Regulation of Eukaryotic Protein Synthesis Initiation

描述（由申请人提供）：该项目的长期目标是 了解真核生物 mRNA 募集过程中发生的步骤 核糖体及其调控方式。这个过程决定了 蛋白质合成的总体速率以及翻译的 mRNA 谱。 四个具体目标主要涉及两个相互作用的启动因素 这个过程的核心是：eIF4E，一种结合 含有 7-甲基鸟苷的 mRNA 帽和连接蛋白 eIF4G 所有已知参与 mRNA 募集的起始因子一起。的 线虫秀丽隐杆线虫为研究这种线虫提供了无与伦比的优势 复杂的生化过程作为蛋白质合成的起始。令人惊讶的是， 它表达五种 eIF4E 亚型，称为 IFE 蛋白，它们具有不同的功能 帽结合特性。在具体目标 1 中，我们将尝试推断他们的 通过研究它们与 m7G- 和 m3 的生化相互作用来发挥生物学作用 含有 '2'7G 的帽类似物和寡核苷酸，与 mRNA 以及其他 蛋白质，在后一种情况下利用 MALDI-TOF 质谱法。具体来说 目标2，我们将确定人eIF4E磷酸化对蛋白质的影响 合成率。 eIF4G 与 eIF4E 和聚腺苷酸结合蛋白（即 两个 C 作用效应子用于 mRNA 募集，但 eIF4G 和 eIF4G 的许多亚型 在哺乳动物细胞中观察到 eIF4G mRNA。在具体目标 3 中，我们将 确定这些 eIF4G 亚型的结构、功能和起源， 描述已知起始因子与它们的结合。特定目标 4、我们将确定eIF4G亚型在体内的功能，特别是 强调 eIF4G 裂解在小核糖核酸病毒诱导的中的相对重要性 宿主蛋白质合成的关闭。因为 eIF4E 和 eIF4G 会导致细胞恶性转化，并且因为自然发生 肿瘤中 eIF4E 水平大幅升高，这些研究具有相关性 到癌症。它们还与小核糖核酸病毒引起的疾病相关，例如 如脊髓灰质炎病毒（脊髓灰质炎）、鼻病毒（普通感冒）和柯萨奇病毒（心脏病） 失败）。

DESCRIPTION (provided by applicant): The long-term goals of this project are to understand the steps that occur during the recruitment of eukaryotic mRNA to the ribosome and how they are regulated. This process determines both the overall rate of protein synthesis and also the spectrum of mRNAs translated. The four Specific Aims principally concern two interacting initiation factors that are central to this process: eIF4E, the protein that binds the 7-methylguanosine-containing cap of mRNA, and eIF4G, the protein that links together all initiation factors known to be involved in mRNA recruitment. The nematode C. elegans provides unparalleled advantages for studying such a complex biochemical process as initiation of protein synthesis. Surprisingly, it expresses five eIF4E isoforms, termed IFE proteins, which have differing cap-binding properties. In Specific Aim 1, we wi2ll attempt to infer their biological roles by studying their biochemical interactions with m7G- and m3 '2'7G-containing cap analogues and oligonucleotides, with mRNAs, and with other proteins, utilizing MALDI-TOF mass spectrometry in the latter case. In Specific Aim 2, we will determine the effect of human eIF4E phosphorylation on protein synthesis rate. eIF4G binds to both eIF4E and the poly(A)-binding protein, the two C is-acting effectors for mRNA recruitment, yet many isoforms of eIF4G and eIF4G mRNA are observed in mammalian cells. In Specific Aim 3, we will determine the structure, function, and origin of these eIF4G isoforms and characterize the binding of known initiation factors to them. In Specific Aim 4, we will determine the function of eIF4G isoforms in vivo, with special emphasis on the relative importance of eIF4G cleavage in picomavirus-induced host shut-off of protein synthesis. Because overexpression of both eIF4E and eIF4G causes malignant transformation of cells, and because naturally occurring tumors contain greatly elevated levels of eIF4E, these studies have relevance to cancer. They also have relevance for diseases caused by picornaviruses such as poliovirus (polio), rhinovirus (the common cold) and Coxsackievirus (heart failure).