Transduction of envelope stress in Escherichia coli by the Cpx two-component system
Transduction of envelope stress in Escherichia coli by the Cpx two-component system
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DOI：
10.1128/jb.179.24.7724-7733.1997
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发表时间：
1997-12-01
影响因子：
3.2
通讯作者：
Silhavy, TJ
中科院分区：
文献类型：
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作者：
Raivio, TL;Silhavy, TJ
埃希氏菌细胞包膜（内膜、周质、外膜）中正常蛋白质运输的破坏可以激活两条平行但不同的信号转导途径。这种激活刺激了许多基因的表达，其产物的功能是折叠或降解错误定位的蛋白质。这些信号转导途径之一是由组氨酸激酶 CpxA 和反应调节器 CpxR 组成的双组分调节系统。在本研究中，我们对功能获得性 Cpx* 突变体进行了表征，以了解有关 Cpx 信号转导的更多信息。测序表明 cpx* 突变聚集在 CpxA 的周质、跨膜或 II-box 结构域中。有趣的是，大多数周质 cpx* 功能获得突变聚集在该结构域的中心区域，其中一个编码 32 个氨基酸的缺失。含有这些突变的菌株对正常激活信号不敏感。野生型 CpxA 和代表性 cpx* 突变体 CpxA101 之间的麦芽糖结合蛋白融合体的体内和体外表征表明，突变体 CpxA 在与 CpxR 的磷酸转移反应中发生了改变。具体而言，虽然 CpxA 和 CpxA101 均充当自激酶和 CpxR 激酶，但 CpxA101 缺乏野生型蛋白中通常存在的 CpxR-P 磷酸酶活性。总而言之，这些数据支持 Cpx 介导的信号转导模型，其中激酶/磷酸酶比率因压力而升高。此外，周质 cpx* 突变的序列和表型表明，与周质信号分子的相互作用通常可能导致非应激条件下激酶/磷酸酶比率降低。
Disruption of normal protein trafficking in the Escherichia call cell envelope (inner membrane, periplasm, outer membrane) can activate two parallel, but distinct, signal transduction pathways. This activation stimulates the expression of a number of genes whose products function to fold or degrade the mislocalized proteins. One of these signal transduction pathways is a two-component regulatory system comprised of the histidine kinase CpxA and the response regulator, CpxR. In this study we characterized gain-of-function Cpx* mutants in order to learn more about Cpx signal transduction. Sequencing demonstrated that the cpx* mutations cluster in either the periplasmic, the transmembrane, or the II-box domain of CpxA. Intriguingly, most of the periplasmic cpx* gain-of-function mutations cluster in the central region of this domain, and one encodes a deletion of 32 amino acids. Strains harboring these mutations are rendered insensitive to a normally activating signal. In vivo and in vitro characterization of maltose-binding-protein fusions between the wild-type CpxA and a representative cpx* mutant, CpxA101, showed that the mutant CpxA is altered in phosphotransfer reactions with CpxR. Specifically, while both CpxA and CpxA101 function as autokinases and CpxR kinases, CpxA101 is devoid of a CpxR-P phosphatase activity normally present in the wild-type protein. Taken together, the data support a model for Cpx-mediated signal transduction in which the kinase/phosphatase ratio is elevated by stress. Further, the sequence and phenotypes of periplasmic cpx* mutations suggest that interactions with a periplasmic signaling molecule may normally dictate a decreased kinase/phosphatase ratio under nonstress conditions.