Internal Initiation in the Translation of Cellular mRNAs

细胞 mRNA 翻译的内部起始

• 批准号: 6520303
• 负责人: VINCENT P MAURO
• 金额: $ 29.63万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-05-01 至 2005-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6520303
• 关键词: cell free system; gel mobility shift assay; genetic library; genetic regulatory element; genetic translation; intracellular transport; messenger RNA; molecular cloning; nucleic acid sequence; ribosomes; stress proteins

DESCRIPTION (applicant's abstract): Some picornaviral and cellular mRNAs initiate translation in a CAP-independent manner at internal ibosome entry sites (IRESes) contained within the mRNA. While extensively studied in picornaviruses, little is known about internal initiation in cellular mRNAs. The boundaries of cellular IRESes have been difficult to define and sequence comparisons show no obvious sequence similarities among cellular IRESes or between cellular and picomaviral IRESes. The Mauro laboratory has analyzed two cellular IRESes contained within the 5' UTRs of the mRNAs that encode Gtx, a homeodomain protein and Rbm3, a cold stress induced protein that is of special interest because it appears to enhance the activity of its own IRES. These studies indicated that some cellular IRE Ses were composed of shorter cis-acting regulatory sequences, some as short as 7-nucleotides, that could function independently to internally initiate translation (IRES-module), or to enhance internal initiation (enhancer element). In addition, a diversity of IRES-modules was selected from a library containing short random nucleotide sequences using a method that was developed in this laboratory. Multiple copies of some naturally-occurring and selected IRESnodules increased internal initiation synergistically. The working hypothesis is that some cellular IRESes are composed of shorter elements that can function independently. The proposed studies will identify naturally-occurring IRES-modules and regulatory elements from cellular mRNAs, while selection studies will attempt to identify sequences with different expression properties. These sequences will be analyzed to determine if they can be categorized, to investigate the rules governing their activity, and to examine if the3 recruit the translation machinery directly by interacting with rRNA or with ribosomal proteins, or indirectly through intermediary proteins. The Rbm3 protein will be studied as a potential example of a trans-acting protein. The proposed studies should provide new insights into our understanding of internal initiation. Inasmuch as several clinically important cellular genes contain IRESes, this increased understanding may allow useful therapeutic manipulations. Moreover, the use of IRE S-modules and enhancers has already resulted in synthetic IRESes that are shorter and more efficient than the large viral IRESes currently used ir dicistronic constructs and should have broad applications in research, gene therapy, and biotechnology.

描述（申请人的摘要）：一些picornaviral和细胞mRNA 在内部ibosome条目下以帽独立的方式启动翻译 mRNA中包含的位点（IRES）。当广泛研究 Picornavirus，对细胞mRNA中的内部启动知之甚少。 细胞激素的边界很难定义和序列 比较在细胞激素之间没有明显的序列相似性或 在细胞和皮科马病毒之间。毛罗实验室已经分析了两个 蜂窝激素包含在编码GTX的mRNA的5'UTR中 同源域蛋白和RBM3，一种冷应激诱导的蛋白质，是特殊的 兴趣是因为它似乎增强了自己的IRES的活动。这些 研究表明，某些细胞IRE SES由较短 顺式作用调节序列，有些短于7-核苷酸，可以 独立发挥内部启动翻译（IRES模块）或 增强内部启动（增强子元素）。另外，多样性 从包含短随机核苷酸的库中选择IRES模型 使用该实验室开发的方法的序列。多个副本 在某些自然出现和选定的IRESNODULES中，内部有增加 启动协同。工作的假设是一些细胞围局 由可以独立起作用的较短元素组成。提议 研究将确定自然存在的IRES模型和调节元件 从细胞mRNA中，选择研究将尝试识别序列 具有不同的表达特性。这些序列将分析为 确定他们是否可以分类，以调查管理他们的规则 活动，并检查该3是否直接募集翻译机械 与rRNA或核糖体蛋白相互作用，或间接通过 中介蛋白。 RBM3蛋白将作为一个潜在的例子进行研究 跨作用蛋白。拟议的研究应提供新的见解 进入我们对内部启动的理解。在临床上有几个 重要的细胞基因包含IRES，这种增加的理解可能允许 有用的治疗操作。此外，使用IRE S模块和 增强剂已经导致了较短且更多的合成卷 比目前使用的IR双重构造的大型病毒IRES效率 并且应该在研究，基因治疗和 生物技术。