ANALYSIS OF THE HEAT SHOCK RESPONSE IN E. COLI

该提案的目标是了解热休克的生理学 (hs) 大肠杆菌中的反应。 我们将研究其机制 反应的诱导和回归以及确切的作用 hs 蛋白在正常细菌生长和保护细菌中的作用 细胞在高温下。 由于巨大的保护 整个生命世界的 hs 反应，预计许多 拟议的大肠杆菌研究得出的结论将有助于 加深我们对法规及其重要性的理解 所有生物体都有 hs 反应。 我们将讨论几个具体的研究领域。 （一）我们将 通过克隆表征编码未鉴定的 hs 蛋白的基因， 绘制并分离其中的突变。 这些基因在 E 中的作用。 首先通过分析大肠杆菌的表型来研究大肠杆菌的生理学 正常和 hs 条件下的突变体。 基因产物将是 纯化并进行生化分析。 (b) 我们会研究规管 通过研究分子水平的 hs 反应 htpR、dnaK 和其他 hs 蛋白参与调节 hs 反应。 我们 将通过分离许多基因改变的突变体来做到这一点 兴趣，检查其各种表型，并分离 原始突变的基因间抑制因子。 我们的生化方法 将纯化野生型和突变蛋白并研究 hs 使用标准转录技术进行体外基因表达或 转录和翻译耦合。 (c) λ噬菌体感染 导致在没有a的情况下诱导大肠杆菌的hs反应 温度变化。 我们将确定负责的噬菌体基因 通过分析突变体、纯化其产物、分析 上述体外系统中蛋白质的作用。 这个 系统可以深入了解动物病毒、 如腺病毒、多瘤病毒和SV40，诱导某些宿主的合成 感染期间的 hs 蛋白。

The goal of this proposal is to understand the physiology of the heat shock (hs) response in Escherichia coli. We will investigate the mechanism of induction and regression of the response and in addition, the exact roles of the hs proteins both in normal bacterial growth and in protection of the cells at high temperature. Because of the tremendous conservation of the hs response throughout the living world, it is expected that many of the conclusions reached by the proposed studies on E. coli will contribute to increasing our understanding of the regulation and the significance of the hs response in all organisms. We will address several specific areas of research. (a) We will characterize the genes coding for the unidentified hs proteins by cloning, mapping and isolating mutations in them. The roles of these genes in E. coli physiology will be studied first by analyzing the phenotypes of mutants in both normal and hs conditions. The gene products will be purified and analyzed biochemically. (b) We will study the regulation of the hs response at the molecular level by investigating the roles of the htpR, dnaK, and other hs proteins in the modulation of the hs response. We will do this by the isolation of many mutants altered in the genes of interest, examination of their various phenotypes, and isolation of intergenic suppressors of the original mutations. Our biochemical approach will be to purify the wild-type and mutant proteins and also to study hs gene expression in vitro using standard techniques for transcription or coupled transcription and translation. (c) Bacteriophage lambda infection results in the induction of the hs response of E. coli in the absence of a temperature shift. We will identify the bacteriophage gene(s) responsible for this effect by analysis of mutants, purify their products, analyze the effects of the proteins in the in vitro system described above. This system may provide insight into the mechanisms by which animal viruses, such as adenovirus, polyoma, and SV40, induce the synthesis of certain host hs proteins during infection.