的控制之下。含有该构建体的细胞在四环素存在下具有运动性，而在没有诱导剂时则不运动。当细胞在没有诱导剂的情况下生长时，在电子显微镜下看不到鞭毛。鞭毛调节子的1类、2类和3类启动子受到时间调控。加入四环素后，1类flhDC操纵子立即转录。flgM（从2类和3类启动子转录）的转录在诱导后15分钟开始。诱导后20分钟，2类fliA启动子激活，细胞内FliA蛋白水平增加；诱导后30分钟，3类fliC启动子被激活。fliC基因表达的诱导与生长培养基中FlgM抗σ因子的出现同时发生。这也与钩-基体结构的完成同时发生。滚动细胞在诱导后35分钟首次出现，此时在生长培养基中也发现了过量的钩蛋白（FlgE）。诱导后45分钟，在电子显微照片中可见新生的鞭毛丝，超过40%的细胞表现出一些游动行为。在主控操纵子诱导后，多个鞭毛在单个细胞上组装并生长。这些结果证实，鼠伤寒沙门氏菌的鞭毛调节层次在诱导后受到时间调控。FlgM分泌和3类基因表达均在钩-基体结构完成后发生。
