Ferrario M, Ernsting B R, Borst D W, Wiese D E, Blumenthal R M, Matthews R G
Department of Biological Chemistry, University of Michigan, Ann Arbor 48109-1055.
J Bacteriol. 1995 Jan;177(1):103-13. doi: 10.1128/jb.177.1.103-113.1995.
The two major porins of Escherichia coli K-12 strains, OmpC and OmpF, are inversely regulated with respect to one another. The expression of OmpC and OmpF has been shown to be influenced by the leucine-responsive regulatory protein (Lrp): two-dimensional gel electrophoresis of proteins from strains with and strains without a functional Lrp protein revealed that OmpC expression is increased in an lrp strain, while OmpF expression is decreased. In agreement with these findings, we now present evidence that transcriptional (operon) fusions of lacZ+ to ompC and micF are negatively regulated by Lrp. Lrp binds specifically to the intergenic region between micF and ompC, as indicated by mobility shift assays and by DNase I footprinting. The expression of an ompF'-lacZ+ gene (translational) fusion is increased 3.7-fold in an lrp+ background compared with an lrp background, but expression of an ompF-lacZ+ operon fusion is not. Studies of in vivo expression of the outer membrane porins during growth on glucose minimal medium showed that the OmpF/OmpC ratio is higher in lrp+ strains than it is in isogenic lrp strains. The effect of Lrp was not seen in a strain containing a deletion of micF. Our studies suggest that the positive effect of Lrp on OmpF expression stems from a negative effect of Lrp on the expression of micF, an antisense RNA that inhibits ompF translation.
大肠杆菌K-12菌株的两种主要孔蛋白OmpC和OmpF相互呈反向调节。研究表明,OmpC和OmpF的表达受亮氨酸应答调节蛋白(Lrp)影响:对具有和不具有功能性Lrp蛋白的菌株进行蛋白质二维凝胶电泳分析发现,在lrp菌株中OmpC表达增加,而OmpF表达降低。与这些发现一致,我们现在提供证据表明,lacZ⁺与ompC和micF的转录(操纵子)融合受Lrp负调控。迁移率变动分析和DNase I足迹分析表明,Lrp特异性结合micF和ompC之间的基因间区域。与lrp背景相比,在lrp⁺背景下ompF'-lacZ⁺基因(翻译)融合的表达增加了3.7倍,但ompF-lacZ⁺操纵子融合的表达没有增加。在葡萄糖基本培养基上生长期间对外膜孔蛋白的体内表达研究表明,lrp⁺菌株中的OmpF/OmpC比率高于同基因lrp菌株。在含有micF缺失的菌株中未观察到Lrp的作用。我们的研究表明,Lrp对OmpF表达的正向作用源于Lrp对micF表达的负向作用,micF是一种抑制ompF翻译的反义RNA。