Initiation of mRNA decay in Bacillus subtilis

枯草芽孢杆菌中 mRNA 降解的启动

• 批准号: 6470300
• 负责人: DAVID H BECHHOFER
• 金额: $ 32.04万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-01-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6470300
• 关键词: Bacillus subtilis; bacterial genetics; chemical cleavage; enzyme activity; exoribonucleases; gene expression; gene mutation; genetic mapping; genetic translation; messenger RNA; microarray technology; microorganism culture; ribonuclease III

DESCRIPTION: (provided by applicant): Our laboratory seeks to understand the control of mRNA decay in Bacillus subtilis. For the Gram-positive bacteria, much needs to be learned about the features of an mRNA that determine its half-life, the ribonucleolytic reactions involved in mRNA decay, the genes that encode ribonuc the regulation of their expression. Since the B. subtilis genome does not have sequence homologues for several of the major ribonuclease genes of Escherichia coli, it is altogether uncertain whether models for mRNA decay based on the well-studied E. coli system will pertain to B. subtilis. Small RNA molecules will be used to probe three facets of mRNA turnover in B. subtilis: 1) the entry site for 3' exonucleolytic degradation; 2) endonucleolytic cleavage that initiates decay; and 3) role of the 5' end in determining mRNA half-life. These small RNA molecules are designed such that analysis of their decay will begin to clarify how mRNA turnover is achieved. To identify additional ribonuclease genes (three have been cloned thus far), biochemical experiments are proposed to isolate several proteins predicted to be involved in mRNA decay, i.e., at least one additional 3'-to-5' exoribonuclease, a putative endoribonuclease, and poly(A) polymerase. Once the identities of these proteins are known, the genes encoding them will be disrupted in order to study the effects on mRNA decay. To develop our understanding of ribonuclease function in the Gram-positive bacteria, we will study the function of Bs-RNase III, a narrow-specificity endoribonuc lease that has been shown to be essential in B. subtilis. Genetic means will be employed in an effort to understand the role of Bs-RNase Ill that is critical for viability. The basis for control of Bs-R.Nase III activity in the cell will be investigated, providing the first look at ribonuclease gene regulation in B. subtilis.

描述：（由申请人提供）：我们的实验室致力于了解 枯草芽孢杆菌中 mRNA 衰减的控制。对于革兰氏阳性菌， 关于 mRNA 的特征，我们需要了解很多，这些特征决定了它的 半衰期，参与 mRNA 衰变的核糖核酸分解反应， 编码核糖核酸并调节其表达。由于枯草芽孢杆菌基因组 几个主要核糖核酸酶基因没有序列同源物 大肠杆菌的 mRNA 衰减模型是否完全不确定 基于经过充分研究的大肠杆菌系统，该系统将属于枯草芽孢杆菌。 小 RNA 分子将用于探测 B. 枯草芽孢杆菌：1) 3' 核酸外切降解的进入位点； 2） 引发衰变的核酸内切裂解； 3) 5'端的作用 确定 mRNA 半衰期。这些小 RNA 分子的设计使得 对它们衰变的分析将开始阐明 mRNA 更新是如何实现的。 为了鉴定其他核糖核酸酶基因（迄今为止已克隆了三个）， 建议进行生化实验来分离几种预计可 参与 mRNA 衰变，即至少有一个额外的 3'-to-5' 核糖核酸外切酶、推定的核糖核酸内切酶和聚腺苷酸聚合酶。一旦 这些蛋白质的身份是已知的，编码它们的基因将是 破坏以研究对 mRNA 衰变的影响。 加深我们对革兰氏阳性菌中核糖核酸酶功能的理解 细菌，我们将研究 Bs-RNase III 的功能，这是一种窄特异性 endoribonuc 租赁已被证明对枯草芽孢杆菌至关重要。遗传 将采用多种方法来努力理解 Bs-RNase III 的作用 对于生存能力至关重要。控制 Bs-R.Nase III 活性的基础 将研究细胞，首次观察核糖核酸酶基因 枯草芽孢杆菌中的调节。