Molecular mechanisms of translational regulation in aging

全基因组微阵列和RNA测序研究揭示了数百种基因表达的变化， 基因在不同生物体衰老过程中的变化。转录调控显然在转录调控中起着重要作用。 在衰老过程中基因表达的控制;然而，翻译效率可能在衰老过程中起着同样重要的作用。 确定蛋白质丰度，但不幸的是在这方面研究不足。在这里，我们建议研究 翻译的变化，与增加寿命，并检查机制后， 衰老中的转录基因调控，使用酵母作为模型系统。我们将测试假设，在重新- 响应于延长寿命的遗传改变，mRNA结合蛋白（RBP）协调调节双 通过影响它们的翻译效率来逆转细胞保护基因。查明管制所涉及的限制性商业惯例， 在这些过程中，我们将应用RNA-Seq和核糖体分析与下一代测序相结合 并描述了一组长寿基因缺失突变体的转录和翻译变化。 在全基因组筛选中进行筛选。我们建议整合翻译谱数据获得的长寿亩- 与RBP识别的结构和序列元件的信息，并建立一个调控因子， 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， 长寿命突变株，以全面确定其表达受翻译调控影响的基因， 衰老。最后，我们将利用尖端的微流体技术来验证和扩展这些发现- 在单细胞水平上。比较年轻和复制老化的野生型酵母中的翻译谱， 多个长寿缺失突变体将揭示与长寿相关的遗传特征， 使我们能够确定新的RBPs参与衰老过程中的翻译调控。然后我们将描述 RBP和直接识别其mRNA结合的目标，使用CLIP-Seq。这些数据将帮助我们揭开 特定的机制，并确定顺式调节元件，负责翻译的变化， 在长寿的突变体中服役我们还将使用荧光显微镜和微流控细胞捕获， 监测个体母细胞中RBPs的丰度如何随年龄变化。最后，我们将测试 从CLIP-Seq和微流体实验中鉴定的候选RBP在介导细胞凋亡中起因果作用。 通过遗传上位性分析延长寿命，以确定候选RBP是否是nec- 对于延长寿命是必要的和足够的。成功完成这项研究将增加宝贵的见解， 翻译调节衰老，并可能提供更好的理解的分子机制， 调节人类的衰老

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.