Investigating the process of exiting mRNAs out of translation

研究 mRNA 退出翻译的过程

• 批准号: 7887958
• 负责人: Jeffery Coller
• 金额: $ 18.13万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-10 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7887958
• 关键词: Amino Acids; Binding; Biochemical; Biological; Boxing; Cells; Cellular Stress; Complex; Comprehension; Congenital Abnormality; Cues; Data; Defect; Dissociation; Embryo; Event; Excision; Gametogenesis; Gene Expression; Gene Expression Regulation; Generations; Genetic; Genetic Translation; Genetic screening method; Glucose; Goals; Homologous Gene; Human; Investigation; Lead; Length; Maternal Messenger RNA; Mediating; Mediator of activation protein; Messenger RNA; MicroRNAs; Molecular; Movement; Mus; Mutation; Nature; Normal Cell; Nutrient; Oocytes; Poly A; Poly(A) Tail; Poly(A)-Binding Proteins; Polyribosomes; Population; Process; Proteins; Public Health; RNA Helicase; Regulation; Reporting; Repression; Ribosomes; Role; Starvation; Sterility; Stress; Translating; Translational Regulation; Translations; base; carcinogenesis; cell growth; design; helicase; insight; mRNA Expression; messenger ribonucleoprotein; metaplastic cell transformation; research study; response

DESCRIPTION (provided by applicant): The long term goal of this proposal is to understand, in detail, the process of cessation of mRNA translation. We have uncovered compelling evidence that shutting down mRNA translation is not simply a passive and default event in the lifetime of an mRNA, but, rather, that mRNA is actively removed from the translational machinery in response to specific cues. Importantly, we have identified the ATP-dependent RNA helicase, Dhh1 p, as a factor required for the dissociation of mRNA from translation. Furthermore, data suggests that Dhhlp functions to exit an mRNA from translation following loss of the poly(A) tail from the 3' end of the mRNA. We have also demonstrated that Dhhlp is used by the cell to achieve global regulation of mRNA expression under conditions of nutrient-starvation. Dhhlp acts, therefore, both as a cis-acitng . regulator of mRNA translation under normal cell growth conditions and a global regulator of mRNA expression upon cellular stress. We propose to extend our initial identification of this unexplored step of gene regulation under three specific aims. First, we will examine the role of Dhhlp in the repression of global mRNA translation that is induced by nutrient starvation. Our preliminary observations indicate that under either glucose and amino acid starvation, mRNA 3' poly(A) tail status or function is rapidly and radically altered. We will investigate how this dramatic alteration in poly(A) tail function is achieved and if it is required for the movement of mRNA into translational quiescence by Dhhlp upon stress. Second, we will dissect the nature of the observed interaction between Dhhlp and mRNA undergoing translation. In particular, we will determine the mode by which Dhhlp interacts with translating mRNAs, and if Dhhlp specifically interacts with either ribosomes or with the mRNA itself. Lastly, we will investigate the molecular mechanisms by which Dhhlp promotes the exit of mRNA out of translation under normal and nutrient stress conditions. Preliminary evidence suggests that the substrate for Dhhlp activity is the 40S ribosomal subunit itself. The nature of this interaction will be more rigorously tested, and genetic and biochemical analyses will be preformed to identify additional substrates of Dhhlp. In aggregate, we will explore further the mechanisms by which mRNA is removed from translation dependent upon Dhhlp. Importantly, Dhhlp activity appears to be functionally conserved. Homologues of Dhhlp are critical for translational silencing of maternal mRNAs in oocytes, and mutations lead to precocious activation of translationally silenced maternal mRNA and the manifestation of embryonic defects. Consistent with these observations, mouse and human Dhhlp has been implicated in gametogenesis and carcinogenesis by impacting mRNA translation. The proposed studies, therefore, can be anticipated to provide relevant insight into public health issues such as the manifestation of birth defects, sterility, and cellular transformation.

描述（由申请人提供）：该提案的长期目标是详细了解 mRNA 翻译停止的过程。我们发现了令人信服的证据，表明关闭 mRNA 翻译不仅仅是 mRNA 生命周期中的被动和默认事件，而是 mRNA 响应特定提示而主动从翻译机制中移除。重要的是，我们已经鉴定出 ATP 依赖性 RNA 解旋酶 Dhh1 p 是 mRNA 从翻译中解离所需的因子。此外，数据表明，在 mRNA 3' 末端的 Poly(A) 尾丢失后，Dhlp 的功能是使 mRNA 退出翻译。我们还证明了细胞利用 Dhlp 在营养饥饿的条件下实现 mRNA 表达的全局调节。因此，Dhlp 既可以作为顺式酸，也可以作为顺式酸。正常细胞生长条件下 mRNA 翻译的调节因子和细胞应激时 mRNA 表达的全局调节因子。我们建议在三个具体目标下扩展我们对这一未经探索的基因调控步骤的初步识别。首先，我们将研究 Dhlp 在抑制营养饥饿引起的整体 mRNA 翻译中的作用。我们的初步观察表明，在葡萄糖和氨基酸饥饿下，mRNA 3'poly(A) 尾部状态或功能会迅速而彻底地改变。我们将研究这种 Poly(A) 尾部功能的显着改变是如何实现的，以及在应激时 Dhlp 是否需要这种改变才能使 mRNA 进入翻译静止状态。其次，我们将剖析观察到的 Dhlp 和正在进行翻译的 mRNA 之间相互作用的本质。特别是，我们将确定 Dhlp 与翻译 mRNA 相互作用的模式，以及 Dhlp 是否与核糖体或 mRNA 本身特异性相互作用。最后，我们将研究 Dhhlp 在正常和营养胁迫条件下促进 mRNA 退出翻译的分子机制。初步证据表明 Dhlp 活性的底物是 40S 核糖体亚基本身。这种相互作用的性质将受到更严格的测试，并且将进行遗传和生化分析以鉴定 Dhlp 的其他底物。总的来说，我们将进一步探索 mRNA 依赖于 Dhlp 从翻译中去除的机制。重要的是，Dhlp 活性似乎在功能上是保守的。 Dhhlp 同源物对于卵母细胞中母体 mRNA 的翻译沉默至关重要，突变会导致翻译沉默的母体 mRNA 过早激活和胚胎缺陷的表现。与这些观察结果一致，小鼠和人类 Dhlp 通过影响 mRNA 翻译而与配子发生和癌发生有关。因此，拟议的研究预计将为公共卫生问题提供相关见解，例如出生缺陷、不育和细胞转化的表现。