Simpson D A, Hammarton T C, Roberts I S
Department of Microbiology and Immunology, University of Leicester, United Kingdom.
J Bacteriol. 1996 Nov;178(22):6466-74. doi: 10.1128/jb.178.22.6466-6474.1996.
The transcriptional organization and regulation of region 1 expression of the Escherichia coli K5 capsule gene cluster were studied. Region 1 was transcribed as an 8.0-kb polycistronic mRNA which was processed to form a separate 1.3-kb transcript encoding the 3'-most gene kpsS. Transcription of region 1 of the E. coli K5 capsule gene cluster was directed from a single promoter 225 bp upstream of a previously unidentified gene, kpsF. The promoter had -35 and -10 consensus sequences typical of an E. coli sigma 70 promoter, with no similarities to binding sites for other sigma factors. Two integration host factor (IHF) binding site consensus sequences were identified 110 bp upstream and 130 bp downstream of the transcription start site. In addition, two AT-rich regions separated by 16 bp identified upstream of the region 1 promoter were conserved upstream of the region 3 promoter. The kpsF gene was 98.8% identical with the kpsF gene identified in the E. coli K1 antigen gene cluster and confirms that the kpsF gene is conserved among group II capsule gene clusters. An intragenic Rho-dependent transcriptional terminator was discovered within the kpsF gene. No essential role for KpsF in the expression of the K5 antigen could be established. The temperature regulation of region 1 expression was at the level of transcription, with no transcription detectable in cells grown at 18 degrees C. Mutations in regulatory genes known to control temperature-dependent expression of a number of virulence genes had no effect on the temperature regulation of region 1 expression. Likewise, RfaH, which is known to regulate expression of E. coli group II capsules had no effect on the expression of region 1. Mutations in the himA and himD genes which encode the subunits of the IHF led to a fivefold reduction in the expression of KpsE at 37 degrees C, confirming a regulatory role for IHF in the expression of region 1 genes.
对大肠杆菌K5荚膜基因簇区域1表达的转录组织和调控进行了研究。区域1转录为一个8.0 kb的多顺反子mRNA，该mRNA经过加工形成一个单独的1.3 kb转录本，编码最下游的基因kpsS。大肠杆菌K5荚膜基因簇区域1的转录由一个位于先前未鉴定基因kpsF上游225 bp处的单一启动子指导。该启动子具有大肠杆菌σ70启动子典型的-35和-10共有序列，与其他σ因子的结合位点没有相似性。在转录起始位点上游110 bp和下游130 bp处鉴定出两个整合宿主因子（IHF）结合位点共有序列。此外，在区域1启动子上游鉴定出的两个相隔16 bp的富含AT的区域在区域3启动子上游保守。kpsF基因与在大肠杆菌K1抗原基因簇中鉴定出的kpsF基因有98.8%的同一性，证实kpsF基因在II组荚膜基因簇中是保守的。在kpsF基因内发现了一个基因内的依赖Rho的转录终止子。未发现KpsF在K5抗原表达中起关键作用。区域1表达的温度调控发生在转录水平，在18℃生长的细胞中未检测到转录。已知控制许多毒力基因温度依赖性表达的调控基因突变对区域1表达的温度调控没有影响。同样，已知调控大肠杆菌II组荚膜表达的RfaH对区域1的表达也没有影响。编码IHF亚基的himA和himD基因突变导致37℃时KpsE表达降低五倍，证实了IHF在区域1基因表达中的调控作用。
