Environmental and programmed regulation of start codon recognition

描述（由申请人提供）： 翻译后摘要：环境条件的变化可以影响基因表达在许多层面上。一般来说，转录的改变是环境扰动最广泛探索的结果，但特定mRNA翻译水平的变化也可能对细胞应对环境应激和毒素或其他外部因素造成的损害的能力产生深远影响。此外，决定细胞命运和功能的关键遗传程序（包括许多发育过程）的执行通常始于翻译水平。蛋白质合成起始阶段的失调在从癌症到神经退行性疾病的多种人类疾病的病因学中起作用。通过翻译机制选择mRNA中的起始密码子是基因表达的关键阶段，因为它决定了将要产生的蛋白质的N-末端和解码的阅读框架。有相当多的间接证据表明，起始密码子识别的保真度可能受到环境条件的影响-例如，暴露于应激源和毒素或特定营养素水平的变化-并且这些变化以影响基因表达的方式改变基因表达。 细胞的生理学。起始密码子识别保真度的变化可能导致产生具有不同N-末端的蛋白质同种型，所述不同N-末端由mRNA的5 '前导序列中的替代起始密码子的翻译起始产生。它还可以通过调节mRNA上游调控开放阅读框的翻译来改变蛋白质的表达水平。这些基因表达的改变可能对细胞有害，如果是非编程的，或者是有益的，如果它们是转录后基因调控机制的一部分，允许细胞对其改变的环境或内部或外部信号做出反应。使用已建立的高通量双荧光素酶测定来测量起始密码子识别的保真度，我们将：1）测试外部条件、信号和内部线索可以调节起始密码子识别的保真度以在翻译水平上改变基因表达的假设; 2）确定起始密码子识别的保真度的强制改变对细胞生理学和基因表达的影响。这些研究有可能揭示一种全新的转录后基因调控模式。它们还将提供重要的见解，以了解环境条件如何在翻译水平上影响基因表达，以及这些变化对细胞生理学的影响。

DESCRIPTION (provided by applicant): Abstract: Changes in environmental conditions can affect gene expression at many levels. In general, alterations in transcription have been the most widely explored consequences of environmental perturbations, but changes at the level of the translation of specific mRNAs can also have profound effects on a cell's ability to respond to environmental stresses and to damage caused by toxins or other external agents. In addition, execution of critical genetic programs that determine a cell's fate and function - including many developmental processes - frequently begins at the translational level. Disregulation of the initiation phase of protein synthesis plays a role in the etiology of a variety of human diseases, from cancers to neurodegenerative disorders. The selection of the start codon in an mRNA by the translational machinery is a critical phase of gene expression because it determines both the N-terminus of the protein that will be produced and the reading frame of decoding. There is considerable circumstantial evidence that the fidelity of start codon recognition can be affected by environmental conditions - for example, exposure to stressors and toxins or changes in the levels of specific nutrients - and that these changes alter gene expression in ways that affect the physiology of the cell. Changes in the fidelity of start codon recognition could result in production of protein isoforms with different N-termini created by translation initiation from alternative start codons in the 5'-leaders of mRNAs. It could also change expression levels of proteins by modulating translation of regulatory upstream open reading frames in mRNAs. These alterations in gene expression could be harmful to the cell, if unprogramed, or beneficial, if they are part of a posttranscriptional gene regulatory mechanism that allows the cell to respond to its altered environment or internal or external signals. Using an established, high-throughput dual-luciferase assay for measuring the fidelity of start codon recognition we will: 1) Test the hypothesis that external conditions, signals and internal cues can modulate the fidelity of start codon recognition to alter gene expression at the translational level; 2) Determine the effects of forcing changes in the fidelity of start codon recognition on cell physiology and gene expression. These studies have the potential to uncover a completely new mode of post-transcriptional gene regulation. They will also provide important insights into how environmental conditions affect gene expression at the translational level, as well as the consequences of these changes for cell physiology.