mRNA selection by eIF4E isoforms and their sequestering factors.

这个项目探索了“未定”的干细胞如何通过改变它们在细胞内制造的蛋白质来发展成一个确定的细胞身份。在生物体的早期发育过程中，细胞必须决定它们成为执行特定功能的器官的路径。每个细胞内的信使RNA（mRNA）是蛋白质的蓝图（翻译）。这种mRNA被选择性地用于仅引入适当的专门功能。该项目将定义翻译因子用于仔细选择mRNA以在适当的时间和地点进行解码的机制，从而获得有关细胞如何决定其最终命运的有用信息。高中，本科，硕士和博士生将获得最现代的生物信息学，重组，分子和生物化学技术的实践经验和知识，使他们在追求科学事业的同时进入劳动力市场具有竞争力。蛋白质合成在早期动物发育中受到高度调节，在每种细胞/组织类型中发生不同，以产生具有适当结构和细胞活性的功能器官。许多实验室对发育过程中基因的转录调控进行了深入研究。然而，转录模式往往不匹配的空间和时间的蛋白质的要求。蛋白质的实际外观很大程度上取决于mRNA选择/翻译控制，其机制更为模糊。多年来，Keiper实验室通过独特形式的eIF 4翻译因子研究了mRNA的选择性。假设eIF 4 E和eIF 4G的同种型积极地和选择性地将休眠的mRNP募集到核糖体以进行有效的蛋白质合成。本项目重点研究了两个生殖系eIF 4 E（IFE-1和IFE-3）。elegans是一种简单的蠕虫，是生殖发育的理想遗传/转基因模型。该项目的第一个目标是使用解析的多核糖体RNA Seq，这是PI实验室开发的一种技术，用于识别依赖于单个IFE进行有效翻译的所有RNA。第二种使用CRISPR/Cas9荧光标记每个IFE以确定其体内定位，并允许通过MALDI-tof蛋白质组学表征其储存和检索复合物。其结果是：一个动态的“全基因组”蛋白质合成蓝图，促进生育，生殖，胚胎和器官发育

This project explores how "undecided" stem cells progress to a defined cellular identity by changing the proteins they make within the cell. During the early development of an organism, cells must decide their path to become an organ that performs a specialized function. The messenger RNAs (mRNAs) within each cell are blueprints for the proteins to be made (translation). Such mRNAs are used selectively to introduce only the appropriate specialized functions. This project will define the mechanisms that translation factors use to carefully select mRNAs to decode at appropriate times and places, yielding useful information about how cells determine their final fate. High school, undergraduate, Masters, and PhD students will gain hands-on experience and knowledge of the most contemporary bioinformatics, recombinant, molecular and biochemical techniques, making them competitive for entering the workforce while pursuing science careers. Protein synthesis is highly regulated in early animal development, occurring differently in each cell/tissue type to create functional organs with appropriate architecture and cellular activities. Transcriptional regulation of genes during development is well studied by many labs. However, transcriptional patterns often don't match the spatial and temporal protein requirements. The actual appearance of proteins is dictated largely by mRNA selection/translational control, whose mechanisms are more obscure. Over many years the Keiper lab has studied mRNA selectivity by unique forms of the eIF4 translation factors. The hypothesis is that isoforms of eIF4E and eIF4G positively and selectively recruit dormant mRNPs to ribosomes for efficient protein synthesis. This project focusses on two germ line eIF4E's (IFE-1 and IFE-3) in C. elegans, a simple worm that is an ideal genetic/transgenic model for reproductive development. The first goal of this project is to use resolved polysome RNA Seq, a technology developed in the PIs lab, to identify all RNAs that rely on individual IFEs for efficient translation. The second uses CRISPR/Cas9 to fluorescently tag each IFE to determine its localization in vivo, and allow characterization of its storage and retrieval complexes by MALDI-tof proteomics. The result: a blueprint of dynamic and "whole genomic" protein synthesis, which promotes fertility, reproduction, embryo and organ development