Yeast eLF2A: A Suppressor of Internal Initiation

酵母 eLF2A：内部启动的抑制剂

• 批准号: 6990517
• 负责人: WILLIAM C. MERRICK
• 金额: $ 24.57万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-01-01 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6990517
• 关键词: binding proteins; crosslink; fungal genetics; fungal proteins; gel mobility shift assay; gene deletion mutation; gene expression; gene induction /repression; genetic models; genetic regulation; genetic strain; genetic translation; immunoprecipitation; linkage mapping; messenger RNA; microarray technology; protein protein interaction; protein quantitation /detection; protein sequence; protein structure function; stress proteins; yeast two hybrid system; yeasts

DESCRIPTION (provided by applicant): Recent observations indicate that the expression of Ure2p occurs in two manners. The predominant form is via cap-dependent initiation of translation. However, under conditions of "stress", a shorter form of Ure2p is expressed via an IRES that is contained within the coding region of the Ure2 mRNA (corresponding to the Nterminal region of the protein). An apparently unrelated finding has been the identification of the yeast homolog to the human eIF2A gene which is non-essential for yeast growth. However, when the expression of the smaller form of the Ure2p protein (via a reporter system fused to lacZ) was monitored in the eIF2A deletion strain, expression oflacZ was increased 6 to 10-fold. In the simplest of terms, eIF2A would appear to be a suppressor of the IRES function found in the Ure2 mRNA. We propose the following 4 questions to more fully evaluate these findings: 1. What are the cis-acting sequences in Ure2p mRNA that are required to allow internal initiation? 2. What other yeast mRNAs are initiated via internal initiation? 3. How is control of internal initiation regulated, through covalent modification or protein content? 4. What portion of eIF2A is required to suppress internal initiation and with what proteins does it interact in this process? Given the general observation that in many systems, "stress" leads to cap-independent initiation (i.e. internal initiation), we feel that we have discovered an ideal model system in which to study the switch from capdependent to cap-independent translation. It is anticipated that the mechanism defined in the yeast system will have broad applicability and will likely provide insights into mammalian systems where there have been numerous reports on the utilization of cap-independent translation (heat shock, apoptosis, viral infection, etc.)

描述（由申请人提供）： 最近的观察表明 Ure2p 的表达以两种方式发生。主要形式是通过帽依赖的翻译起始。然而，在“应激”条件下，较短形式的 Ure2p 通过包含在 Ure2 mRNA 编码区（对应于蛋白质的 N 末端区域）内的 IRES 表达。一个明显不相关的发现是人类 eIF2A 基因的酵母同源物的鉴定，该基因对于酵母生长来说不是必需的。然而，当在 eIF2A 缺失菌株中监测较小形式的 Ure2p 蛋白的表达（通过与 lacZ 融合的报告系统）时，lacZ 的表达增加了 6 至 10 倍。用最简单的术语来说，eIF2A 似乎是 Ure2 mRNA 中 IRES 功能的抑制因子。我们提出以下 4 个问题来更全面地评估这些发现： 1. Ure2p mRNA 中允许内部启动所需的顺式作用序列是什么？ 2. 还有哪些酵母 mRNA 是通过内部启动来启动的？ 3. 如何通过共价修饰或蛋白质含量来调节内部起始的控制？ 4. eIF2A 的哪一部分需要抑制内部起始以及在此过程中它与哪些蛋白质相互作用？ 鉴于在许多系统中“压力”导致与帽无关的启动（即内部启动）的普遍观察，我们认为我们已经发现了一个理想的模型系统，可以在其中研究从帽依赖型到帽独立型翻译的转换。预计酵母系统中定义的机制将具有广泛的适用性，并且可能会为哺乳动物系统提供深入的了解，在哺乳动物系统中，有大量关于利用帽独立翻译（热休克、细胞凋亡、病毒感染等）的报告。