Molecular mechanisms of translational regulation in aging

衰老转化调控的分子机制

• 批准号: 9564583
• 负责人: Vyacheslav M Labunskyy
• 金额: $ 63.5万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-30 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9564583
• 关键词: Address; Affect; Age; Aging; Ally; Applications Grants; Binding; Binding Proteins; Binding Sites; Biological Models; Biotin; Cells; Chimeric Proteins; Complex; Data; Data Set; Elements; Eukaryota; Fluorescence Microscopy; Gene Deletion; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Epistasis; Genetic Transcription; Goals; Human; Individual; Label; Link; Longevity; Magnetism; Mediating; Messenger RNA; Microfluidics; Modeling; Molecular; Molecular Profiling; Monitor; Mothers; Mutation; Organism; Pathway interactions; Pattern; Play; Post-Transcriptional Regulation; Process; Protein Biosynthesis; Proteins; Regulation; Regulatory Element; Reporter; Role; Saccharomycetales; Sorting - Cell Movement; System; Technology; Testing; Transcriptional Regulation; Translating; Translational Regulation; Translations; Work; Yeast Model System; Yeasts; aged; base; cell type; crosslinking and immunoprecipitation sequencing; experimental study; genetic signature; genome-wide; genome-wide analysis; insight; mutant; next generation sequencing; novel; protein function; public health relevance; response; ribosome profiling; tool; transcription factor; transcriptome sequencing

Genome-wide microarray and RNA sequencing studies have revealed changes in the expression of hundreds of genes during aging in diverse organisms. Transcriptional regulation clearly plays an important role in the control of gene expression during aging; however, translation efficiency likely plays an equally important role in determining protein abundance, but has been woefully understudied in this context. Here we propose to study translational changes that are associated with increased longevity and examine the mechanisms of post- transcriptional gene regulation in aging using yeast as a model system. We will test the hypothesis that, in re- sponse to genetic alterations that extend lifespan, mRNA-binding proteins (RBPs) coordinately regulate di- verse cytoprotective genes by affecting their translation efficiency. To identify RBPs involved in regulation of these processes, we will apply RNA-Seq and ribosome profiling combined with next-generation sequencing and characterize transcriptional and translational changes in a panel of long-lived gene deletion mutants identi- fied in genome-wide screens. We propose to integrate translational profiling data obtained for long-lived mu- tants with information about structural and sequence elements recognized by RBPs and build a regulatory in- teraction network. We also propose to carry out ribosome profiling in replicatively aged wild-type cells and long-lived mutant strains to globally identify genes whose expression is affected by translational regulation dur- ing aging. Finally, we will utilize cutting-edge microfluidic technologies to validate and extend these discover- ies at the single-cell level. Comparing translational profiles in young and replicatively aged wild-type yeast and multiple long-lived deletion mutants will reveal genetic signatures associated with increased longevity and will allow us to identify novel RBPs involved in translational regulation during aging. We will then characterize RBPs and directly identify their mRNA-binding targets using CLIP-Seq. These data will allow us to uncover specific mechanisms and identify cis-regulatory elements that are responsible for translational changes ob- served in long-lived mutants. We will also use fluorescence microscopy and microfluidic cell trapping in order to monitor how the abundance of RBPs changes with age in individual mother cells. Finally, we will test if the candidate RBPs identified from CLIP-Seq and microfluidics experiments play a causal role in mediating the lifespan extension through genetic epistasis analysis in order to determine whether candidate RBPs are nec- essary and sufficient for lifespan extension. Successful completion of this study will add valuable insight into translational regulation of aging, and may provide a better understanding of the molecular mechanisms that regulate aging in humans.

全基因组微阵列和RNA测序研究揭示了数百个基因表达的变化 在不同的生物体中衰老过程中的基因。转录调控显然在转录调控中发挥着重要作用。 在衰老过程中对基因表达的控制；然而，翻译效率可能在 确定蛋白质丰度，但可悲的是，在这方面研究不足。在这里，我们建议研究 与寿命延长相关的翻译变化，并检查后 以酵母为模型系统研究衰老过程中的转录基因调控。我们将检验这一假设，在重新- 为了应对延长寿命的遗传改变，mRNA结合蛋白(RBPs)协调调节细胞周期。 反义细胞保护基因通过影响其翻译效率。确定参与监管的限制性商业惯例 在这些过程中，我们将应用RNA-Seq和核糖体分析结合下一代测序 并表征了一组长寿基因缺失突变体的转录和翻译变化。 在全基因组筛选中被证实。我们建议将获得的长寿命小鼠的翻译剖面数据整合在一起。 与限制性商业惯例识别的结构和序列元件有关的信息的抗菌剂，并在- 互动网络。我们还建议在复制老化的野生型细胞中进行核糖体分析，并 长寿突变株在全球范围内识别其表达受翻译调控影响的基因 我老了。最后，我们将利用尖端微流控技术来验证和扩展这些发现- IIS处于单细胞级别。比较年轻和复制老化的野生型酵母和 多个长寿缺失突变体将揭示与延长寿命相关的基因签名，并将 使我们能够识别在衰老过程中参与翻译调节的新限制性商业惯例。然后我们将描述 并利用CLIP-SEQ直接鉴定它们的mRNA结合靶点。这些数据将使我们能够发现 具体的机制，并确定负责翻译变化的顺式调控元件。 在长寿的变种人中服役。我们还将依次使用荧光显微镜和微流控细胞捕获 监测个体母细胞中限制性商业惯例的丰度如何随年龄变化。最后，我们将测试是否 从CLIP-SEQ和微流体学实验中确定的候选限制性商业惯例在调节 通过遗传上位性分析延长寿命，以确定候选限制性商业惯例是否为NEC- 对于延长寿命来说，这是必要的，也是足够的。这项研究的成功完成将增加对 对衰老的翻译调节，并可能提供更好的理解 调节人类的衰老。