Erythroid stage-specific transcriptome expression, dynamics, and regulation

红系阶段特异性转录组表达、动态和调控

基本信息

批准号：
8335204
负责人：
JOHN G CONBOY
金额：
$ 45.11万
依托单位：
UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-20 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8335204
关键词：
Address Affect Alternative Splicing Basophilic Erythroblast Binding Biochemical Bioinformatics Biological Models Biological Process Biology Bone Marrow Cell division Cell physiology Cells Characteristics Complex Computer Analysis Cultured Cells Cytoskeleton Data Data Analyses Defect Detection Development Ensure Erythroblasts Erythrocytes Erythroid Erythroid Cells Erythropoiesis Event Evolution Exhibits Exons Fetal Liver Foundations Future Gene Expression Profile Genome Gifts Goals Health Hereditary Disease Human Human Biology Human Genetics Introns Joints Knockout Mice Knowledge Lead Length Methods Mining Modeling Mus Myoblasts Nucleotides Oligonucleotides Pathway interactions Pattern Physiological Population Post-Transcriptional Regulation Pronormoblasts Property Protein Isoforms Proteins Proteome RNA RNA Processing RNA Splicing RNA analysis Regulation Regulatory Element Research Research Personnel Shapes Sorting - Cell Movement Splicing Regulation Pathway Staging Structure System Systems Analysis Testing Tissues Transcript Transcriptional Regulation Work base chromatin modification comparative genomics erythroid differentiation expectation genetic regulatory protein human disease improved innovation insight iron metabolism mRNA Precursor novel programs response tool

项目摘要

DESCRIPTION (provided by applicant): Primary focus will be on global characterization of late erythroid transcriptome(s) with emphasis on detection of new transcripts and new isoforms of known transcripts, dynamic evolution of the transcriptome during late erythropoiesis, and mechanisms by which an evolutionarily conserved alternative splicing program shapes the transcriptome. Differentiating erythroblasts execute a diverse and dynamic pre-mRNA alternative splicing program that cooperates with the transcriptional program to ensure synthesis of the appropriate stage-specific proteome as cells acquire specialized functional properties. Proper regulation of alternative splicing is extremely relevant to human health, for misregulation is a major contributor to many human diseases yet the erythroid splicing program, its regulation, and its importance in erythroid biology remain poorly understood. This project proposes a global analysis of the stage-specific erythroid transcriptome by RNA-Seq analysis and advanced bioinformatic strategies to address these issues and to generate a wealth of new information of use to many other investigators studying erythroid differentiation and erythroid biology. To explore the hypothesis that a conserved mammalian erythroid alternative splicing program regulates critical erythroid functions, investigators with expertise in erythroid differentiation, alternative splicing regulation, and computational analysis of deep sequencing data have come together to propose three specific research aims. Aim 1 will define the mouse erythroid stage-specific transcriptome using highly purified FACS-sorted erythroid cells from bone marrow (proerythroblasts as well as basophilic, polychromatic, and orthochromatic erythroblast stages). Advanced computational analysis of RNA-seq data enables comparison among the differentiation stages and between erythroid and non-erythroid cells, to characterize erythroid isoform diversity and stage-specific switches in alternative splicing that imply functional changes in the encoded proteome. Aim 2 will perform a similar analysis of human erythroblasts that are highly purified by FACS sorting. Comparison of human and mouse data will facilitate the definition of evolutionarily conserved erythroid-specific and dynamic switches in isoform expression that suggests critical erythroid functions, in addition to highlighting isoform differences that exist between mouse and human cells. Aim 3 proposes a mechanistic study of the conserved alternative splicing events defined in Aims 1 and 2 by using computational and biochemical approaches to analyze cis- regulatory sequences and splicing factor proteins that direct these splicing networks. Ultimately, this work should reveal the splicing regulatory network(s) that orchestrate programmed splicing in differentiating erythroid cells. Long term benefits anticipated from this work include greatly improved insights into regulation of biological processes in erythroid cells by alternative protein isoforms. Moreover, the RNA-Seq data itself may stimulate studies of the transcriptional and post-transcriptional regulation of this transcriptome.

描述（由申请人提供）：主要关注晚期红细胞转录组的整体特征，重点是检测新转录本和已知转录本的新亚型、晚期红细胞生成过程中转录组的动态进化，以及进化过程中的机制。保守的选择性剪接程序塑造转录组。分化的成红细胞执行多样化且动态的前 mRNA 选择性剪接程序，该程序与转录程序配合，以确保在细胞获得特殊功能特性时合成适当的阶段特异性蛋白质组。选择性剪接的正确调节与人类健康极为相关，因为错误调节是许多人类疾病的主要原因，但红细胞剪接程序、其调节及其在红细胞生物学中的重要性仍然知之甚少。该项目提出通过 RNA-Seq 分析和先进的生物信息学策略对阶段特异性红细胞转录组进行全局分析，以解决这些问题，并为研究红细胞分化和红细胞生物学的许多其他研究人员提供大量新信息。为了探索保守的哺乳动物红细胞选择性剪接程序调节关键红细胞功能的假设，在红细胞分化、选择性剪接调节和深度测序数据计算分析方面具有专业知识的研究人员聚集在一起，提出了三个具体的研究目标。目标 1 将使用来自骨髓的高度纯化的 FACS 分选的红细胞（原红细胞以及嗜碱性、多染性和正染红细胞阶段）来定义小鼠红细胞阶段特异性转录组。 RNA-seq 数据的高级计算分析能够在分化阶段之间以及红系细胞和非红系细胞之间进行比较，以表征红系亚型多样性和选择性剪接中的阶段特异性开关，这意味着编码蛋白质组中的功能变化。目标 2 将对通过 FACS 分选高度纯化的人类成红细胞进行类似的分析。人类和小鼠数据的比较将有助于定义进化上保守的红细胞特异性和亚型表达的动态开关，这表明了关键的红细胞功能，此外还强调了小鼠和人类细胞之间存在的亚型差异。目标 3 提出通过使用计算和生化方法来分析指导这些剪接网络的顺式调控序列和剪接因子蛋白，对目标 1 和 2 中定义的保守选择性剪接事件进行机制研究。最终，这项工作应该揭示在分化红系细胞中协调程序性剪接的剪接调节网络。这项工作预期的长期好处包括大大提高对替代蛋白质亚型对红系细胞生物过程调节的认识。此外，RNA-Seq数据本身可能会刺激对该转录组的转录和转录后调控的研究。