Chen Shuo, Zhang Aixia, Blyn Lawrence B, Storz Gisela
IBIS Therapeutics, ISIS Pharmaceuticals, Inc., Carlsbad, California, USA.
J Bacteriol. 2004 Oct;186(20):6689-97. doi: 10.1128/JB.186.20.6689-6697.2004.
In a previous bioinformatics-based search for novel small-RNA genes encoded by the Escherichia coli genome, we identified a region, IS063, located between the ompN and ydbK genes, that encodes an approximately 100-nucleotide small-RNA transcript. Here we show that the expression of this small RNA is increased at a low temperature and in minimal medium. Twenty-two nucleotides at the 5' end of this transcript have the potential to form base pairs with the leader sequence of the mRNA encoding the outer membrane protein OmpC. The deletion of IS063 increased the expression of an ompC-luc translational fusion 1.5- to 2-fold, and a 10-fold overexpression of the small RNA led to a 2- to 3-fold repression of the fusion. Deletion and overexpression of the IS063 RNA also resulted in increases and decreases, respectively, in OmpC protein levels. Taken together, these results suggest that IS063 is a regulator of OmpC expression; thus, the small RNA has been renamed MicC. The antisense regulation was further demonstrated by the finding that micC mutations were suppressed by compensatory mutations in the ompC mRNA. MicC was also shown to inhibit ribosome binding to the ompC mRNA leader in vitro and to require the Hfq RNA chaperone for its function. We suggest that the MicF and MicC RNAs act in conjunction with the EnvZ-OmpR two-component system to control the OmpF/OmpC protein ratio in response to a variety of environmental stimuli.
在之前基于生物信息学的对大肠杆菌基因组编码的新型小RNA基因的搜索中，我们鉴定出一个位于ompN和ydbK基因之间的区域IS063，它编码一个约100个核苷酸的小RNA转录本。在此我们表明，这种小RNA的表达在低温和基本培养基中会增加。该转录本5'端的22个核苷酸有可能与编码外膜蛋白OmpC的mRNA的前导序列形成碱基对。IS063的缺失使ompC - luc翻译融合体的表达增加了1.5至2倍，而小RNA的10倍过表达导致该融合体受到2至3倍的抑制。IS063 RNA的缺失和过表达还分别导致OmpC蛋白水平的增加和降低。综合这些结果表明，IS063是OmpC表达的调节因子；因此，这种小RNA已被重新命名为MicC。ompC mRNA中的补偿性突变抑制了micC突变，这一发现进一步证明了反义调节作用。MicC在体外也被证明能抑制核糖体与ompC mRNA前导序列的结合，并且其功能需要Hfq RNA伴侣蛋白。我们认为，MicF和MicC RNA与EnvZ - OmpR双组分系统协同作用，以响应多种环境刺激来控制OmpF/OmpC蛋白比例。
