Thermogenetic tools to monitor temperature-dependent gene expression in bacteria
Thermogenetic tools to monitor temperature-dependent gene expression in bacteria
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DOI：
10.1016/j.jbiotec.2012.01.007
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发表时间：
2012-07-31
影响因子：
4.1
通讯作者：
Narberhaus, Franz
中科院分区：
文献类型：
--
作者：
Klinkert, Birgit;Cimdins, Annika;Narberhaus, Franz
自由生活的细菌不断监测它们的环境温度。剧烈的偏离会立即引起被称为冷休克或热休克反应的保护性反应。许多哺乳动物的病原体使用温度监测系统来识别宿主成功入侵的体温，通常是37摄氏度。温度反应基因的翻译可以由RNA温度计(RNAT)调节。RNAT主要在其转录本的5‘非翻译区形成复杂的结构。大多数RNAT在低温下阻断核糖体结合部位。翻译是由于RNA结构的熔化而在温度升高时诱导的。对这种依赖温度的RNA元件的分析需要在适当的温度范围内发挥作用的足够的测试系统。在此，我们对已建立的基于经典的β-半乳糖苷酶LacZ、耐热的β-半乳糖苷酶BgaB和绿色荧光蛋白GFP的报告基因系统进行综述。我们通过测试已知的RNAT来验证这些系统，并描述了一个优化的BGaB系统的构建和应用。最后，将介绍来自大肠杆菌和沙门氏菌的两种新型RNA温度计。(C)2012爱思唯尔B.V.保留所有权利。
Free-living bacteria constantly monitor their ambient temperature. Drastic deviations elicit immediate protective responses known as cold shock or heat shock response. Many mammalian pathogens use temperature surveillance systems to recognize the successful invasion of a host by its body temperature, usually 37 degrees C. Translation of temperature-responsive genes can be modulated by RNA thermometers (RNATs). RNATs form complex structures primarily in the 5'-untranslated region of their transcripts. Most RNATs block the ribosome binding site at low temperatures. Translation is induced at increasing temperature by melting of the RNA structure. The analysis of such temperature-dependent RNA elements calls for adequate test systems that function in the appropriate temperature range. Here, we summarize previously established reporter gene systems based on the classical beta-galactosidase LacZ, the heat-stable beta-galactosidase BgaB and the green fluorescent protein GFP. We validate these systems by testing known RNATs and describe the construction and application of an optimized bgaB system. Finally, two novel RNA thermometer candidates from Escherichia coli and Salmonella will be presented. (C) 2012 Elsevier B.V. All rights reserved.