Barker Clive S, Prüss Birgit M, Matsumura Philip
Department of Microbiology and Immunology, College of Medicine, University of Illinois at Chicago, Chicago, Illinois, USA.
J Bacteriol. 2004 Nov;186(22):7529-37. doi: 10.1128/JB.186.22.7529-7537.2004.
The flhD operon is the master operon of the flagellar regulon and a global regulator of metabolism. The genome sequence of the Escherichia coli K-12 strain MG1655 contained an IS1 insertion sequence element in the regulatory region of the flhD promoter. Another stock of MG1655 was obtained from the E. coli Genetic Stock Center. This stock contained isolates which were poorly motile and had no IS1 element upstream of the flhD promoter. From these isolates, motile subpopulations were identified after extended incubation in motility agar. Purified motile derivatives contained an IS5 element insertion upstream of the flhD promoter, and swarm rates were sevenfold higher than that of the original isolate. For a motile derivative, levels of flhD transcript had increased 2.7-fold, leading to a 32-fold increase in fliA transcript and a 65-fold increase in flhB::luxCDABE expression from a promoter probe vector. A collection of commonly used lab strains was screened for IS element insertion and motility. Five strains (RP437, YK410, MC1000, W3110, and W2637) contained IS5 elements upstream of the flhD promoter at either of two locations. This correlated with high swarm rates. Four other strains (W1485, FB8, MM294, and RB791) did not contain IS elements in the flhD regulatory region and were poorly motile. Primer extension determined that the transcriptional start site of flhD was unaltered by the IS element insertions. We suggest that IS element insertion may activate transcription of the flhD operon by reducing transcriptional repression.
flhD操纵子是鞭毛调节子的主操纵子和新陈代谢的全局调节因子。大肠杆菌K-12菌株MG1655的基因组序列在flhD启动子的调节区域含有一个IS1插入序列元件。从大肠杆菌遗传菌种保藏中心获得了另一批MG1655菌株。这批菌株包含运动能力较差的分离株，且在flhD启动子上游没有IS1元件。在这些分离株中，在运动性琼脂中长时间培养后鉴定出了运动亚群。纯化的运动衍生物在flhD启动子上游含有一个IS5元件插入，群体游动速率比原始分离株高7倍。对于一个运动衍生物，flhD转录本水平增加了2.7倍，导致fliA转录本增加32倍，并且来自启动子探针载体的flhB::luxCDABE表达增加65倍。对一批常用实验室菌株进行了IS元件插入和运动性筛选。五株菌株（RP437、YK410、MC1000、W3110和W2637）在flhD启动子上游的两个位置之一含有IS5元件。这与高群体游动速率相关。另外四株菌株（W1485、FB8、MM294和RB791）在flhD调节区域不含有IS元件，运动能力较差。引物延伸实验确定flhD的转录起始位点未因IS元件插入而改变。我们认为IS元件插入可能通过减少转录抑制来激活flhD操纵子的转录。
