Mechanism of action of codon pair bias

密码子对偏倚的作用机制

• 批准号: 8460843
• 负责人: BRUCE Bruce FUTCHER
• 金额: $ 25.56万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2015-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8460843
• 关键词: Affect; Amino Acids; Attenuated; Attenuated Live Virus Vaccine; Attenuated Vaccines; Biological Models; Code; Codon Nucleotides; Data; Development; Escherichia coli; Flushield; Gene Expression; Genes; Goals; Human; Human poliovirus; Knowledge; Life; Maps; Measures; Messenger RNA; Methods; Modeling; Molecular; Movement; Mutate; Phenotype; Polioviruses; Polyribosomes; Procedures; Proteins; Ribosomes; Speed; System; Testing; Time; Translating; Translations; Vaccines; Viral Vaccines; Virus; Work; Yeasts; attenuation; base; density; design; fitness; gene function; influenza virus vaccine; influenzavirus; multicatalytic endopeptidase complex; mutant; protein degradation; protein misfolding; research study

DESCRIPTION (provided by applicant): Because an amino acid can be encoded by as many as six codons, there are many different ways to encode any particular protein. Biases exist in the way codons are used. One well- known bias is the codon bias, which is simply that some codons are used more than others. Less well-known is that there is also a "codon pair bias", such that some codons "prefer" to be adjacent to certain other codons. This codon pair bias is completely separate and independent from the codon bias. Recently, we have found that when viruses are re-coded to have a bad codon pair bias, the viruses are attenuated, in extreme cases to inviability. It appears that attenuation of a virus via a bad codon pair bias can be used to make a live, attenuated vaccine. However, while the procedure works, nothing whatever is known about the mechanim by which codon pair bias causes attenuation, and this lack of knowledge is slowing the work with viral vaccines. In this proposal, we will investigate, for the first time, the mechanism of attenuation by bad codon pair bias. This will be done in yeast, where we have recently shown there are strong codon pair bias effects. We will confirm the effects of codon pair bias in yeast using two codon pair de-optimized synthetic yeast genes, dHIS3 and dLYS2. We will test the idea that bad codon pair bias slows translation using polysome profiling and ribosome density mapping. We will test the idea that bad codon pair bias causes inaccurate translation and protein degradation by turning off proteasomal degradation, and also using other methods. We have already selected yeast mutants that are apparently less sensitive than wild-type to codon pair bias, and we will characterize these mutants and identify the mutant genes. Finally we will do a high- throughput study of (initially) 22,000 different encodings of HIS3 to correlate particular encodings with the strength of gene function. Our long term goal is to understand the mechanism of attenuation by bad codon pair bias to facilitate the development of live, attenuated viral vaccines, and also to facilitate the tuning of gene expression.

描述（由申请人提供）：因为氨基酸可以由多达六个密码子编码，所以有许多不同的方法来编码任何特定的蛋白质。密码子的使用方式存在偏差。一种众所周知的偏差是密码子偏差，即某些密码子比其他密码子使用得更多。不太为人所知的是，还存在“密码子对偏差”，即某些密码子“更喜欢”与某些其他密码子相邻。这种密码子对偏倚与密码子偏倚完全分开且独立。最近，我们发现，当病毒被重新编码为具有不良密码子对偏差时，病毒就会被减毒，在极端情况下甚至会变得无法生存。看来，通过不良密码子对偏差来减毒病毒可用于制造活的减毒疫苗。然而，虽然该程序有效，但对密码子对偏倚导致减毒的机制一无所知，而这种知识的缺乏正在减慢病毒疫苗的工作。在本提案中，我们将首次研究不良密码子对偏差造成的衰减机制。这将在酵母中完成，我们最近表明酵母存在强烈的密码子对偏差效应。我们将使用两个密码子对去优化的合成酵母基因 dHIS3 和 dLYS2 来确认酵母中密码子对偏倚的影响。我们将使用多核糖体分析和核糖体密度图来测试坏密码子对偏差会减慢翻译的想法。我们将通过关闭蛋白酶体降解以及使用其他方法来测试不良密码子对偏差会导致不准确的翻译和蛋白质降解的想法。我们已经选择了明显比野生型对密码子对偏倚更不敏感的酵母突变体，我们将表征这些突变体并鉴定突变基因。最后，我们将对（最初）22,000 种不同的 HIS3 编码进行高通量研究，以将特定编码与基因功能的强度相关联。我们的长期目标是了解不良密码子对偏倚造成的减毒机制，以促进活减毒病毒疫苗的开发，并促进基因表达的调整。