Regulation of the Translational Response to Cellular Stress

项目摘要/摘要 对基因表达的适当调节是确定和维持细胞特性和 对细胞外环境做出反应。在真核生物中，这需要一个多步骤的途径从 转录到翻译和最终的降解，以及细胞对不断变化的条件的反应通常会诱导 在这些步骤中的许多步骤中高度协调的变化。对不利压力条件的反应，如缺乏 为了确保细胞存活，营养物质或氧化应激在各种各样的生物体中往往是必要的。 鉴于其巨大的能量需求，蛋白质翻译的变化在细胞应激中起着特别关键的作用。 反应，导致数千个基因的翻译发生变化。这种反应也变得不受控制 许多疾病的病理学。值得注意的是，在压力下规避翻译规则的能力是 癌症进展的标志，促进了细胞的持续生长，并改变了应激反应和 翻译机器也会导致衰老。尽管存在这些疾病联系，但这些巨大的变化如何 翻译是否规范还不能很好地理解。 DED1是一种保守的RNA解旋酶，在翻译起始过程中起着关键作用。人类的变化 Ded1的同源基因DDX3已经在许多癌症中被发现，包括儿童脑癌 髓母细胞瘤和自然杀伤/T细胞淋巴瘤。DDX3的突变也与认知功能障碍有关 DDX3参与包括艾滋病毒在内的几种病毒的复制。这些发现 强调了解Ded1/DDX3正常功能的重要性，因为这也可以揭示 与疾病相关的功能。在稳态条件下，Ded1刺激翻译启动；然而， 最近的研究表明，Ded1在应激条件下对翻译的抑制中起着重要作用， 尤其是当细胞的中央营养感受器TOR途径失活时。这项提案探索了 DED1及其相关因子在控制对细胞应激的翻译反应中的这一功能。 具体地说，目的I将描述Ded1在TOR途径失活时的应激功能。 这一角色的机制，包括关键翻译脚手架的重塑和退化 因子eIF4G，将被定义，并对此机制的下游后果进行具体的翻译 MRNAs将被确定。应激期间Ded1和eIF4G的上游调节因子和辅助因子将 也被识别和表征。然后，AIM II将检查DED1参与应激的另一部分 应激反应、应激颗粒的形成、胞浆内信使核糖核酸及相关蛋白的积累。会的 还要测试包含Ded1/DDX3突变的细胞中的应激反应是否受到影响 髓母细胞瘤。这项工作将极大地增强我们对细胞翻译反应的理解 应激，并将为未来的研究提供信息，说明它在癌症和其他疾病中的错误调控。

PROJECT SUMMARY/ABSTRACT Proper regulation of gene expression is required for determining and maintaining cellular identity and for responding to the extracellular environment. In eukaryotes, this entails a multi-step pathway for mRNA from transcription to translation and eventual degradation, and cellular responses to changing conditions often induce highly coordinated changes in many of these steps. The response to adverse stress conditions, such as lack of nutrients or oxidative stress, is often necessary in a wide diversity of organisms in order to ensure cell survival. Given its large energy requirements, changes in protein translation play a particularly critical role in cellular stress responses, resulting in altered translation of thousands of genes. This response also becomes misregulated in the pathologies of a number of diseases. Notably, the ability to circumvent translation regulation during stress is a hallmark of cancer progression that promotes continued cell growth, and alterations to stress responses and the translation machinery also contribute to aging. Despite these disease links, how these massive changes to translation are regulated is not well understood. Ded1 is a conserved RNA helicase that plays critical roles in translation initiation. Alterations in the human homolog of Ded1, DDX3, have been found in a number of cancers, including the pediatric brain cancer medulloblastoma and natural killer/T-cell lymphoma. Mutations of DDX3 are also linked to a cognitive developmental disorder, and DDX3 is involved in replication of several viruses, including HIV. These findings underscore the importance of understanding the normal functioning of Ded1/DDX3 since this can also shed light on its disease-associated functioning. In steady-state conditions, Ded1 stimulates translation initiation; however, recent research has revealed that Ded1 has a major role in the repression of translation during stress conditions, specifically when the TOR pathway, the central nutrient-sensor of the cell, is inactivated. This proposal explores this function of Ded1 and associated factors in controlling the translational response to cellular stress. Specifically, Aim I will characterize the stress function of Ded1 in response to TOR pathway inactivation. The mechanism of this role, which involves remodeling and degradation of the critical translation scaffolding factor eIF4G, will be defined, and the downstream consequences of this mechanism on translation of specific mRNAs will be determined. Upstream regulators and accessory factors for Ded1 and eIF4G during stress will also be identified and characterized. Aim II will then examine Ded1 involvement in another part of the stress response, the formation of stress granules, cytoplasmic accumulations of mRNA and associated proteins. It will also test whether the stress response is affected in cells containing Ded1/DDX3 mutations associated with medulloblastoma. This work will greatly enhance our understanding of the translational response to cellular stress and will inform future studies of its misregulation in cancer and other diseases.